Elephant
Bibliographic
Database

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Elephant Bibliographic Database
www.elephantcare.org

References updated October 2009 by date of publication, most recent first.

Behr, B., Rath, D., Hildebrandt, T.B., Goeritz, F., Blottner, S., Portas, T.J., Bryant, B.R., Sieg, B., Knieriem, A., de Graaf, S.P., Maxwell, W.M., Hermes, R., 2009. Germany/Australia index of sperm sex sortability in elephants and rhinoceros. Reprod. Domest. Anim 44, 273-277.
Abstract: Flow cytometric sexing of spermatozoa followed by application in artificial insemination or in vitro fertilization provides a unique opportunity to predetermine the sex of offspring and might enhance the conservation management of endangered species in captivity such as the elephant and rhinoceros. To obtain an indication of the sortability of spermatozoa from these species, the relative DNA differences between X and Y chromosome bearing spermatozoa (fresh, frozen thawed, epididymal) from three rhinoceros species [white (Ceratotherium simum), black (Diceros bicornis), Indian (Rhinoceros unicornis)] and both elephant species, the Asian and the African elephant (Elephas maximus, Loxodonta Africana), were determined through separation of spermatozoa into X and Y chromosome bearing populations, using a modified high speed flow cytometer. The head profile areas of spermatozoa from all five species were measured using light microscopy. By multiplying the relative DNA differences and the head profile areas, the sperm sorting indices were calculated to be 47, 48 and 51 for white, black and Indian rhinoceros respectively. The calculated sorting index for the Asian elephant was 66. In the African elephant, we determined the highest sorting index of 76. These results indicate the practicability of flow cytometric sex sorting of spermatozoa from the tested rhinoceros species and both elephant species. The lower sorting indices in rhinos indicate that sex sorting of spermatozoa from the rhinoceros will be more challenging than in elephants

Brodie, J.F., Helmy, O.E., Brockelman, W.Y., Maron, J.L., 2009. Bushmeat poaching reduces the seed dispersal and population growth rate of a mammal-dispersed tree. Ecological Applications 19, 854-863.
Abstract: Myriad tropical vertebrates are threatened by overharvest. Whether this harvest has indirect effects on nonhunted organisms that interact with the game species is a critical question. Many tropical birds and mammals disperse seeds. Their overhunting in forests can cause zoochorous trees to suffer from reduced seed dispersal. Yet how these reductions in seed dispersal influence tree abundance and population dynamics remains unclear. Reproductive parameters in long-lived organisms often have very low elasticities; indeed the demographic importance of seed dispersal is an open question. We asked how variation in hunting pressure across four national parks with seasonal forest in northern Thailand influenced the relative abundance of gibbons, muntjac deer, and sambar deer, the sole dispersers of seeds of the canopy tree Choerospondias axillaris. We quantified how variation in disperser numbers affected C. axillaris seed dispersal and seedling abundance across the four parks. We then used these data in a structured population model based on vital rates measured in Khao Yai National Park (where poaching pressure is minimal) to explore how variation in illegal hunting pressure might influence C. axillaris population growth and persistence. Densities of the mammals varied strongly across the parks, from relatively high in Khao Yai to essentially zero in Doi Suthep-Pui. Levels of C. axillaris seed dispersal and seedling abundance positively tracked mammal density. If hunting in Khao Yai were to increase to the levels seen in the other parks, C. axillaris population growth rate would decline, but only slightly. Extinction of C. axillaris is a real possibility, but may take many decades. Recent and ongoing extirpations of vertebrates in many tropical forests could be creating an extinction debt for zoochorous trees whose vulnerability is belied by their current abundance.

Brown, J.L., Kersey, D.C., Freeman, E.W., Wagener, T., 2009. Assessment of diurnal urinary cortisol excretion in Asian and African elephants using different endocrine methods. Zoo. Biol.
Abstract: Longitudinal urine samples were collected from Asian and African elephants to assess sample processing and immunoassay techniques for monitoring adrenal activity. Temporal profiles of urinary cortisol measured by RIA and EIA, with and without dichloromethane extraction, were similar; all correlation coefficients were >0.90. However, based on regression analyses, cortisol immunoactivity in extracted samples was only 72-81% of that of unextracted values. Within assay technique, RIA values were only 74-81% of EIA values. Collection of 24-hr urine samples demonstrated a clear diurnal pattern of glucocorticoid excretion, with the lowest concentrations observed just before midnight and peak concentrations occurring around 0600-0800 hr. These results indicate that elephants fit the pattern of a diurnal species, and that glucocorticoid production is affected by a sleep-wake cycle similar to that described for other terrestrial mammals. Cortisol can be measured in both extracted and unextracted urine using RIA and EIA methodologies. However, unexplained differences in quantitative results suggest there may be sample matrix effects and that data generated using different techniques may not be directly comparable or interchangeable. Zoo Biol 28:1-10, 2009. (c) 2009 Wiley-Liss, Inc

Freeman, E.W., Schulte, B.A., Brown, J.L., 2009. Investigating the impact of rank and ovarian activity on the social behavior of captive female African elephants . Zoo Biology 0, 1-14.
Abstract: Over a third of captive female African elephants in North America fail to exhibit normal estrous cycles based on long-term serum progestagen analyses. Why acyclicity occurs is unknown; however, the majority of noncycling females are ranked by keepers as the dominant individual within the group. To investigate the relationship between ovarian cyclicity status and keeper-determined social rank, observations were conducted on 33 female African elephants (18 cycling, 15 noncycling). Based on keeper evaluations, five cycling elephants were ranked dominant, seven in the middle and six as subordinate. In contrast, 10 noncycling elephants were ranked as dominate and five as subordinate with none ranked as middle. When comparing the behavior of the elephants by their keeper-determined rank, the dominant females dominant were significantly more likely to approach, displace and push. Similarly, keeper-determined subordinate females more frequently presented their hind end and held their ears erect. Behaviors initiated by one elephant toward another did not vary between cycling and noncycling females, except when the interaction with social rank was tested. Dominant, noncycling females initiated a higher percentage of approach and displace behaviors than both cycling and noncycling, subordinate elephants. Subordinate, noncycling elephants displayed the highest percentage of ears erect. Social rank drives the interactions of ex situ female African elephants more than ovarian cyclicity status. Thus, behavioral interactions cannot be used to predict which cycling elephants are most likely to become acyclic.

Freeman, E.W., Schulte, B.A., Brown, J.L., 2009. Investigating the impact of rank and ovarian activity on the social behavior of captive female African elephants . Zoo Biology 0, 1-14.
Abstract:
Over a third of captive female African elephants in North America fail to exhibit normal estrous cycles based on long-term serum progestagen analyses. Why acyclicity occurs is unknown; however, the majority of noncycling females are ranked by keepers as the dominant individual within the group. To investigate the relationship between ovarian cyclicity status and keeper-determined social rank, observations were conducted on 33 female African elephants (18 cycling, 15 noncycling). Based on keeper evaluations, five cycling elephants were ranked dominant, seven in the middle and six as subordinate. In contrast, 10 noncycling elephants were ranked as dominate and five as subordinate with none ranked as middle. When comparing the behavior of the elephants by their keeper-determined rank, the dominant females dominant were significantly more likely to approach, displace and push. Similarly, keeper-determined subordinate females more frequently presented their hind end and held their ears erect. Behaviors initiated by one elephant toward another did not vary between cycling and noncycling females, except when the interaction with social rank was tested. Dominant, noncycling females initiated a higher percentage of approach and displace behaviors than both cycling and noncycling, subordinate elephants. Subordinate, noncycling elephants displayed the highest percentage of ears erect. Social rank drives the interactions of ex situ female African elephants more than ovarian cyclicity status. Thus, behavioral interactions cannot be used to predict which cycling elephants are most likely to become acyclic.

Freeman, E.W., Whyte, I., Brown, J.L., 2009. Reproductive evaluation of elephants culled in Kruger National Park, South Africa between 1975 and 1995. African Journal of Ecology 47, 192-201.
Abstract: To reduce elephant densities and preserve biological diversity, 14,629 elephants were culled from Kruger National Park, South Africa (1967-1999). Data were catalogued between 1975 and 1996 on 2737 male and female elephants, including pregnancy and lactational status for 1620 females (>= 5 years of age) and, uterine and/or ovarian characteristics for 1279. This study used these data to investigate the effects of age and precipitation on reproduction. The youngest age of conception was 8 years (n = 6) and by 12 years of age all females were sexually mature. From the age of 14 years, the percentage of reproductively active females (pregnant and/or lactating) was > 90%; however, this percentage declined when females reached 50 years of age. Overall, one-tenth of females were nonreproductive (not pregnant or lactating) at any given time, mostly in the youngest (< 15 years) and oldest (> 50 years) age classes. Eighteen (3.3%) of the nonpregnant females had reproductive tract pathologies, including endometrial, uterine or ovarian cysts. There was a seasonal distribution of mating activity that correlated with the rainy season. As has been demonstrated in other populations of free-ranging African elephants, most of the females in Kruger National Park were reproductively active; however, age and climate affected reproductive activity.

Freeman, E.W., Guagnano, G., Olson, D., Keele, M., Brown, J.L., 2009. Social factors influence ovarian acyclicity in captive African elephants (Loxodonta africana). Zoo. Biol. 28, 1-15.
Abstract: Nearly one-third of reproductive age African elephants in North America that are hormonally monitored fail to exhibit estrous cycle activity, which exacerbates the nonsustainability of the captive population. Three surveys were distributed to facilities housing female African elephants to determine how social and environmental variables contribute to cyclicity problems. Forty-six facilities returned all three surveys providing information on 90% of the SSP population and 106 elephants (64 cycling, 27 noncycling and 15 undetermined). Logistic analyses found that some physiological and social history variables were related to ovarian acyclicity. Females more likely to be acyclic had a larger body mass index and had resided longer at a facility with the same herdmates. Results suggest that controlling the weight of an elephant might be a first step to helping mitigate estrous cycle problems. Data further show that transferring females among facilities has no major impact on ovarian activity. Last, social status appears to impact cyclicity status; at 19 of 21 facilities that housed both cycling and noncycling elephants, the dominant female was acyclic. Further studies on how social and environmental dynamics affect hormone levels in free-living, cycling elephants are needed to determine whether acyclicity is strictly a captivity-related phenomenon

Freeman, E.W., Schulte, B.A., Brown, J.L., 2009. Using behavioral observations and keeper questionnaires to assess social relationships among captive female African elephants
60. Zoo. Biol. 28, 1-14.
Abstract: Free-ranging African elephants are highly social animals that live in a society where age, size, kinship, and disposition all contribute to social rank. Although captive elephant herds are small and largely comprises of unrelated females, dominance hierarchies are common. The goal of this study was to delineate how the behavior of captive female African elephants varies with respect to age and social rank based on a combination of keeper questionnaires and behavioral observations. "Body movements" and "trunk to" behaviors of 33 nonpregnant female African elephants housed at 14 North American zoos were recorded over 8 hr. Keepers at each facility also rated each elephant based on a series of questions about interactions with herdmates. The assessment of social rank based on observations correlated strongly with ranks assigned by keepers via the questionnaires. Observations and questionnaire responses indicated that body weight of the female, and to a lesser extent age, were significantly related to rates and types of "body movements" and that these demographic factors dictate the captive elephant hierarchy, similar to that observed in the wild. Many of the observed "body movements," such as back away, displace, push, and present, were correlated with keeper questionnaire responses about elephant interactions. However, none of the "trunk to" behaviors were related to age, size, or questionnaire responses even though they occurred frequently. In conclusion, we demonstrated that short-term behavioral observations and keeper questionnaires provided similar behavioral profiles for female African elephants housed in North American zoos. Zoo Biol 28:1-14, 2009. (c) 2009 Wiley-Liss, Inc

Fulka, J., Jr., Loi, P., Ptak, G., Fulka, H., John, J.S., 2009. Hope for the mammoth? Cloning Stem Cells 11, 1-4.

Hakeem, A.Y., Sherwood, C.C., Bonar, C.J., Butti, C., Hof, P.R., Allman, J.M., 2009. Von Economo neurons in the elephant brain. Anat. Rec. (Hoboken. ) 292, 242-248.
Abstract: Von Economo neurons (VENs), previously found in humans, all of the great ape species, and four cetacean species, are also present in African and Indian elephants. The VENs in the elephant are primarily found in similar locations to those in the other species. They are most abundant in the frontoinsular cortex (area FI) and are also present at lower density in the anterior cingulate cortex. Additionally, they are found in a dorsolateral prefrontal area and less abundantly in the region of the frontal pole. The VEN morphology appears to have arisen independently in hominids, cetaceans, and elephants, and may reflect a specialization for the rapid transmission of crucial social information in very large brains

Hermes, R., Behr, B., Hildebrandt, T.B., Blottner, S., Sieg, B., Frenzel, A., Knieriem, A., Saragusty, J., Rath, D., 2009. Sperm sex-sorting in the Asian elephant (Elephas maximus). Anim Reprod. Sci. 112, 390-396.
Abstract: In captive Asian elephants, there is a strong need for production of female offspring to enhance reproduction, counter premature aging processes in female animals and reduce challenging management situations derived from husbandry of several bulls in one institution. Artificial insemination of flow cytometrically sex-sorted spermatozoa offers the possibility to predetermine the sex of offspring with high accuracy. The aims of this study were to determine a suitable semen extender and basic parameters for flow cytometrical sex-sorting of Asian elephant spermatozoa. In total 18 semen samples were collected by manual rectal stimulation from one bull. Sperm quality parameters and sex sortability of spermatozoa were evaluated after dilution in three semen extenders (MES-HEPES-skim milk, MES-HEPES, TRIS-citric acid) and DNA staining. MES-HEPES-skim milk was the only semen extender found suitable to sex Asian elephant spermatozoa. From 18 ejaculates collected, 12 were successfully sorted with a purity of 94.5+/-0.7% at an average sort rate of 1945.5+/-187.5 spermatozoa per second. Sperm integrity, progressive and total motility were 42.6+/-3.9%, 48.1+/-3.3%, 59.4+/-3.8% after DNA labelling, and 64.8+/-3.2%, 58.0+/-5.0%, 70.8+/-4.4% after sorting, respectively. After liquid storage of sorted spermatozoa for 12h at 4 degrees C, sperm integrity, progressive and total motility were 46.4+/-5.2%, 32.2+/-4.2% and 58.2+/-3.9%, respectively. The obtained results provide a promising base to inseminate Asian elephants with sexed semen

Leighty, K.A., Soltis, J., Wesolek, C.M., Savage, A., Mellen, J., Lehnhardt, J., 2009. GPS determination of walking rates in captive African elephants (Loxodonta africana)
79. Zoo. Biol. 28, 16-28.
Abstract: The movements of elephants in captivity have been an issue of concern for animal welfare activists and zoological professionals alike in recent years. In order to fully understand how movement rates reflect animal welfare, we must first determine the exact distances these animals move in the captive environment. We outfitted seven adult female African elephants (Loxodonta africana) at Disney's Animal Kingdom with collar-mounted global positioning recording systems to document their movement rates while housed in outdoor guest viewing habitats. Further, we conducted preliminary analyses to address potential factors impacting movement rates including body size, temperature, enclosure size, and social grouping complexity. We found that our elephants moved at an average rate of 0.409+/-0.007 km/hr during the 9-hr data collection periods. This rate translates to an average of 3.68 km traveled during the observation periods, at a rate comparable to that observed in the wild. Although movement rate did not have a significant relationship with an individual's body size in this herd, the movements of four females demonstrated a significant positive correlation with temperature. Further, females in our largest social group demonstrated a significant increase in movement rates when residing in larger enclosures. We also present preliminary evidence suggesting that increased social group complexity, including the presence of infants in the herd, may be associated with increased walking rates, whereas factors such as reproductive and social status may constrain movements

Mason, G.J., Veasey, J.S., 2009. How should the psychological well-being of zoo elephants be objectively investigated?
47. Zoo. Biol.
Abstract: Animal welfare (sometimes termed "well-being") is about feelings - states such as "suffering" or "contentment" that we can infer but cannot measure directly. Welfare indices have been developed from two main sources: studies of suffering humans, and of research animals deliberately subjected to challenges known to affect emotional state. We briefly review the resulting indices here, and discuss how well they are understood for elephants, since objective welfare assessment should play a central role in evidence-based elephant management. We cover behavioral and cognitive responses (approach/avoidance; intention, redirected and displacement activities; vigilance/startle; warning signals; cognitive biases, apathy and depression-like changes; stereotypic behavior); physiological responses (sympathetic responses; corticosteroid output - often assayed non-invasively via urine, feces or even hair; other aspects of HPA function, e.g. adrenal hypertrophy); and the potential negative effects of prolonged stress on reproduction (e.g. reduced gametogenesis; low libido; elevated still-birth rates; poor maternal care) and health (e.g. poor wound-healing; enhanced disease rates; shortened lifespans). The best validated, most used welfare indices for elephants are corticosteroid outputs and stereotypic behavior. Indices suggested as valid, partially validated, and/or validated but not yet applied within zoos include: measures of preference/avoidance; displacement movements; vocal/postural signals of affective (emotional) state; startle/vigilance; apathy; salivary and urinary epinephrine; female acyclity; infant mortality rates; skin/foot infections; cardio-vascular disease; and premature adult death. Potentially useful indices that have not yet attracted any validation work in elephants include: operant responding and place preference tests; intention and vacuum movements; fear/stress pheromone release; cognitive biases; heart rate, pupil dilation and blood pressure; corticosteroid assay from hair, especially tail-hairs (to access endocrine events up to a year ago); adrenal hypertrophy; male infertility; prolactinemia; and immunological changes. Zoo Biol 28:1-19, 2009. (c) 2009 Wiley-Liss, Inc

Owens, M.J., Owens, D., 2009. Early age reproduction in female savanna elephants (Loxodonta africana) after severe poaching. African Journal of Ecology 47, 214-222.
Abstract: A 10-year study revealed that after severe poaching (> 93% killed) of elephants (Loxodonta africana) in Zambia's North Luangwa National Park (NLNP) during the 1970s and 1980s, the age of reproduction in females was greatly reduced. Fifty-eight per cent of births were delivered by females aged 8.5-14 years, an age at which elephants were reported to be sexually immature in nearby South Luangwa National Park (SLNP) before poaching. The mean age of females at first birth (AFB) (1993, 1994) was 11.3 years. Prior to poaching, the mean age AFB in SLNP was 16 years. The NLNP age structure and sex ratio were skewed, mean family unit size was reduced, and 37% of family units contained no females older than 15 years. Twenty-eight per cent of family units were comprised entirely of a single mother and her calf, and 8% of units consisted only of orphans who would have been considered sexually immature prior to poaching. Only 6% of survivors were older than 20 years, the age at which females in little-poached populations generally become most reproductively active. After a community-based conservation programme and the UN-CITES ban on the ivory trade were introduced, no elephants were recorded killed. In spite of a high reproductive rate, 6 years after poaching decreased, the density of the NLNP population had not increased, supporting predictions that the removal of older matriarchs from family units will have serious consequences on the recovery of this species.

Pinter-Wollman, N., Isabell, L.A., Hart, L.A., 2009. Assessing translocation outcome: Comparing behavioral and physiological aspects of translocated and resident African elephants (Loxodonta africana). Biological Conservation 142, 1116-1124.
Abstract: Evaluating translocation outcomes is important for improving wildlife management and conservation actions. Often, when quick decisions need to be made and long-lived animals with slow reproductionrates are translocated, traditional assessment methods such as long-term survival and reproductive successcannot be used for assessing translocation outcomes. Thus, alternative, seldom used, measures suchas comparing the behavior and physiology of translocated animals to those of local residents should beemployed to assess the translocated animals' acclimation to their new home. Here we monitored the survival,physiology, and behavior of translocated African elephants (Loxodonta africana) and comparedthese measures to the local resident population at the release site. Adult male and female translocatedelephants' death rates were higher than those of the local population. Furthermore, the mortality rateof translocated adult males and calves was greater than expected based on their proportion in the translocatedelephant population. No difference was found in stress hormone levels between the two populations,but the body condition of the translocated elephants was significantly poorer than that of the localpopulation throughout the study period. The behavioral time budgets of the translocated elephants convergedwith those of the local population over time. Finally, translocated elephants utilized habitat thatwas similar to their source site (hills and permanent rivers) more than did the local population. Based on these findings we recommend careful consideration of timing, release location, and individuals targetedin future elephant translocations. More broadly, we introduce and explore seldom used translocation assessment techniques.

Saragusty, J., Hermes, R., Goritz, F., Schmitt, D.L., Hildebrandt, T.B., 2009. Skewed birth sex ratio and premature mortality in elephants. Anim Reprod. Sci. 115, 247-254.
Abstract: Sex allocation theories predict equal offspring number of both sexes unless differential investment is required or some competition exists. Left undisturbed, elephants reproduce well and in approximately even numbers in the wild. We report an excess of males are born and substantial juvenile mortality occurs, perinatally, in captivity. Studbook data on captive births (CB, n=487) and premature deaths (PD, <5 years of age; n=164) in Asian and African elephants in Europe and North America were compared with data on Myanmar timber (Asian) elephants (CB, n=3070; PD, n=738). Growth in CB was found in three of the captive populations. A significant excess of male births occurred in European Asian elephants (ratio: 0.61, P=0.044) and in births following artificial insemination (0.83, P=0.003), and a numerical inclination in North American African elephants (0.6). While juvenile mortality in European African and Myanmar populations was 21-23%, it was almost double (40-45%) in all other captive populations. In zoo populations, 68-91% of PD were within 1 month of birth with stillbirth and infanticide being major causes. In Myanmar, 62% of juvenile deaths were at >6 months with maternal insufficient milk production, natural hazards and accidents being the main causes. European Asian and Myanmar elephants PD was biased towards males (0.71, P=0.024 and 0.56, P<0.001, respectively). The skewed birth sex ratio and high juvenile mortality hinder efforts to help captive populations become self-sustaining. Efforts should be invested to identify the mechanism behind these trends and seek solutions for them.

Saragusty, J., Hildebrandt, T.B., Behr, B., Knieriem, A., Kruse, J., Hermes, R., 2009. Successful cryopreservation of Asian elephant (Elephas maximus) spermatozoa. Anim Reprod. Sci. 115, 255-266.
Abstract: Reproduction in captive elephants is low and infant mortality is high, collectively leading to possible population extinction. Artificial insemination was developed a decade ago; however, it relies on fresh-chilled semen from just a handful of bulls with inconsistent sperm quality. Artificial insemination with frozen-thawed sperm has never been described, probably, in part, due to low semen quality after cryopreservation. The present study was designed with the aim of finding a reliable semen freezing protocol. Screening tests included freezing semen with varying concentrations of ethylene glycol, propylene glycol, trehalose, dimethyl sulfoxide and glycerol as cryoprotectants and assessing cushioned centrifugation, rapid chilling to suprazero temperatures, freezing extender osmolarity, egg yolk concentration, post-thaw dilution with cryoprotectant-free BC solution and the addition of 10% (v/v) of autologous seminal plasma. The resulting optimal freezing protocol uses cushioned centrifugation, two-step dilution with isothermal 285 m Osm/kg Berliner Cryomedium (BC) with final glycerol concentration of 7% and 16% egg yolk, and freezing in large volume by the directional freezing technique. After thawing, samples are diluted 1:1 with BC solution. Using this protocol, post-thaw evaluations results were: motility upon thawing: 57.2+/-5.4%, motility following 30 min incubation at 37 degrees C: 58.5+/-6.0% and following 3h incubation: 21.7+/-7.6%, intact acrosome: 57.1+/-5.2%, normal morphology: 52.0+/-5.8% and viability: 67.3+/-6.1%. With this protocol, good quality semen can be accumulated for future use in artificial inseminations when and where needed

Sherwood, C.C., Stimpson, C.D., Butti, C., Bonar, C.J., Newton, A.L., Allman, J.M., Hof, P.R., 2009. Neocortical neuron types in Xenarthra and Afrotheria: implications for brain evolution in mammals. Brain Struct. Funct. 213, 301-328.
Abstract: Interpreting the evolution of neuronal types in the cerebral cortex of mammals requires information from a diversity of species. However, there is currently a paucity of data from the Xenarthra and Afrotheria, two major phylogenetic groups that diverged close to the base of the eutherian mammal adaptive radiation. In this study, we used immunohistochemistry to examine the distribution and morphology of neocortical neurons stained for nonphosphorylated neurofilament protein, calbindin, calretinin, parvalbumin, and neuropeptide Y in three xenarthran species-the giant anteater (Myrmecophaga tridactyla), the lesser anteater (Tamandua tetradactyla), and the two-toed sloth (Choloepus didactylus)-and two afrotherian species-the rock hyrax (Procavia capensis) and the black and rufous giant elephant shrew (Rhynchocyon petersi). We also studied the distribution and morphology of astrocytes using glial fibrillary acidic protein as a marker. In all of these species, nonphosphorylated neurofilament protein-immunoreactive neurons predominated in layer V. These neurons exhibited diverse morphologies with regional variation. Specifically, high proportions of atypical neurofilament-enriched neuron classes were observed, including extraverted neurons, inverted pyramidal neurons, fusiform neurons, and other multipolar types. In addition, many projection neurons in layers II-III were found to contain calbindin. Among interneurons, parvalbumin- and calbindin-expressing cells were generally denser compared to calretinin-immunoreactive cells. We traced the evolution of certain cortical architectural traits using phylogenetic analysis. Based on our reconstruction of character evolution, we found that the living xenarthrans and afrotherians show many similarities to the stem eutherian mammal, whereas other eutherian lineages display a greater number of derived traits

Soltis, J., 2009. Vocal communication in African Elephants (Loxodonta africana)
61. Zoo. Biol. 28, 1-18.
Abstract: Research on vocal communication in African elephants has increased in recent years, both in the wild and in captivity, providing an opportunity to present a comprehensive review of research related to their vocal behavior. Current data indicate that the vocal repertoire consists of perhaps nine acoustically distinct call types, "rumbles" being the most common and acoustically variable. Large vocal production anatomy is responsible for the low-frequency nature of rumbles, with fundamental frequencies in the infrasonic range. Additionally, resonant frequencies of rumbles implicate the trunk in addition to the oral cavity in shaping the acoustic structure of rumbles. Long-distance communication is thought possible because low-frequency sounds propagate more faithfully than high-frequency sounds, and elephants respond to rumbles at distances of up to 2.5 km. Elephant ear anatomy appears designed for detecting low frequencies, and experiments demonstrate that elephants can detect infrasonic tones and discriminate small frequency differences. Two vocal communication functions in the African elephant now have reasonable empirical support. First, closely bonded but spatially separated females engage in rumble exchanges, or "contact calls," that function to coordinate movement or reunite animals. Second, both males and females produce "mate attraction" rumbles that may advertise reproductive states to the opposite sex. Additionally, there is evidence that the structural variation in rumbles reflects the individual identity, reproductive state, and emotional state of callers. Growth in knowledge about the communication system of the African elephant has occurred from a rich combination of research on wild elephants in national parks and captive elephants in zoological parks. Zoo Biol 28:1-18, 2009. (c) 2009 Wiley-Liss, Inc

Steinetz, B., Lasano, S., de Haas van, D.F., Glickman, S., Bergfelt, D., Santymire, R., Songsassen, N., Swanson, W., 2009. Relaxin concentrations in serum and urine of endangered and crazy mixed-up species
66. Ann. N. Y. Acad. Sci. 1160, 179-185.
Abstract: The human population explosion has pushed many mammalian wildlife species to the brink of extinction. Conservationists are increasingly turning to captive breeding as a means of preserving the gene pool. We previously reported that serum immunoactive relaxin provided a reliable means of distinguishing between true and pseudopregnancy in domestic dogs, and this method has since been found to be a reliable indicator of true pregnancy in endangered Asian and African elephants and Sumatran rhinoceroses. Our canine relaxin radioimmunoassay (RIA) has now been adapted and validated to measure relaxin in the serum and urine of felids, including domestic and wild species. Moreover, a commercially available canine serum relaxin kit (Witness) Relaxin Kit; Synbiotics, San Diego, CA), has been adapted for reliable detection of relaxin in urine of some felid species. Our porcine relaxin RIA has also been utilized to investigate the role of relaxin in reproductive processes of the spotted hyena, a species in which the female fetuses are severely masculinized in utero. Indeed, this species might well now be extinct were it not for the timely secretion of relaxin to enable copulation and birth of young through the clitoris. Additional studies have suggested relaxin may be a useful marker of pregnancy in the northern fur seal and the maned wolf (the former species has been designated as "depleted" and the latter as "near threatened"). Given appropriate immunoassay reagents, relaxin determination in body fluids thus provides a powerful tool for conservationists and biologists investigating reproduction in a wide variety of endangered and exotic species

Thitaram, C., Chansitthiwet, S., Pongsopawijit, P., Brown, J.L., Wongkalasin, W., Daram, P., Roongsri, R., Kalmapijit, A., Mahasawangkul, S., Rojanasthien, S., Colenbrander, B., van der Weijden, G.C., van Eerdenburg, F.J., 2009. Use of genital inspection and female urine tests to detect oestrus in captive Asian elephants
116. Anim Reprod. Sci. 115, 267-278.
Abstract: Captive Asian elephant (Elephas maximus) populations are decreasing due to low birth rates compared to wild elephants. Improving oestrous detection in female elephants is required to ensure successful mating in captive and semi-captive herds. Responsive behaviours of eight semi-captive bull elephants to the uro-genital area (genital inspection test) or urinary pheromones (urine test) of 14 female elephants throughout the oestrous cycle were evaluated. Weekly blood samples were collected for 27 consecutive months (14 months for the genital inspection test and 13 months for the urine test) from female elephants to characterize the patterns of circulating progestagen. Responsive behaviours of bulls were compared between females in the follicular versus the luteal phase of the cycle. The sensitivity and specificity of the genital inspection test were 65% and 68%, while those of the urine test were 52% and 61%, respectively. The bulls showed significantly higher "genital inspection", "flehmen from genital area" and "trunk on back" behaviours during the genital inspection test, and "flehmen" behaviours during the urine test in oestrous than in non-oestrous females. In sum, this study showed that monitoring sexual behaviours of Asian elephant bulls towards females or their urine can be used to detect the oestrous period. Although the sensitivity and specificity of both tests were not as high as expected, still, these methods appear to be more efficient at detecting oestrous than traditional methods based on mahout estimations of female receptivity. The use of genital inspection and urine tests may lead to more successful matings and thus to creating self-sustaining populations of captive elephants in range countries

Thitaram, C., Pongsopawijit, P., Chansitthiwet, S., Brown, J.L., Nimtragul, K., Boonprasert, K., Homkong, P., Mahasawangkul, S., Rojanasthien, S., Colenbrander, B., van der Weijden, G.C., van Eerdenburg, F.J., 2009. Induction of the ovulatory LH surge in Asian elephants (Elephas maximus): a novel aid in captive breeding management of an endangered species
49. Reprod. Fertil. Dev. 21, 672-678.
Abstract: A unique feature of the reproductive physiology of Asian elephants (Elephas maximus) is the occurrence of two LH surges before ovulation, instead of one. An anovulatory LH (anLH) surge, the function of which is unknown, occurs consistently 3 weeks before the ovulatory LH (ovLH) surge that induces ovulation. Thus, the ability to induce an ovLH surge would be useful for scheduling natural mating or artificial insemination. The present study tested the efficacy of a gonadotrophin-releasing hormone agonist (GnRH-Ag) to induce LH surges during the follicular phase of the oestrous cycle, which resulted in varied LH responses, but generally none were as high as previously documented natural surges. Thus, for the ovulation-induction trials, nine females were administered 80 microg GnRH-Ag intravenously at three time periods during the oestrous cycle, namely the anovulatory follicular phase, the ovulatory follicular phase and the luteal phase. During the late anovulatory follicular phase, nine of 10 females (90%) responded with an immediate LH surge followed 15-22 days later by an ovLH surge or a post-ovulatory increase in progestagens. In contrast, despite responding to the GnRH-Ag with an immediate increase in LH, none of the females treated during other periods of the oestrous cycle exhibited subsequent ovLH surges. One cow got pregnant from natural mating following the induced ovLH surge. In conclusion, ovLH induction is possible using a GnRH-Ag, but only during a specific time of the anovulatory follicular phase

Thongtip, N., Mahasawangkul, S., Thitaram, C., Pongsopavijitr, P., Kornkaewrat, K., Pinyopummin, A., Angkawanish, T., Jansittiwate, S., Rungsri, R., Boonprasert, K., Wongkalasin, W., Homkong, P., Dejchaisri, S., Wajjwalku, W., Saikhun, K., 2009. Successful artificial insemination in the Asian elephant (Elephas maximus) using chilled and frozen-thawed semen. Reprod. Biol. Endocrinol. 7, 75.
Abstract: BACKGROUND: Artificial insemination (AI) using frozen-thawed semen is well established and routinely used for breeding in various mammalian species. However, there is no report of the birth of elephant calves following AI with frozen-thawed semen. The objective of the present study was to investigate the fertilizing ability of chilled and frozen-thawed semen in the Asian elephant following artificial insemination (AI). METHODS: Semen samples were collected by from 8 bulls (age range, 12-to 42-years) by manual stimulation. Semen with high quality were either cooled to 4 degrees C or frozen in liquid nitrogen (-196 degrees C) before being used for AI. Blood samples collected from ten elephant females (age range, 12-to 52-years) were assessed for estrus cycle and elephants with normal cycling were used for AI. Artificial insemination series were conducted during 2003 to 2008; 55 and 2 AI trials were conducted using frozen-thawed and chilled semen, respectively. Pregnancy was detected using transrectal ultrasonography and serum progestagen measurement. RESULTS: One female (Khod) inseminated with chilled semen became pregnant and gave birth in 2007. The gestation length was 663 days and the sex of the elephant calf was male. One female (Sao) inseminated with frozen-thawed semen showed signs of pregnancy by increasing progestagen levels and a fetus was observed for 5 months by transrectal ultrasonography. CONCLUSION: This is the first report showing pregnancy following AI with frozen-thawed semen in the Asian elephant. Successful AI in the Asian elephant using either chilled or frozen-thawed semen is a stepping stone towards applying this technology for genetic improvement of the elephant population.

Trimble, M.J., Ferreira, S.M., van Aarde, R.J., 2009. Drivers of megaherbivore demographic fluctuations: inference from elephants. Journal of Zoology 1-9.
Abstract: Environmentally induced variation in survival and fecundity generates demographic fluctuations that affect population growth rate. However, a general pattern of the comparative influence of variation in fecundity and juvenile survival on elephant population dynamics has not been investigated at a broad scale. We evaluated the relative importance of conception, gestation, first year survival and subsequent survivorship for controlling demographic variation by exploring the relationship between past environmental conditions determined by integrated normalized difference vegetation index (INDVI) and the shape of age distributions at 17 sites across Africa. We showed that, generally, INDVI during gestation best explained anomalies in age structure. However, in areas with low mean annual rainfall, INDVI during the first year of life was critical. The results challenge Eberhardt's paradigm for population analysis that suggests that populations respond to limited resource availability through a sequential decrease in juvenile survival, reproductive rate and adult survival. Contrastingly, elephants appear to respond first through a reduction in reproductive rate. We conclude that this discrepancy is likely due to the evolutionary significance of extremely large body size - an adaptation that increases survival rate but decreases reproductive potential. Other megaherbivores may respond similarly to resource limitation due to similarities in population dynamics. Knowing how vital rates vary with changing environmental conditions will permit better forecasts of the trajectories of megaherbivore populations.

Tripp, K.M., Dubois, M., Delahaut, P., Verstegen, J.P., 2009. Detection and identification of plasma progesterone metabolites in the female Florida manatee (Trichechus manatus latirostris) using GC/MS/MS
54. Theriogenology 72, 365-371.
Abstract: Florida manatees (Trichechus manatus latirostris) have relatively low peripheral concentrations of progesterone (P4). The objective of this study was to determine if these relatively low P4 concentrations are associated with a high ratio of progestin metabolites and to document metabolite concentrations from individual blood samples obtained from manatees during diestrus or pregnancy. Metabolites known to exist in elephants-terrestrial manatee relatives-were targeted. These included 5alpha-reduced progestins (5alpha-pregnane-3,20-dione [5alpha-DHP] and 3alpha-hydroxy-5alpha-pregnan-20-one [5alpha-P3-OH]) and 17alpha-hydroxyprogesterone (17alpha-OHP), which occurs in Asian elephants. An additional, inactive metabolite, 20alpha-hydroxyprogesterone (20alpha-OHP), indicative of P4 overproduction, was also targeted. Progesterone itself was the predominant progestin detected in pregnant and nonpregnant manatee plasma (n = 10) using gas chromatography-mass spectrometry with tandem quadrupole detectors (GC/MS/MS). Progesterone concentrations in pregnant females varied from early (moderate to high) through mid and late (low) pregnancy. Progesterone concentrations ranged from low to high in nonpregnant, nonlactating females. The most commonly detected metabolite was 5alpha-P3-OH (n = 7), which occurred in pregnant (lower limit of detection [LLOD] to high) and nonpregnant (trace to high) females. The 5alpha-DHP metabolite was also detected in pregnant (LLOD to moderate) and nonpregnant (low) females. The 17alpha-OHP metabolite was not detected in any tested female. The 20alpha-OHP metabolite was detected in one nonpregnant, nonlactating, captive female (LLOD). Metabolites were most prevalent during early pregnancy, concurrent with maximum P4 concentrations. Based on their concentrations in peripheral circulation, we inferred that these metabolites may have, opposite to elephants, a limited physiologic role during luteal, pregnant, and nonpregnant phases in the manatee

Wallis, M., 2009. Prolactin in the Afrotheria: characterization of genes encoding prolactin in elephant (Loxodonta africana), hyrax (Procavia capensis) and tenrec (Echinops telfairi). J. Endocrinol. 200, 233-240.
Abstract: Pituitary prolactin shows an episodic pattern of molecular evolution, with occasional short bursts of rapid change imposed on a generally rather slow evolutionary rate. In mammals, episodes of rapid change occurred in the evolution of primates, cetartiodactyls, rodents and the elephant. The bursts of rapid evolution in cetartiodactyls and rodents were followed by duplications of the prolactin gene that gave rise to large families of prolactin-related proteins including placental lactogens, while in primates the burst was followed by corresponding duplications of the related GH gene. The position in elephant is less clear. Extensive data relating to the genomic sequences of elephant and two additional members of the group Afrotheria are now available, and have been used here to characterize the prolactin genes in these species and explore whether additional prolactin-related genes are present. The results confirm the rapid evolution of elephant (Loxodonta africana) prolactin - the sequence of elephant prolactin is substantially different from that predicted for the ancestral placental mammal. Hyrax (Procavia capensis) prolactin is even more divergent but tenrec (Echinops telfairi) prolactin is strongly conserved. No evidence was obtained from searches of public databases for additional genes encoding prolactin-like proteins in any of these species. Detailed analysis of evolutionary rates, and other factors, indicates that the episode of rapid change in hyrax, and probably elephant, was adaptive, though the nature of the associated biological change(s) is not clear

Weissenbock, N.M., Schwammer, H.M., Ruf, T., 2009. Estrous synchrony in a group of African elephants (Loxodonta africana) under human care. Anim Reprod. Sci. 113, 322-327.
Abstract: Synchrony of estrous, and consequently of conception and birth of young, may be of adaptive significance for certain mammals. Among the species in which estrous synchrony has been suspected several times are elephants, but clear evidence is still missing. We determined estrous cycles of African elephants (Loxodonta africana) (n=4) at the Vienna Zoo, Austria, between June 2003 and January 2006 by measuring serum progesterone levels from weekly blood samples. Except for the dominant female when she was intensively lactating, all animals showed clear cycles or progesterone release with a mean period of 105.3+/-15.37 days. For most of the study period, estrous cycles were asynchronous between females. However, after re-occurrence of the progesterone cycle in the dominant female following the first period of lactation, all four females showed high synchrony of progesterone release over the two subsequent cycles. Large changes in individual period lengths indicated that synchronization was due to the adjustment of cycle length in subdominants to that of the dominant female. We used a bootstrap procedure, based on resampling measured times of progesterone peaks, to determine if this apparent synchrony could have been caused by chance alone. This statistical analysis indicated that between-individual variances of the timing of progesterone peaks were much smaller that to be expected by chance (P=0.009). This finding represents the first evidence for estrous synchrony between elephants. We discuss various hypotheses to explain the biological function of cycle synchrony in elephants

Archie, E.A., Maldonado, J.E., Hollister-Smith, J.A., Poole, J.H., Moss, C.J., Fleischer, R.C., Alberts, S.C., 2008. Fine-scale population genetic structure in a fission-fusion society. Mol. Ecol. 17, 2666-2679.
Abstract: Nonrandom patterns of mating and dispersal create fine-scale genetic structure in natural populations - especially of social mammals - with important evolutionary and conservation genetic consequences. Such structure is well-characterized for typical mammalian societies; that is, societies where social group composition is stable, dispersal is male-biased, and males form permanent breeding associations in just one or a few social groups over the course of their lives. However, genetic structure is not well understood for social mammals that differ from this pattern, including elephants. In elephant societies, social groups fission and fuse, and males never form permanent breeding associations with female groups. Here, we combine 33 years of behavioural observations with genetic information for 545 African elephants (Loxodonta africana), to investigate how mating and dispersal behaviours structure genetic variation between social groups and across age classes. We found that, like most social mammals, female matrilocality in elephants creates co-ancestry within core social groups and significant genetic differentiation between groups (Phi(ST) = 0.058). However, unlike typical social mammals, male elephants do not bias reproduction towards a limited subset of social groups, and instead breed randomly across the population. As a result, reproductively dominant males mediate gene flow between core groups, which creates cohorts of similar-aged paternal relatives across the population. Because poaching tends to eliminate the oldest elephants from populations, illegal hunting and poaching are likely to erode fine-scale genetic structure. We discuss our results and their evolutionary and conservation genetic implications in the context of other social mammals

Aupperle, H., Reischauer, A., Bach, F., Hildebrandt, T., Goritz, F., Jager, K., Scheller, R., Klaue, H.J., Schoon, H.A., 2008. Chronic endometritis in an Asian elephant (Elephas maximus). J. Zoo. Wildl. Med. 39, 107-110.
Abstract: A 48-yr-old female Asian elephant with a history of pododermatitis developed recurrent hematuria beginning in 2002. Transrectal ultrasonography and endoscopic examination in 2004 identified the uterus as the source of hematuria and excluded hemorrhagic cystitis. Treatment with Desloreline implants, antibiotics, and homeopathic drugs led to an improved general condition of the elephant. In July 2005, the elephant was suddenly found dead. During necropsy, the severely enlarged uterus contained about 250 L of purulent fluid, and histopathology revealed ulcerative suppurative endometritis with high numbers of Streptococcus equi ssp. zooepidemicus and Escherichia coli identified on aerobic culture. Additional findings at necropsy included: multifocal severe pododermatitis, uterine leiomyoma, and numerous large calcified areas of abdominal fat necrosis. Microbiologic culture of the pododermatitis lesion revealed the presence of Streptococcus agalactiae, Streptococcus equi ssp. zooepidemicus, Staphylococcus sp., Corynebacterium sp., and Entercoccus sp

Drews, B., Hermes, R., Goritz, F., Gray, C., Kurz, J., Lueders, I., Hildebrandt, T.B., 2008. Early embryo development in the elephant assessed by serial ultrasound examinations. Theriogenology 69, 1120-1128.
Abstract: The elephant has an extraordinary long pregnancy, lasting 21 months. However, knowledge on embryo development is limited. To date, only single morphological observations of elephant embryo development associated with placentation are available, all lacking correlation to gestational age. The present study describes morphological characteristics of early embryo development in the elephant with exact biometric staging. Six pregnancies in five Asian and one African elephants with known conception dates were followed by 2D and 3D ultrasound, covering the embryonic period from ovulation to day 116 post-ovulation. The embryonic vesicle was earliest observed was on day 50 p.o. The proper embryo was not detected until day 62 p.o. Embryonic heartbeat was first observed on day 71 p.o. The allantois, which became visible as a single sacculation on day 71 p.o. was subdivided in four compartments on day 76 p.o. By day 95 p.o., head, rump, front and hind legs were clearly distinguished. Between days 95 and 103 p.o. the choriovitelline placenta was replaced by the chorioallantoic placenta. A physiological midgut herniation was transiently present between days 95 and 116 p.o. On the basis of the late appearance of the embryonic vesicle, delayed implantation in the elephant is discussed. The study provides a coherent description of elephant embryonic development, formation of the extraembryonic organs and their role in placenta formation, all of which are of interest for both comparative evolutionary studies and the improvement of assisted reproduction techniques

Evans, K., Harris, S., 2008. Adolescence in male African elephants, Loxodonta africana, and the importance of sociality. Animal Behavior 76, 779-787.
Abstract: The degree of sociality during an animal's life changes as it modulates its behaviour to reflect different lifestages. Only a few species of mammal undergo a period of adolescence, but for these species it is probablyone of their most important life stages. It is when individuals acquire skills and develop relationships thatare of both immediate and long-term benefit to their survival and reproductive success, particularly in polygynous males in which sexual selection favours size and dominance. We collected focal and observationaldata on male African elephants in the Okavango Delta, Botswana, to assess behaviour and socialinteractions during adolescence. Adolescent males (10-15 and 16-20 years of age) were the most sociableage group, showing preferences for larger social groupings and being in closer proximity to other elephants;later adolescent males (ages 16-20) showed a tendency for higher social levels. Males of all agespreferred to have males 36 years of age as their nearest neighbour. We argue that this proximity to oldermales provides opportunities for males to learn from more experienced individuals. It has long been recognized that matriarchs are the repositories of social and ecological knowledge within elephant breedingherds: we suggest that mature males are reservoirs for such knowledge within bull society.

Gobush, K.S., Mutayoba, B.M., Wasser, S.K., 2008. Long-term impacts of poaching on relatedness, stress physiology, and reproductive output of adult female african elephants. Conserv. Biol. 22, 1590-1599.
Abstract: Widespread poaching prior to the 1989 ivory ban greatly altered the demographic structure of matrilineal African elephant (Loxodonta africana) family groups in many populations by decreasing the number of old, adult females. We assessed the long-term impacts of poaching by investigating genetic, physiological, and reproductive correlates of a disturbed social structure resulting from heavy poaching of an African elephant population in Mikumi National Park, Tanzania, prior to 1989. We examined fecal glucocorticoid levels and reproductive output among 218 adult female elephants from 109 groups differing in size, age structure, and average genetic relatedness over 25 months from 2003 to 2005. The distribution in group size has changed little since 1989, but the number of families with tusked old matriarchs has increased by 14.2%. Females from groups that lacked an old matriarch, first-order adult relatives, and strong social bonds had significantly higher fecal glucocorticoid values than those from groups with these features (all females R(2)= 0.31; females in multiadult groups R(2)= 0.46). Females that frequented isolated areas with historically high poaching risk had higher fecal glucocorticoid values than those in low poaching risk areas. Females with weak bonds and low group relatedness had significantly lower reproductive output (R(2)[U]=0.21). Females from disrupted groups, defined as having observed average group relatedness 1 SD below the expected mean for a simulated unpoached family, had significantly lower reproductive output than females from intact groups, despite many being in their reproductive prime. These results suggest that long-term negative impacts from poaching of old, related matriarchs have persisted among adult female elephants 1.5 decades after the 1989 ivory ban was implemented

Hermes, R., Saragusty, J., Schaftenaar, W., Goritz, F., Schmitt, D.L., Hildebrandt, T.B., 2008. Obstetrics in elephants. Theriogenology 70, 131-144.
Abstract: Obstetrics, one of the oldest fields in veterinary medicine, is well described and practiced in domestic and exotic animals. However, when providing care during elephant birth or dystocia, veterinary intervention options differ greatly from any domestic species, and are far more limited due to the dimensions and specific anatomy of the elephant reproductive tract. In addition, aging of captive elephant populations and advanced age of primiparous females make active birth management increasingly important. Intrauterine infection, uterine inertia and urogenital tract pathologies are emerging as major causes for dystocia, often leading to foetal and dam death. This paper reviews the current knowledge on elephant birth and the factors associated with dystocia. It then summarises recommendations for birth and dystocia management. As Caesarean section, the most common ultima ratio in domestic animal obstetrics, is lethal and therefore not an option in the elephant, non-invasive medical treatment, induction of the Fergusson reflex or the conscious decision to leave a retained foetus until it is expelled voluntarily, are key elements in elephant obstetrics. Surgical strategies such as episiotomy and foetotomy are sometimes inevitable in order to try to save the life of the dam, however, these interventions result in chronic post-surgical complications or even fatal outcome. Limited reliable data on serum calcium concentrations, and pharmacokinetics and effect of exogenous oestrogen, oxytocin, and prostaglandins during birth provide the scope of future research, necessary to advance scientific knowledge on obstetrics in elephants

Hollister-Smith, J.A., Alberts, S.C., Rasmussen, L.E.L., 2008. Do male African elephants, Loxodonta africana, signal musth via urine dribbling? Animal Behavior 76, 1829-1841.
Abstract: The phenomenon of musth in male elephants involves increased sexual activity, heightened aggression and nearly continuous dribbling of pungent smelling urine. Urine chemistry during musth is altered, suggesting that urine may signal the musth status of the individual. Signalling musth remotely may benefit individuals if it reduces the likelihood of physical confrontation between males, which can lead to injury and even death. Few studies, however, have asked whether and how male elephants respond to urine of other males. We tested two predictions of the hypothesis that urine signals musth status to male conspecifics: (1) that male African elephants differentiate musth and nonmusth urine, and (2) that males differentiate between urine dribbled during early and late musth. The second prediction stems from the observation that males lose weight and presumably body condition during musth. We conducted two related bioassays with 26 captive nonmusth males ranging from 13 to 52 years of age. In each assay, subjects were simultaneously presented with three urine samples (nonmusth, early musth, late musth), each from a different donor male, and a control. We found that subjects differentiated between musth and nonmusth samples using their vomeronasal organ system, but did not discriminate between the samples using their main olfactory system. Males did not differentiate early from late musth. In addition, we found that subject contextual factors, specifically age, dominance status and social grouping, significantly predicted response. We discuss these results within the framework of male elephant longevity and social relationships and their importance to reproductive success.

Lotfy, W.M., Brant, S.V., DeJong, R.J., Le, T.H., Demiaszkiewicz, A., Rajapakse, R.P., Perera, V.B., Laursen, J.R., Loker, E.S., 2008. Evolutionary origins, diversification, and biogeography of liver flukes (Digenea, Fasciolidae). American Journal of Tropical Medicine and Hygiene 79, 248-255.
Abstract: Fasciolid flukes are among the largest and best known digenetic trematodes and have considerable historical and veterinary significance. Fasciola hepatica is commonly implicated in causing disease in humans. The origins, patterns of diversification, and biogeography of fasciolids are all poorly known. We have undertaken a molecular phylogenetic study using 28S, internal transcribed spacer 1 and 2 (ITS-1 and ITS-2) of nuclear ribosomal DNA, and mitochondrial nicotinamide dehydrogenase subunit 1 (nad1) that included seven of the nine recognized species in the family. The fasciolids examined comprise a monophyletic group with the most basal species recovered from African elephants. We hypothesize fasciolids migrated from Africa to Eurasia, with secondary colonization of Africa. Fasciolids have been conservative in maintaining relatively large adult body size, but anatomical features of their digestive and reproductive systems are available. These flukes have been opportunistic, with respect to switching to new snail (planorbid to lymnaeid) and mammalian hosts and from intestinal to hepatic habitats within mammals

Lynch, V.J., Tanzer, A., Wang, Y., Leung, F.C., Gellersen, B., Emera, D., Wagner, G.P., 2008. Adaptive changes in the transcription factor HoxA-11 are essential for the evolution of pregnancy in mammals. Proc. Natl. Acad. Sci. U. S. A 105, 14928-14933.
Abstract: Evolutionary change in gene regulation can result from changes in cis-regulatory elements, leading to differences in the temporal and spatial expression of genes or in the coding region of transcription factors leading to novel functions or both. Although there is a growing body of evidence supporting the importance of cis-regulatory evolution, examples of protein-mediated evolution of novel developmental pathways have not been demonstrated. Here, we investigate the evolution of prolactin (PRL) expression in endometrial cells, which is essential for placentation/pregnancy in eutherian mammals and is a direct regulatory target of the transcription factor HoxA-11. Here, we show that (i) endometrial PRL expression is a derived feature of placental mammals, (ii) the PRL regulatory gene HoxA-11 experienced a period of strong positive selection in the stem-lineage of eutherian mammals, and (iii) only HoxA-11 proteins from placental mammals, including the reconstructed ancestral eutherian gene, are able to up-regulate PRL from the promoter used in endometrial cells. In contrast, HoxA-11 from the reconstructed therian ancestor, opossum, platypus, and chicken are unable to up-regulate PRL expression. These results demonstrate that the evolution of novel gene expression domains is not only mediated by the evolution of cis-regulatory elements but can also require evolutionary changes of transcription factor proteins themselves

Meyer, J., Goodwin, T., Schulte, B., 2008. Intrasexual chemical communication and social responses ofcaptive female African elephants. Animal Behavior 76, 163-174.
Abstract: In matrilineal societies, competition between females can occur within and between social units. Dominance hierarchies reduce costly conflicts when reliable cues of status are available, and reproductive condition may alter individual or group status. Female African elephants live in matriarchal groups with linear dominance hierarchies occurring within and between groups; elephants use chemical signals to mediate social interactions. If reproductive condition has important implications for inter- or intragroup behaviour, then females should discriminate between chemical signal sources that reveal reproductive condition. We examined whether trunk-tip contacts between females within a social group were related to phase of oestrus. Observations were conducted on 21 reproductively viable females at nine zoological facilities in North America. Females in the follicular phase received contacts to the urogenital region at a higher rate than did luteal phase females, and contacts increased with approaching ovulation. This supports the existence of an oestrous signal. We also examined whether an oestrous signal was evident by female investigation of urine collected from the luteal and follicular phases of unfamiliar conspecifics. Elephants responded to unfamiliar urine more than to the control, but response rates to the urine types did not differ. Females within a social unit detected differences in oestrus, but they did not show such discrimination to urinary signals from unfamiliar females. Further evaluation of the existence of a female-to-female oestrous pheromone requires assessing responses to urine from familiar individuals. Understanding the relationship between oestrous condition and dominance status can shed light on the adaptive value of sociality.

Meyers, D.A., Isaza, R., MacNeill, A. Evaluation of acute phase proteins for diagnosis of inflammation in Asian elephants ( Elephas maximus). Proc American Associaton of Zoo Veterinarians and Assoc of Reptile and Amphibian Veterinarians.  128. 2008. 11-10-2008.
Ref Type: Conference Proceeding
Abstract: In many domestic species, routine hematology assays are useful diagnostic tools to diagnose inflammatory conditions. Unlike other species, these hematologic tests apparently are insensitive indicators of inflammation in elephants.1 We studied a novel group of blood proteins, called acute phase proteins, which increase during inflammatory conditions, for their usefulness in diagnosing elephants with inflammatory diseases. Although these proteins currently are useful in humans and domestic animals, each species has a different set of important proteins that must be individually investigated.2 We tested several acute phase proteins (C-reactive protein, alpha-1 glycoprotein, alpha-1 antitrypsin, serum amyloid A, haptoglobin, fibrinogen, ceruloplasmin, and albumin) as well as complete blood counts, chemistry panels, serum protein electrophoresis, and 3-D gel electrophoresis to determine their usefulness for diagnosing different types of inflammatory conditions in Asian elephants (Elephas maximus). Animals with inflammatory conditions were classified as those individuals with known illnesses such as mycobacteriosis, arthritis, nail bed abscesses, and malignant tumors. Control animals were thoseanimals that were suspected to not have any inflammation and be healthy at the time of testing as determined by physical examination and obtaining a thorough medical history.
LITERATURE CITED
1. Lyashchenko, K., R. Greenwald, J. Esfandiari, J. Olsen, R. Ball, G. Dumonceaux, F. Dunker, C. Buckley, M.
Richard, S. Murray, J.B. Payeur, P. Anderson, J.M. Pollock, S. Mikota, M. Miller, D. Sofranko, and W.R.
Waters. 2006. Tuberculosis in Elephants: Antibody responses to defined antigens of Mycobacterium
tuberculosis, potential for early diagnosis, and monitoring of treatment. Clin. Vacc. Immunol. 13: 722-732.
2. Murata H., N. Shimada, M. Yoshioka. 2004. Current research on acute phase proteins in veterinary diagnosis:
an overview. Vet J. 168: 28-40.

Miller, J., McClean, M. Pharmacokinetics of enrofloxacin in African elephants (Loxodonta africana) after a single rectal dose. Proc American Associaton of Zoo Veterinarians and Assoc of Reptile and Amphibian Veterinarians.  224-225. 2008. 11-10-2008.
Ref Type: Conference Proceeding
Abstract: Captive African elephants (Loxodonta Africana) are susceptible to many types of gram negative bacterial infections such as Escherichia coli, Mycoplasma  spp., Salmonella spp., Klebsiella spp., Pseudomonas spp., and Proteus spp. Enrofloxacin (Baytril®, Bayer Health Care, Animal Health Division, P.O. Box 390, Shawnee Mission, KS 66201) is a potentially effective antibiotic for
treatment of these bacterial infections in elephants. Very limited data exists on the pharmacokinetics of enrofloxacin in elephants2 and most of the dosage regimes for gastrointestinal absorption are based on horse dosages since they share a similar  gastrointestinal tract. Three African elephants from Wildlife Safari in Winston, Oregon, two females both 37-yr-old and one male 26-yr-old, were used to determine whether therapeutic levels of enrofloxacin could be achieved thru rectal administration of liquid injectable enrofloxacin (Baytril 100®, 100 mg/ml, Bayer Health Care, Animal Health Division, P.O. Box 390, Shawnee Mission, KS 66201) at a dosage of 2.5 mg/kg. A pretreatment baseline blood sample was collected. Following administration, blood samples were collected at 45 min, 1.5hr, 2.5hr, 5hr, 9hr, 23hr, 36hr to determine plasma enrofloxacin levels. Plasma enrofloxacin levels were measured at North Carolina State University, College of Veterinary Medicine using high performance liquid chromatography (HPLC) analysis. Plasma ciprofloxacin levels were measured concurrently. Results indicate plasma concentrations of enrofloxacin did not reach adequate bacteriocidal levels for any of the the following common bacterial isolates in captive elephants: Mycoplasma
spp., Escherichia coli, Salmonella spp., Klebsiella spp., Pseudomonas spp., and Proteus spp. The study determined that a rectally administered dosage of 2.5 mg/kg of liquid injectable enrofloxacin was insufficient to obtain therapeutic levels in African elephants. The low plasma levels of enrofloxacin in all three elephants may be a result of poor absorption in the distal large intestine. A future study will determine if oral administration will provide a more efficient mode of drug delivery and absorption in African elephants. It is also possible that the current dosage of 2.5 mg/kg is too low to achieve adequate therapeutic levels.
ACKNOWLEDGMENTS
I would like to thank the elephant and veterinary staff at Wildlife Safari for their participation in conducting this study. Thanks to Doctors: Modesto McClean, Jason Bennett, Andi Chariffe, Tessa Lohe, Benji Alacantar. Also thanks to Dinah Wilson, Carol Matthews, Anthony Karels, Mary Iida, Shawn Finnell, Patches Stroud, Katie Alayan.
LITERATURE CITED
1. Haines, G.R., et. al. 2000. Serum concentrations and pharmacokinetics of enrofloxacin after intravenous and intragastric administration to mares. Can. J.Vet. Res. 64(3):171-177.
2. Sanchez, C.R., et. al. 2005. Pharmacokinetics of a single dose of enrofloxacin administered orally to captive Asian elephants (Elephas maximus). Am. J. Vet. Res. 66:1948-1953.

Nicholls, H., 2008. Darwin 200: Let's make a mammoth. Nature 456, 310-314.

Rasmssen, H.B., Ganswindt, A., Douglas-Hamilton, I., Vollrath, F., 2008. Endocrine and behavioral changes in male African elephants: Linking hormone changes to sexual state and reproductive tactics. Hormones and Behavior.
Abstract: Endocrine and behavioral changes in male African elephants: Linking hormone changes to sexual state and reproductive tactics.Henrik B Rasmussen, Andre Ganswindt, Iain Douglas-Hamilton, and Fritz VollrathHormones and Behavior, May 22, 2008                     
Hormones play a crucial role in mediating genetic and environmental effects into morphological and behavioral phenotypes. In systems with alternative reproductive tactics (ART) shifts between tactics are hypothesized to be under proximate hormonal control. Most studies of the underlying endocrine changes behind ART have focused on fish and amphibians rather than mammals and few have investigated the potential interaction between different endocrine axes in regulating shifts between conditional dependent tactics. Using a combination of endocrine and behavioral data from male African elephants we expand on our previously published analysis and show that the initial increase in androgens predates the behavioral shifts associated with reproductively active periods, supporting the role of androgens in activating sexually active periods in males. A strong interactive effect between androgens and glucocorticoids was found to determine the presence or absence of temporal gland secretion and urine dribbling, signals associated with the competitive reproductive tactic of musth, with elevated glucocorticoids levels suppressing the occurrence of musth signals. In addition external environmental conditions affected hormone levels. The presence of receptive females resulted in elevated androgens in dominant musth males but increased glucocorticoids in subordinate non-musth males. The presented data on hormones, behavior and reproductive tactics strongly support an underlying endocrine mechanism for mediating the translation of intrinsic as well as extrinsic local conditions into the conditional dependent reproductive tactics in male elephants via interactions between the hypothalamic-pituitary-gonadal and -adrenal axes.

Rasmussen, H.B., Ganswindt, A., Douglas-Hamilton, I., Vollrath, F., 2008. Endocrine and behavioral changes in male African elephants: linking hormone changes to sexual state and reproductive tactics. Horm. Behav. 54, 539-548.
Abstract: Hormones play a crucial role in mediating genetic and environmental effects into morphological and behavioral phenotypes. In systems with alternative reproductive tactics (ART) shifts between tactics are hypothesized to be under proximate hormonal control. Most studies of the underlying endocrine changes behind ART have focused on fish and amphibians rather than mammals and few have investigated the potential interaction between different endocrine axes in regulating shifts between conditional dependent tactics. Using a combination of endocrine and behavioral data from male African elephants we expand on our previously published analysis and show that the initial increase in androgens predates the behavioral shifts associated with reproductively active periods, supporting the role of androgens in activating sexually active periods in males. A strong interactive effect between androgens and glucocorticoids was found to determine the presence or absence of temporal gland secretion and urine dribbling, signals associated with the competitive reproductive tactic of musth, with elevated glucocorticoids levels suppressing the occurrence of musth signals. In addition external environmental conditions affected hormone levels. The presence of receptive females resulted in elevated androgens in dominant musth males but increased glucocorticoids in subordinate non-musth males. The presented data on hormones, behavior and reproductive tactics strongly support an underlying endocrine mechanism for mediating the translation of intrinsic as well as extrinsic local conditions into the conditional dependent reproductive tactics in male elephants via interactions between the hypothalamic-pituitary-gonadal and -adrenal axes

Schmitt, D., Charmason, S., Wiedner, E. Use of luteinizing hormone ELISAs  in breeding elephants. Proc American Associaton of Zoo Veterinarians and Assoc of Reptile and Amphibian Veterinarians.  120-121. 2008. 11-10-2008.
Ref Type: Conference Proceeding
Abstract: Successful artificial insemination (AI) of elephants depends heavily on determining the unique luteinizing hormone (LH) surges that occur during the follicular phase of the elephant's estrous cycle. Natural breeding of elephants also can benefit from a rapid and accurate determination of the two LH surges found in elephants. There are three ELISAs available for determining the LH
surge; two are commercially-available assays and one is a laboratory in-house assay. Each vary in their cost, time to complete the assay, and ease of performing the procedures. Detection of the initial non-ovulatory peak in luteinizing hormone (LH1) is best accomplished by use of an in house LH assay, or use of the LH assay available from Dr. Nancy Dahl (UC-Davis, Davis, CA
95616 USA), both of which are quantitative assays for detection of LH. For cow-side use during estrus, the qualitative ELISA Witness® LH Ovulation Timing Test Kit (Symbiotics Corporation, Kansas City, MO 64163 USA) detects LH in elephants within 20 min. This assay requires a minimum of laboratory precision to detect the ovulatory LH peak (LH2).
Introduction
Elephants are the only species known to exhibit a double LH peak during a single estrous cycle.2,4 Increased success of artificial insemination in elephants occurred partly in response to the ability to detect the LH1 surge about 21 days prior to the ovulatory LH2 surge that occurs at the end of a two to three day estrus.1 The first reports regarding detection of the double LH
surges were performed in laboratories using custom ELISA technology that require exacting procedures and two days to complete the quantitative assays.2,4 A semi-quantitative elephant LH ELISA that can be performed in the field in about 2.5 hr was developed at UC-Davis.3 A qualitative LH assay was developed for use in dogs and cats that uses a latex strip ELISA. The time for development of the test is 20 min and detects a LH surge greater than 1 ng/ml using serum. Elephants have LH1 and LH2 surges in the 4-16 ng/ml range,2,4 well within the detectable range for all of the assays described. The detection of the LH1 peak usually is from daily samples submitted weekly; this allows some efficiency of assay resources and provides at least a two-wk notice of LH2. However, accurate and timely detection of LH2 is needed at least daily and at times twice daily during estrus. The use of an LH assay which can be performed 'cow-side' and accurately detect LH2 is essential for successful AI and can be helpful in determining estrus status for natural breeding. The Witness® LH Ovulation Timing Test Kit from Symbiotics was developed for use in dogs and cats, but is effective in other species, including elephants, and meets these requirements.
Discussion
Detection of LH1 provides information for predicting the LH2 surge and performance of assays that require more laboratory time and precision are useful since detection of LH1 is not as timesensitive as LH2 detection. Both of the quantitative assays have unique advantages. An inhouse assay can be set up, but requires greater preparation time, precision of laboratory procedures is more demanding, often takes two days to perform, and is more susceptible to environmental variables. The assay developed by UC-Davis costs about $5.00 per well, takes about 2.5 hr to perform and is more stable. However, for quantitative results the overhead costs of the standard curve requires about 16 wells ($90), plus two wells for each unknown sample. The UC-Davis assay can be set up as a qualitative test with high and low controls and no standard curve. This requires from three to six wells for a single sample. The Witness® LH Ovulation Timing Test Kit has a control built into each test strip and costs about $25.00 per sample. Because 'cow-side' testing possible using the Witness® LH Ovulation Timing Test Kit, I recommend its use for detection of LH2, although the UC-Davis Elephant ELISA is competitively priced and can be performed in a nearby temporary laboratory. Because timing is
critical in detecting LH2 and performing subsequent AI, I recommend using the Witness® LH Ovulation Timing Test Kit at the time of estrus, preceded by either one of the other assays for detecting LH1, depending on availability of laboratory labor and equipment.
LITERATURE CITED
1. Brown, J. L., F. Goritz, N. Pratt-Hawkes, R. Hermes, M. Galloway, L. H. Graham, C. Gray, S. L. Walker, A. Gomez, R. Moreland, S. Murray, D. L. Schmitt, J. G. Howard, J. Lehnhardt, B. Beck, A. Bellem, R. Montali, and T. B. Hildebrandt. 2004. Successful artificial insemination of an Asian elephant at the National Zoological Park. Zoo Biol. 23: 45-63.
2. Brown, J. L., D. L. Schmitt, A. Bellem, L. H. Graham, and J. Lehnhardt. 1999. Hormone secretion in the Asian elephant (Elephas maximus): Characterization of ovulatory and anovulatory luteinizing hormone surges. Biol. Reprod. 61: 1294-1299.
3. Dahl, N. J., D. Olson, D. L. Schmitt, D. R. Blasko, R. S. Kristipati, and J. F. Roser. 2004. Development of an enzyme-linked immunosorbent assay (ELISA) in the elephant (Loxodonta africana and Elephas maximus). Zoo Biol. 23: 65-78.
4. Kapustin, N., J. K. Critser, D. Olson, and P. V. Malven. 1996. Nonluteal estrous cycles of 3-week duration are initiated by anovulatory luteinizing hormone peaks in African elephants. Biol. Reprod. 55:1147-1154.

Slade-Cain, B.E., Rasmussen, L.E., Schulte, B.A., 2008. Estrous state influences on investigative, aggressive, and tail flicking behavior in captive female Asian elephants
78. Zoo. Biol. 27, 167-180.
Abstract: Females of species that live in matrilineal hierarchies may compete for temporally limited resources, yet maintain social harmony to facilitate cohesion. The relative degree of aggressive and nonaggressive interactions may depend on the reproductive condition of sender and receiver. Individuals can benefit by clearly signaling and detecting reproductive condition. Asian elephants (Elephas maximus) live in social matrilineal herds. Females have long estrous cycles (14-16 weeks) composed of luteal (8-12 weeks) and follicular (4-8 weeks) phases. In this study, we observed the behavior of four captive Asian elephant females during multiple estrous cycles over 2 years. We evaluated whether investigative, aggressive, and tail flicking behaviors were related to reproductive condition. Investigative trunk tip contacts showed no distinct pattern by senders, but were more prevalent toward female elephants that were in their follicular compared with their luteal phase. The genital area was the most frequently contacted region and may release reproductively related chemosignals. Aggression did not differ significantly with estrus; however, rates of aggression were elevated when senders were approaching ovulation and receivers were in the luteal phase. Females in the follicular phase may honestly advertise their condition. Contacts by conspecifics may serve to assess condition and reduce aggression. A behavior termed "tail flicking" was performed mainly during the mid-follicular phase when estrogen and luteinizing hormone levels are known to spike. Tail flicking may disperse chemical signals in urine or mucus as well as act as a tonic signal that could provide a means of anticipating forthcoming ovulation by elephants and also for human observers and caretakers. Zoo Biol 27:167-180, 2008. (c) 2008 Wiley-Liss, Inc

Steinmetz, H.W., Eulenberger, U., Hatt, J.M. Daily clinical examinations in a herd of captive asian elephants. Proc American Associaton of Zoo Veterinarians and Assoc of Reptile and Amphibian Veterinarians.  124. 2008. 11-10-2008.
Ref Type: Conference Proceeding
Abstract: The captive population of Asian elephants (Elephas maximus) is not self-sustaining.2 Poor reproduction and high juvenile mortality are key factors in the decreasing population. Infection with endotheliotropic elephant herpes virus (EEHV) is one of the major causes of death in the captive population, and has resulted in the loss of at least 40 captive animals.1 EEHV has been
responsible for the peracute death of two juvenile males at Zurich Zoo, Switzerland. Mortality due to peracute infection with EEHV mainly is seen in juveniles. Early detection of characteristic clinical signs of EEHV and immediate initiation of therapy are of crucial
importance due to its rapid progression. Based on past fatal EEHV experiences, Zurich Zoo modified its daily clinical health monitoring program to increase staff awareness of EEHV infection. Examinations have been incorporated into the daily routine and include daily evaluation of behaviour, appetite, colour of mucosal membranes and the measurement of body temperature; these examinations are performed by keepers. In our experiences, characteristic signs of acute EEHV infection are lethargy, anorexia, mild
colic, and cyanosis of the mucosal membranes. Results of temperature measurements have shown that best estimations of body temperature are done by measurement of the temperature in the centre of a fecal ball 5-9 min after defecation. Mean values of 36.5°C (± 0.2°C SD) are within published reference values, although adult elephants have shown significantly lower body temperature than juveniles. Establishment of individual reference values for each elephant is essential to detect unusual temperature peaks that may indicate possible EEHV viremia. The present study has shown that daily health examinations increase the awareness of keepers for
early signs of EEHV infection (e.g., peaks in body temperature and cyanotic mucosal membranes).
ACKNOWLEDGMENTS
The authors thank B. Aeschbach and all elephant keepers for taking special care of our elephants. The work and organization of Ms. G. Hürlimann is gratefully appreciated.
LITERATURE CITED
1. Mikota, S. 2007. Endotheliotropic Herpesvirus (EEHV). http://www.elephantcare.org/herpes.htm. cited: 10.04.2008:
2. Wiese, R. J. 2000. Asian elephants are not self-sustaining in North America. Zoo Biol. 19: 299-309.

Thongtip, N., Saikhun, J., Mahasawangkul, S., Kornkaewrat, K., Pongsopavijitr, P., Songsasen, N., Pinyopummin, A., 2008. Potential factors affecting semen quality in the Asian elephant (Elephas maximus). Reprod. Biol. Endocrinol. 6, 9.
Abstract: BACKGROUND: One of the major obstacles in using artificial insemination to manage genetics of elephant population in captivity is the large variations in semen quality among ejaculates within the same and among individuals. The objectives of this study were to determine the influences of (1) age (2) seasonality (3) and circulating testosterone (SrTest), triiodothyronine (SrT3) and tetraiodothyronine (SrT4), as well as seminal (4) testosterone (SpTest), zinc (SpZn) and protein (SpTP) on semen quality in the Asian elephant METHODS: Analyses, including motility, viability and morphology were performed in semen samples collected twice monthly from 13 elephant bulls (age range, 10-to 72-years) by manual stimulation between July 2004 and June 2005. Serum samples obtained monthly were assessed for SrTest, SrT3, SrT4, and seminal plasma samples were evaluated for, SpTest, SpZn and SpTP. RESULTS: The highest semen quality was observed at age 23 to 43 years. Percentages of progressive motility and viable sperm were lowest at age 51 to 70 years (P < 0.05); on the other hand, sperm concentration was lowest at age 10 to 19 years (P < 0.05). Percentage of sperm with normal morphology was highest at age 23 to 43 years. The levels of SrT3, SrTest, SpTest and SpZn were lowest at age 51 to 70 years, whereas SrT4 was lowest at age 23 to 43 years. Seasonality significantly affected semen characteristics in which percentage of viable sperm and cell concentration were highest during rainy season and lowest during summer months (P < 0.05). However, percentage of sperm with normal morphology was highest in summer and lowest in rainy season (P < 0.05). Seasonality significantly influenced SrTest with elevated concentrations observed in rainy season and winter (P < 0.05). CONCLUSION: This study indicates that age and seasonality had influence on semen characteristics in the Asian elephant. The knowledge obtained in this study will improve our understanding of the reproductive biology of this species

Turner, J.W., Rutberg, A.T., Naugle, R.E., Kaur, M.A., Flamagan, D.R., Bertschinger, H.J., Liu, I.K.M., 2008. Controlled-release components of PZP contraceptive vaccine extend duration of infertility. Wildlife Research 35, 555-562.
Abstract: Successful immunocontraception of wildlife relying on repeated access to individuals for boosters has highlighted the need to incorporate primer and booster immunisations into one injection. We have investigated use of controlled-release polymers (lactide-glycolide) in small pellets to provide delayed in vivo delivery of booster porcine zona pellucida (PZP) antigen and adjuvant. This report reviews pellet-making methodology, in vitro testing of controlled-release pellets and in vivo effects of controlled-release PZP vaccine. We assessed 3 different manufacturing approaches for producing reliable, cost-effective pellets: (1) polymer melting and extrusion; (2) solvent evaporation from polymer solution; and (3) punch and die polymer moulding. In vitro testing of release patterns of controlled-release formulations, towards development of a 3-year duration vaccine, provided estimates for in vivo use of pellet preparations. These in vitro studies demonstrated protein release delay up to 22 months using 100% l-lactide or polycaprolactone polymers. For in vivo tests, pellets (1-, 3-, and 12-month release delay) serving as boosters were administered intramuscularly with PZP/adjuvant liquid primer to wild horses (Equus caballus), white-tailed deer (Odocoileus virginanus) and African elephants (Loxodonta africana). Horse. field studies assessed fertility via offspring counts and/or faecal-hormone pregnancy testing. Treatment decreased fertility 5.3-9.3-fold in Year 1 and 3.6-fold in Year 2. In preliminary testing in deer, offspring counts revealed treatment-associated fertility reduction of 7.1-fold Year 1 and 3.3-fold Year 2. In elephants, treatment elevated anti-PZP titres 4.5-6.9- fold from pretreatment (no fertility data).

van der Kolk, J.H., van Leeuwen, J.P., van den Belt, A.J., van Schaik, R.H., Schaftenaar, W., 2008. Subclinical hypocalcaemia in captive Asian elephants (Elephas maximus). Veterinary Record 162, 475-479.
Abstract: The hypothesis that hypocalcaemia may play a role in dystocia in captive Asian elephants (Elephas maximus) was investigated. The objectives of the study were to measure the total calcium concentration in elephant plasma; assess the changes in parameters of calcium metabolism during a feeding trial; investigate a possible relationship between calcium metabolism and dystocia; and assess bone mineralisation in captive Asian elephants in vivo. The following parameters were measured: total and ionised calcium, inorganic phosphorous and magnesium, the fractional excretions of these minerals, intact parathyroid hormone, 25-OH-D(3) and 1,25-OH-D(3). Radiographs were taken from tail vertebrae for assessment of bone mineralisation. The mean (sd) heparinised plasma total calcium concentration was 2.7 (0.33) mmol/l (n=43) ranging from 0.84 to 3.08 mmol/l in 11 Asian elephants. There was no significant correlation between plasma total calcium concentration and age. Following feeding of a calcium rich ration to four captive Asian elephant cows, plasma total and ionised calcium peaked at 3.6 (0.24) mmol/l (range 3.4 to 3.9 mmol/l) and 1.25 (0.07) mmol/l (range 1.17 to 1.32 mmol/l), respectively. Plasma ionised calcium concentrations around parturition in four Asian elephant cows ranged from 0.37 to 1.1 mmol/l only. The present study indicates that captive Asian elephants might be hypocalcaemic, and that, in captive Asian elephants, the normal plasma concentration of total calcium should actually be around 3.6 mmol/l and normal plasma concentration of ionised calcium around 1.25 mmol/l. Given the fact that elephants absorb dietary calcium mainly from the intestine, it could be concluded that elephants should be fed calcium-rich diets at all times, and particularly around parturition. In addition, normal values for ionised calcium in captive Asian elephants should be reassessed

Viijoen, J.J., Ganswindt, A., du Toit, J.T., Langbauer, W.R., 2008. Translocation stress and faecal glucocorticoid metabolite levels in free-ranging African savanna elephants. South African Journal of Wildlife Research 38, 146-152.
Abstract: There are local populations of African elephants (Loxodonta africana) which have increased to levels where they are implicated in altering vegetation types. The local reduction of elephant numbers for wildlife management objectives can involve contraception, killing excess animals, or translocation to alternative habitats. The effects these management decisions can have on the physiological stress response of free-ranging African savanna elephants are still not fully understood. We examined the effect of translocation on faecal glucocorticoid metabolite levels of an African elephant family group, which was translocated within the Kruger National Park, South Africa. We found that translocation resulted in a significant increase in faecal glucocorticoid metabolite levels (up to 646 ng/g wet weight) compared to (1) pre-translocation levels in this group, (2) post-translocation levels in this group, and (3) levels measured in undisturbed 'control' groups in the area. However, the faecal glucocorticoid metabolite levels had returned to <100 ng/g by the time the translocated animals had navigated their way back to their previous home range, covering 300 km in 23 days.

von, A., I, Nimzyk, R., Klemke, M., Bullerdiek, J., 2008. A microRNA encoded in a highly conserved part of the mammalian HMGA2 gene. Cancer Genet. Cytogenet. 187, 43-44.
Abstract: The high mobility group protein HMGA2 plays an important role as a chromatin component of stem cells and as a protein causally related to the development of a variety of benign tumors (e.g., uterine leiomyomas, lipomas, and pleomorphic adenomas of the salivary glands). Herein, the existence of a highly conserved region within intron 3 of HMGA2 encoding a microRNA is described. The co-expression with HMGA2 suggests that as an intronic microRNA, this microRNA may cooperate with HMGA2 in its physiological and/or aberrant functions

Wittemyer, G., Polansky, L., Douglas-Hamilton, I., Getz, W.M., 2008. Disentangling the effects of forage, social rank, and risk on movement autocorrelation of elephants using Fourier and wavelet analyses. Proc. Natl. Acad. Sci. U. S. A 105, 19108-19113.
Abstract: The internal state of an individual-as it relates to thirst, hunger, fear, or reproductive drive-can be inferred by referencing points on its movement path to external environmental and sociological variables. Using time-series approaches to characterize autocorrelative properties of step-length movements collated every 3 h for seven free-ranging African elephants, we examined the influence of social rank, predation risk, and seasonal variation in resource abundance on periodic properties of movement. The frequency domain methods of Fourier and wavelet analyses provide compact summaries of temporal autocorrelation and show both strong diurnal and seasonal based periodicities in the step-length time series. This autocorrelation is weaker during the wet season, indicating random movements are more common when ecological conditions are good. Periodograms of socially dominant individuals are consistent across seasons, whereas subordinate individuals show distinct differences diverging from that of dominants during the dry season. We link temporally localized statistical properties of movement to landscape features and find that diurnal movement correlation is more common within protected wildlife areas, and multiday movement correlations found among lower ranked individuals are typically outside of protected areas where predation risks are greatest. A frequency-related spatial analysis of movement-step lengths reveal that rest cycles related to the spatial distribution of critical resources (i.e., forage and water) are responsible for creating the observed patterns. Our approach generates unique information regarding the spatial-temporal interplay between environmental and individual characteristics, providing an original approach for understanding the movement ecology of individual animals and the spatial organization of animal populations

Yon, L., Chen, J., Moran, P., Lasley, B., 2008. An analysis of the androgens of musth in the Asian bull elephant (Elephas maximus). Gen. Comp Endocrinol. 155, 109-115.
Abstract: During musth in bull elephants, the androgens testosterone (T), dihydrotestosterone (DHT), and androstenedione all increase significantly. Given the unusual endocrine physiology that has been discovered in female elephants, it is also possible that bull elephants produce some unusual androgens. A cell-based androgen receptor assay was used to explore this possibility using two different methods. The first method compared the level of T measured by radioimmunoassay (RIA) with the level of androgen receptor (AR) activity measured in the serum of eight bull elephants during musth and non-musth periods. A ratio was calculated for T/AR activity for non-musth and musth, to determine if there was a change in the ratio between these two states. The second method used HPLC to separate two pooled serum samples (one non-musth and one musth) into fractions using a protocol which separates known androgens into specific, previously identified fractions. Each fraction was then tested with the AR assay to determine the androgenicity of any compounds present. This was done to determine if there were any fractions which had androgenic activity but did not contain any previously identified androgens. Results from the first analysis indicated no change in the T/AR ratio between non-musth and musth states. Clearly whatever active androgens are present during musth, they increase proportionately with T. Findings from the second analysis suggested that the only bioactive androgen present in the serum of non-musth Asian bulls is a low level of T. During musth, the only bioactive androgens detected were T and DHT; of these, T was by far the predominant active androgen present. Taken together, these two analyses suggest that T is by far the predominant active androgen present during musth in Asian bull elephants, and that no previously unidentified bioactive androgen is present

Archie, E.A., Hollister-Smith, J.A., Poole, J.H., Lee, P.C., Moss, C.J., Maldonado, J.E., Fleischer, R.C., Alberts, S.C., 2007. Behavioural inbreeding avoidance in wild African elephants. Molecular Ecology 16, 4138-4148.
Abstract: The costs of inbreeding depression, as well as the opportunity costs of inbreeding avoidance, determine whether and which mechanisms of inbreeding avoidance evolve. In African elephants, sex-biased dispersal does not lead to the complete separation of male and female relatives, and so individuals may experience selection to recognize kin and avoid inbreeding. However, because estrous females are rare and male-male competition for mates is intense, the opportunity costs of inbreeding avoidance may be high, particularly for males. Here we combine 28 years of behavioural and demographic data on wild elephants with genotypes from 545 adult females, adult males, and calves in Amboseli National Park, Kenya, to test the hypothesis that elephants engage in sexual behaviour and reproduction with relatives less often than expected by chance. We found support for this hypothesis: males engaged in proportionally fewer sexual behaviours and sired proportionally fewer offspring with females that were natal family members or close genetic relatives (both maternal and paternal) than they did with nonkin. We discuss the relevance of these results for understanding the evolution of inbreeding avoidance and for elephant conservation.

Brown, J.L., Somerville, M., Riddle, H.S., Keele, M., Duer, C.K., Freeman, E.W., 2007. Comparative endocrinology of testicular, adrenal and thyroid function in captive Asian and African elephant bulls. Gen. Comp Endocrinol. 151, 153-162.
Abstract: Concentrations of serum testosterone, cortisol, thyroxine (free and total T4), triiodothyronine (free and total T3) and thyroid stimulating hormone (TSH) were measured to assess adrenal and thyroid function as they relate to testicular activity and musth in captive elephants. Blood samples were collected approximately weekly from Asian (n=8) and African (n=12) bulls at seven facilities for periods of 4 months to 9.5 years. Age ranges at study onset were 8-50 years for Asian and 10-21 years for African elephants. Based on keeper logs, seven Asian and three African bulls exhibited behavioral and/or physical (temporal gland secretion, TGS, or urine dribbling, UD) signs of musth, which lasted 2.8+/-2.5 months in duration. Serum testosterone was elevated during musth, with concentrations often exceeding 100 ng/ml. Patterns of testosterone secretion and musth varied among bulls with no evidence of seasonality (P>0.05). Only three bulls at one facility exhibited classic, well-defined yearly musth cycles. Others exhibited more irregular cycles, with musth symptoms often occurring more than once a year. A number of bulls (1 Asian, 9 African) had consistently low testosterone (<10 ng/ml) and never exhibited significant TGS or UD. At facilities with multiple bulls (n=3), testosterone concentrations were highest in the oldest, most dominant male. There were positive correlations between testosterone and cortisol for six of seven Asian and all three African males that exhibited musth (range, r=0.23-0.52; P<0.05), but no significant correlations for bulls that did not (P>0.05). For the three bulls that exhibited yearly musth cycles, TSH was positively correlated (range, r=0.22-0.28; P<0.05) and thyroid hormones (T3, T4) were negatively correlated (range, r=-0.25 to -0.47; P<0.05) to testosterone secretion. In the remaining bulls, there were no clear relationships between thyroid activity and musth status. Overall mean testosterone and cortisol concentrations increased with age for all bulls combined, whereas thyroid activity declined. In summary, a number of bulls did not exhibit musth despite being of adequate physical maturity. Cortisol and testosterone were correlated in most bulls exhibiting musth, indicating a possible role for the adrenal gland in modulating or facilitating downstream responses. Data were generally inconclusive as to a role for thyroid hormones in male reproduction, but the finding of discrete patterns in bulls showing clear testosterone cycles suggests they may facilitate expression or control of musth in some individuals

Brown, J.L., Somerville, M., Riddle, H.S., Keele, M., Duer, C.K., Freeman, E.W., 2007. Comparative endocrinology of testicular, adrenal and thyroid function in captive Asian and African elephant bulls. General and Comparative Endocrinology 151, 153-162.
Abstract: Concentrations of serum testosterone, cortisol, thyroxine (free and total T4), triiodothyronine (free and total T3) and thyroid stimulating hormone (TSH) were measured to assess adrenal and thyroid function as they relate to testicular activity and musth in captive elephants. Blood samples were collected approximately weekly from Asian (n = 8) and African (n = 12) bulls at seven facilities for periods of 4 months to 9.5 years. Age ranges at study onset were 8-50 years for Asian and 10-21 years for African elephants. Based on keeper logs, seven Asian and three African bulls exhibited behavioral and/or physical (temporal gland secretion, TGS, or urine dribbling, UD) signs of musth, which lasted 2.8 +/- 2.5 months in duration. Serum testosterone was elevated during musth, with concentrations often exceeding 100 ng/ml. Patterns of testosterone secretion and musth varied among bulls with no evidence of seasonality (P > 0.05). Only three bulls at one facility exhibited classic, well-defined yearly musth cycles. Others exhibited more irregular cycles, with musth symptoms often occurring more than once a year. A number of bulls (I Asian, 9 African) had consistently low testosterone (< 10 ng/ml) and never exhibited significant TGS or UD. At facilities with multiple bulls (n = 3), testosterone concentrations were highest in the oldest, most dominant male. There were positive correlations between testosterone and cortisol for six of seven Asian and all three African males that exhibited musth (range, r = 0.23-0.52; P < 0.05), but no significant correlations for bulls that did not (P > 0.05). For the three bulls that exhibited yearly musth cycles, TSH was positively correlated (range, r = 0.22-0.28; P < 0.05) and thyroid hormones (T3, T4) were negatively correlated (range, r = -0.25 to -0.47; P < 0.05) to testosterone secretion. In the remaining bulls, there were no clear relationships between thyroid activity and musth status. Overall mean testosterone and cortisol concentrations increased with age for all bulls combined, whereas thyroid activity declined. In summary, a number of bulls did not exhibit musth despite being of adequate physical maturity. Cortisol and testosterone were correlated in most bulls exhibiting musth, indicating a possible role for the adrenal gland in modulating or facilitating downstream responses. Data were generally inconclusive as to a role for thyroid hormones in male reproduction, but the finding of discrete patterns in bulls showing clear testosterone cycles suggests they may facilitate expression or control of musth in some individuals.

Dehnhard, M., 2007. Characterisation of the sympathetic nervous system of Asian (Elephas maximus) and African (Loxodonta africana) elephants based on urinary catecholamine analyses. Gen. Comp Endocrinol. 151, 274-284.
Abstract: Assessing the welfare status of captive animals using non-invasive measurements of hormones is of growing interest because this can serve as an effective tool to facilitate the optimization of environmental and husbandry conditions. Both the African elephant (Loxodonta africana) and the Asian elephant (Elephas maximus) exhibit extremely low breeding success in captivity, and because elevated levels of stress may negatively influence reproductive functions, this study sought to establish a method for assessing sympathoadrenal activity in captive female elephants. We found a circadian variation in urinary noradrenaline (norepinephrine, NE), adrenaline (epinephrine, Epi) and dopamine (DA) under short day length. Peak activity of noradrenaline and dopamine was noted at 3 a.m. Adrenaline showed a biphasic pattern with a minor peak recorded at 3 a.m. and a major peak 9 a.m. Under long-day photoperiodic conditions, simultaneous peaks of noradrenaline and adrenaline were again noted at 3 a.m. whereas dopamine does not appear to have a distinct circadian pattern under long-day length. A transfer of two elephant cows resulted in a marked increase in urinary adrenaline and noradrenaline levels, confirming that the transfer represented a stressful event. During the peripartal period, noradrenaline concentrations increased and maximum concentrations were obtained at delivery. Daily measurements of urinary dopamine throughout the follicular phase revealed an increase in dopamine secretion close to ovulation. This increase might indicate a role of dopamine in the ovulatory mechanisms. These results suggest that changes in urinary catecholamine excretion reflect fluctuations in sympathoadrenal activity and may be a useful indicator of stress

Duer, C., Carden, M., Tomasi, T., 2007. Detection of fetal gender differences in maternal serum progesterone concentrations of Asian elephants (Elephas maximus)
486. Anim Reprod. Sci. 97, 278-283.
Abstract: Previous studies have analyzed total testosterone concentrations in maternal serum for a reliable method of fetal gender determination in Asian elephants (Elephas maximus). The present study investigated the possibility that progesterone concentrations in maternal serum may reflect these testosterone patterns. Weekly serum samples were collected from 17 pregnancies in captive Asian elephants and analyzed via radioimmunoassay (RIA) for progesterone concentrations. Nine and eight cows carried male and female calves, respectively. Mean progesterone concentrations in maternal serum of elephants carrying male calves were greater than in those carrying female calves (P<0.01). Mean progesterone concentrations (based on 5-week means) in maternal serum were greater at weeks 20-55 (P<0.01) and 60-65 (P<0.05) for elephants carrying male calves

Hildebrandt, T., Drews, B., Gaeth, A.P., Goeritz, F., Hermes, R., Schmitt, D., Gray, C., Rich, P., Streich, W.J., Short, R.V., Renfree, M.B., 2007. Foetal age determination and development in elephants. Proc. Biol. Sci. 274, 323-331.
Abstract: Elephants have the longest pregnancy of all mammals, with an average gestation of around 660 days, so their embryonic and foetal development have always been of special interest. Hitherto, it has only been possible to estimate foetal ages from theoretical calculations based on foetal mass. The recent development of sophisticated ultrasound procedures for elephants has now made it possible to monitor the growth and development of foetuses of known gestational age conceived in captivity from natural matings or artificial insemination. We have studied the early stages of pregnancy in 10 captive Asian and 9 African elephants by transrectal ultrasound. Measurements of foetal crown-rump lengths have provided the first accurate growth curves, which differ significantly from the previous theoretical estimates based on the cube root of foetal mass. We have used these to age 22 African elephant foetuses collected during culling operations. Pregnancy can be first recognized ultrasonographically by day 50, the presumptive yolk sac by about day 75 and the zonary placenta by about day 85. The trunk is first recognizable by days 85-90 and is distinct by day 104, while the first heartbeats are evident from around day 80. By combining ultrasonography and morphology, we have been able to produce the first reliable criteria for estimating gestational age and ontological development of Asian and African elephant foetuses during the first third of gestation.

Hollister-Smith, J.A., Poole, J.H., Archie, E.A., Vance, E.A., Georgiadis, N.J., Moss, C.J., Alberts, S.C., 2007. Age, musth and paternity success in wild maleAfrican elephants, Loxodonta africana. Animal Behaviour 74, 287-296.
Abstract: Male African elephants experience intense intrasexual selection in gaining access to oestrous females, who represent a very scarce and highly mobile resource. An unusual combination of behavioural and physiologica ltraits in males probably reflects this intense selection pressure. Males show prolonged growth, growing throughout much or perhaps all of their long life span (ca. 60-65 years), and they show musth,a physiological and behavioural condition exclusive to elephants, which is manifested by bouts of elevated testosterone and aggression and heightened sexual activity. Most observed matings are by males over 35years of age and in musth, suggesting that age and musth are both important factors contributing to male reproductive success. Here we report the results of a genetic paternity analysis of a well-studied population of wild African elephants. Patterns of paternity for 119 calves born over a 22-year period showed significant effects of both age and musth on paternity success. Among males in musth, paternity success increased significantly with age until the very oldest age classes, when it modestly declined. When not inmusth, males experienced relatively constant, low levels of paternity success at all ages. Thus, despite the importance of both musth and age in determining male paternity success, adult males both in and out ofmusth, and of all ages, produced calves. In general, however, older males had markedly elevated paternitysuccess compared with younger males, suggesting the possibility of sexual selection for longevity in this species.

Kirkpatrick, J.F., 2007. Measuring the effects of wildlife contraception: the argument for comparing apples with oranges. Reprod. Fertil. Dev. 19, 548-552.
Abstract: There are few wildlife populations existing today that can be supported without some form of management. Wildlife fertility control, as one option, has moved from the research stage to actual application with a number of species, including wild horses, urban deer, captive exotic species and even African elephants, but this approach remains controversial in many quarters. Strident debate has arisen over the possible effects of contraception on behaviour, genetics, stress and even management economics, among other parameters. Part of the debate arises from the fact that critics often fail to recognise that some form of alternative management will be applied, and a second problem arises when critics fail to identify and demand the same concern for the consequences of the alternative management approaches. Thus, any rational debate on the merits or possible effects of contraceptive management of wildlife must also recognise all alternative management approaches and apply the same concern and questions to these alternative approaches--including 'no management'--as are currently being applied to fertility control. Only then will the stewards of wildlife be in a position to make wise and informed decisions about management options

Kusuda, S., Wakimoto, T., Nishimura, K., Kawakami, S., Okuda, K., Saito, E., Shimado, T., Sakamoto, H., Yanagimoto, H., Wada, S., Nishio, K., Fuji, H., Suzuki, T., Hashikawa, H., Kusunoki, H., Doi, O., 2007. Relationship between body temperature and ovarian cycle in Asian and African elephants. J Reprod Dev 53, 1099-1105.
Abstract: The aim of the present study was to investigate whether changes in body temperature are related to the ovarian cycle in elephants. Rectal, tongue or fecal temperature was measured for 2 Asian and 5 African elephants using an electric thermometer. Evaluation of ovarian cycles was based on the changes in serum or fecal progestin. The mean  SD values of the rectal, tongue, and fecal temperatures were 36.3  0.3 (2 Asian), 36.2  0.5 (1 African) and 36.5 0.3 C (4 African), respectively; the fecal temperature was the highest of the 3 temperatures (p<0.01). The longitudinal changes in body temperatures correlated with the ovarian cycle, with higher temperatures occurring during the luteal phase. The fecal temperatures of one acyclic African elephant did not change cyclically. These results suggest that measurement of body temperature can be used to easily evaluate the ovarian cyclicity of an individual animal, although
it might not be able to determine the ovarian cycle length.

Portas, T., Bryant, B., Goritz, F., Hermes, R., Keeley, T., Evans, G., Maxwell, M., Hildebrand, T., 2007. Semen collection in an Asian elephant (Elephas maximus) under combined physical and chemical restraint. Aust Vet J 85, 425-427.

Smit, I.P.J., Grant, C.C., Whyte, I.J., 2007. Landscape-scale sexual segregation in the dry season distribution and resource utilization of elephants in Kruger National Park, South Africa. Diversity And Distributions 13, 225-236.
Abstract:  This study compared the dry season distribution of elephant bull groups and mixed herds and the resources driving these distributions within Kruger National Park, South Africa. It is important to understand what resources drive the distribution of elephants as this may be of relevance to understanding and managing their impact. It is also important to distinguish between resource use by bull groups and mixed herds because their impact on the habitat may differ. Our results indicated that sexual segregation, both in space and in resource selection, did occur in Kruger Park. Bulls roamed more widely in the park, and although their distribution and resource use overlapped with mixed herds, they also occurred in areas that mixed herds apparently did not, or could not, utilize in the dry season. This gave rise to areas used exclusively by bulls but no areas used exclusively by mixed herds. Lower collective feeding requirements as a result of smaller group size, wider habitat tolerance, and increased mobility as a result of bigger body size, as well as conflict avoidance with musth bulls in areas with mixed herds, might have been some of the reasons for bull groups roaming more widely and for the establishment of separate bull areas. Rivers were an important resource driving both the distribution of the mixed herds and bull groups, but with the mixed herds occurring closer to these resource hot spots than the bull groups. Tree cover proved important for mixed herds, probably because of the importance of shade and the higher nutritional requirements of the smaller-sized cows and calves. Artificial waterholes might have opened up previously unutilized areas for bulls in the dry season, especially on the grassy basaltic plains in the north of the park. However, the distribution of the mixed herds suggested that they did not occur in higher densities in areas surrounding waterholes.

Wittemyer, G., Ganswindt, A., Hodges, K., 2007. The impact of ecological variability on the reproductive endocrinology of wild female African elephants. Hormones and Behavior 51, 346-354.
Abstract: Non-invasive endocrine methods enable investigation of the relationship between ecological variation and ovarian activity and how this impacts on demographic processes. The underlying physiological factors driving high variation in inter-calving intervals among multi-parous African elephants offer an interesting system for such an investigation. This study investigates the relationship between Normalized Differential Vegetation Index (NDVI), an ecosystem surrogate measure of primary productivity, and fecal progestin concentrations among wild female elephants. Matched fecal samples and behavioral data on reproductive activity were collected from 37 focal individuals during the two-year study. Linear mixed models were used to explore the relationship between fecal 5 alpha-pregnane-3-ol-20-one concentrations and the independent variables of NDVI, calf sex, female age, gestation day, and time since last parturition. Among both non-pregnant and pregnant females, fecal 5 alpha-pregnane-3-ol-20-one concentrations were significantly correlated with time-specific NDVI indicating a strong relationship between ecological conditions and endocrine activity regulating reproduction. In addition, the age of a female and time since her last parturition impacted hormone concentrations. These results indicate that the identification of an individual's reproductive status from a single hormone sample is possible, but difficult to achieve in practice since numerous independent factors, particularly season, impact fecal hormone concentrations. Regardless of season, however, fecal 5 alpha-pregnane-3-ol-20-one concentrations below 1 mu g/g were exclusively collected from non-pregnant females, which could be used as a threshold value to identify non-pregnant individuals. Collectively the information generated contributes to a better understanding of environmental regulation of reproductive endocrinology in wild elephant populations, information salient to the management and manipulation of population dynamics in this species.

Wittemyer, G., Rasmussen, H.B., Douglas-Hamilton, I., 2007. Breeding phenology in relation to NDVI variability in free-ranging African elephant. Ecography 30, 42-50.
Abstract: The phenology of reproduction is often correlated with resource availability and is hypothesized to be shaped by selective forces in order to maximize lifetime reproductive success. African elephants have the distinctive life history traits of a 22 month gestation and extended offspring investment, necessitating a long-term strategy of energy acquisition and reproductive expenditure to ensure successful offspring recruitment. We investigated the relationship between the reproductive phenology of a wild elephant population and resource availability using remotely sensed Normalized Differential Vegetation Index (NDVI) data as a measure of time-specific primary productivity and hence forage quality. The initiation of female elephants' 3+yr reproductive bout was dependent on conditions during the season of conception but timed so parturition occurred during the most likely periods of high primary productivity 22 months later. Thus, the probability of conception is linked to the stochastic variation in seasonal quality and the phenology of parturition is related to the predictable seasonality of primary productivity, indicating elephants integrate information on known current and expected future conditions when reproducing. Juvenile mortality was not correlated with ecological variability, hence female fecundity rather than calf mortality appears to drive demographic processes in the study population. Extreme climatic events, such as those associated with the El Niño-Southern-Oscillation (ENSO), acted to synchronize female fecundity in the population. This study suggests that the relationship between fecundity and ecological variability instigates the characteristic demographic fluctuations in elephant populations, rather than the mortality-driven fluctuations observed in many ungulate populations.

Wittemyer, G., Ganswindt, A., Hodges, K., 2007. The impact of ecological variability on the reproductive endocrinology of wild female African elephants. Horm. Behav. 51, 346-354.
Abstract: Non-invasive endocrine methods enable investigation of the relationship between ecological variation and ovarian activity and how this impacts on demographic processes. The underlying physiological factors driving high variation in inter-calving intervals among multi-parous African elephants offer an interesting system for such an investigation. This study investigates the relationship between Normalized Differential Vegetation Index (NDVI), an ecosystem surrogate measure of primary productivity, and fecal progestin concentrations among wild female elephants. Matched fecal samples and behavioral data on reproductive activity were collected from 37 focal individuals during the two-year study. Linear mixed models were used to explore the relationship between fecal 5alpha-pregnane-3-ol-20-one concentrations and the independent variables of NDVI, calf sex, female age, gestation day, and time since last parturition. Among both non-pregnant and pregnant females, fecal 5alpha-pregnane-3-ol-20-one concentrations were significantly correlated with time-specific NDVI indicating a strong relationship between ecological conditions and endocrine activity regulating reproduction. In addition, the age of a female and time since her last parturition impacted hormone concentrations. These results indicate that the identification of an individual's reproductive status from a single hormone sample is possible, but difficult to achieve in practice since numerous independent factors, particularly season, impact fecal hormone concentrations. Regardless of season, however, fecal 5alpha-pregnane-3-ol-20-one concentrations below 1 microg/g were exclusively collected from non-pregnant females, which could be used as a threshold value to identify non-pregnant individuals. Collectively the information generated contributes to a better understanding of environmental regulation of reproductive endocrinology in wild elephant populations, information salient to the management and manipulation of population dynamics in this species

Witter, K., Egger, G.F., Boeck, P., 2007. Renaut bodies in nerves of the trunk of the African elephant, Loxodonta africana. J. Morphol. 268, 414-422.
Abstract: Renaut bodies are loosely textured, cell-sparse structures in the subperineurial space of peripheral nerves, frequently found at sites of nerve entrapment. The trunk of the elephant is a mobile, richly innervated organ, which serves for food gathering, object grasping and as a tactile organ. These functions of the trunk lead to distortion and mechanical compression of its nerves, which can therefore be expected to contain numerous Renaut bodies. Samples of the trunk wall of an adult African elephant (Loxodonta africana) were examined histologically using conventional staining methods, immunohistochemistry, and lectin histochemistry. Architecture of nerve plexuses and occurrence of Renaut bodies in the elephant trunk were compared with those in tissues surrounding the nasal vestibule of the pig. Prominent nerve plexuses were found in all layers of the elephant trunk. Almost all (81%) nerve profiles contained Renaut bodies, a basophilic, discrete subperineurial layer resembling cushions around the nerve core. In contrast, Renaut bodies were seen in only 15% of nerve profiles in the porcine nasal vestibule. Within Renaut bodies, fusiform fibroblasts and round, ruff-like cells were placed into a matrix of acidic glycosaminoglycans with delicate collagen and very few reticular fibers. The turgor of this matrix is thought to protect nerves against compression and shearing strain. Renaut bodies are readily stained with alcian blue (pH 2.5) favorably in combination with immunohistochemical markers of nerve fibers. They should be regarded as a physiological response to repeated mechanical insults and are distinct from pathological alterations. alterations

Yon, L., Kanchanapangka, S., Chaiyabutr, N., Stanczyk, F., Meepan, S., Lasley, B., 2007. ACTH stimulation in four Asian bull elephants (Elephas maximus): an investigation of androgen sources in bull elephants. Gen. Comp Endocrinol. 151, 246-251.
Abstract: The phenomenon of musth is a very stressful event, both behaviorally and physiologically. An ACTH stimulation test was conducted in four adult Asian bull elephants to investigate the possibility that the classical hypothalamic-pituitary-adrenal (HPA) axis is active during musth, resulting in an increase in adrenally produced steroids. Serum cortisol, testosterone (T), androstenedione (A4), androstenediol (A5), and dehydroepiandrosterone (DHEA) were measured. Cortisol increased 3-10 times above baseline in response to ACTH stimulation, and DHEA doubled. A4 and A5 were erratic, while testosterone decreased significantly in all bulls. The pattern of results suggests that the adrenal steroid increase which occurs during musth results from some mechanism other than the classical HPA axis

Yon, L., Chen, J., Moran, P., Lasley, B., 2007. An analysis of the androgens of musth in the Asian bull elephant (Elephas maximus). Gen Comp Endocrinol. Mar 24; [Epub ahead of print].
Abstract: During musth in bull elephants, the androgens testosterone (T), dihydrotestosterone (DHT), and androstenedione all increase significantly. Given the unusual endocrine physiology that has been discovered in female elephants, it is also possible that bull elephants produce some unusual androgens. A cell-based androgen receptor assay was used to explore this possibility using two different methods. The first method compared the level of T measured by radioimmunoassay (RIA) with the level of androgen receptor (AR) activity measured in the serum of eight bull elephants during musth and non-musth periods. A ratio was calculated for T/AR activity for non-musth and musth, to determine if there was a change in the ratio between these two states. The second method used HPLC to separate two pooled serum samples (one non-musth and one musth) into fractions using a protocol which separates known androgens into specific, previously identified fractions. Each fraction was then tested with the AR assay to determine the androgenicity of any compounds present. This was done to determine if there were any fractions which had androgenic activity but did not contain any previously identified androgens. Results from the first analysis indicated no change in the T/AR ratio between non-musth and musth states. Clearly whatever active androgens are present during musth, they increase proportionately with T. Findings from the second analysis suggested that the only bioactive androgen present in the serum of non-musth Asian bulls is a low level of T. During musth, the only bioactive androgens detected were T and DHT; of these, T was by far the predominant active androgen present. Taken together, these two analyses suggest that T is by far the predominant active androgen present during musth in Asian bull elephants, and that no previously unidentified bioactive androgen is present.

Yon, L., Kanchanapangka, S., Chaiyabutr, N., Meepan, S., Stanczyk, F.Z., Dahl, N., Lasley, B., 2007. A longitudinal study of LH, gonadal and adrenal steroids in four intact Asian bull elephants (Elephas maximus) and one castrate African bull (Loxodonta africana) during musth and non-musth periods. Gen. Comp Endocrinol. 151, 241-245.
Abstract: During their annual musth cycle, adult African and Asian bull elephants have increased gonadal androgens (testosterone [T], dihydrotestosterone [DHT], androstenedione [A4]). Because musth is a physiologically and psychologically stressful time, this study was conducted to investigate whether the adrenal glands (stimulated by stress) increase production of both glucocorticoids and androgens during musth. Weekly serum samples were taken for 11-15 months from four intact adult Asian bull elephants, and from a castrate African bull elephant who exhibits musth. Testosterone, androstenediol (A5), A4, luteinizing hormone (LH), cortisol, and dehydroepiandrosterone (DHEA) were measured in each sample. In three of the four intact bulls, all hormones measured increased during musth. Adrenal androgens were strongly correlated with LH and testicular androgens, though not to cortisol. None of the hormones measured in the castrate bull increased during his musth cycles. While the significance of adrenal activity in the elephant during musth has yet to be determined, this study provides evidence that the adrenal gland actively produces both glucocorticoids and androgens during musth in the Asian elephant

Allen, W.R., 2006. Ovulation, pregnancy, placentation and husbandry in the African elephant (Loxodonta africana)
470. Philos. Trans. R. Soc. Lond B Biol. Sci. 361, 821-834.
Abstract: The African elephant reproduces so efficiently in the wild that overpopulation is now a serious problem in some game parks in Zimbabwe, Botswana and South Africa. The female reaches puberty between 10 and 12 years of age in the wild and, when in captivity, shows oestrous cycles of 14-15 weeks duration. She readily conceives a singleton in the wild yet her uterus has the capacity for twins. She shows a gestation length of 22 months and, in the wild, shows a population density and feed dependent intercalving interval of 4-8 years. The trophoblast erodes the lumenal epithelium of the endometrium and stimulates upgrowths of blood vessel-containing stromal villi, which develop eventually into the broad, tightly folded lamellae of the zonary, endotheliochorial placenta. Significant quantities of leaked maternal erythrocytes and ferric iron are phagocytosed by specialized trophoblast cells in the haemophagous zones at the lateral edges of the placental band. Although the placenta itself is endocrinologically inert, the foetal gonads, which enlarge greatly during the second half of pregnancy can synthesize 5alpha-dihydryoprogesterone and other 5alpha pregnane derivatives from cholesterol and pregnenolone. These products may synergize with progestagens secreted by the 2-8 large corpora lutea which are always present in the maternal ovaries throughout gestation to maintain the pregnancy state

Bagley, K.R., Goodwin, T.E., Rasmussen, L.E.L., Schulte, B.A., 2006. Male African elephants, Loxodonta africana, can distinguish oestrous status via urinary signals. Animal Behaviour 71, 1445.
Abstract: African elephants are a polygynous species that raise offspring in a matriarchal society. Unlike females, males disperse, spend time in mate groups and search for mates when mature. Urinary chemical signals aid males in detecting reproductively active females. A preovulatory pheromone has been identified in Asian elephants, Elephas maximus, but has not yet been experimentally identified in African elephants. In this study, the goal was to determine whether adult captive male African elephants can distinguish between urine from conspecific females in luteal and periovulatory oestrous stages as an indication that a preovulatory pheromone is released in the urine. Urine was collected from seven different female African elephants during their luteal and periovulatory periods of oestrus. Bioassays were conducted with nine adult male elephants housed at six different facilities. Males were presented with the two urine types and a control sample once a day over 3 days to reduce sample novelty, which can result in misleadingly high responses. All mates showed greater chemosensory responses to the periovulatory urine by trial 3 with the ability to distinguish the urines increasing over the 3 days. This is the first experimental behavioural evidence that African elephants release an oestrous pheromone in the urine. The ability of the captive male elephants to discern between the two urine types bolsters the hypothesis that there is a preovulatory pheromone in African elephants and encourages efforts to identify it.

Ball, R., Fad, O. Serum cortisols in captive Asian elephants (Elephas maximus) in different management systems at Busch Gardens Tampa Bay.  2006 Proceedings American Association of Zoo Veterinarians.  177-180. 2006.
Ref Type: Conference Proceeding
Abstract: Introduction:Cortisol is a widely accepted measure of stress in wild and captive animals.  In the past, captive elephant management systems have been criticized as potential stress inducers. The analysis of fecal cortisols is non-invasive and has been used to give long term evalutions of social and ecologic pressures in elephants and other species.  Salivary cortisols have also been used as a minimally invasive technique to measure social stress in captive elephants. The herd of Asian elephants at Busch Gardens Tampa Bay (BGT) changed from a traditional contact managemnt (free ccontact, FC) to a protected contact (PC) system utilizing positive-reinforcement based operant conditioning in 2004.  Serum cortisols were measured after the change and evaluated along wth banked samples from before. Long term sampling will be utilized to measure this transition but evaluating a single process will hopefully reflect the overall changes that can be expected with this change in management. While the individual variations are notable and other issues potentially confound the issue, it appears that this transition has lowered the serum cortisols in this herd.  In addition to serum cortisol measurements, the actual process of collecting the samples appears to be less stressful behaviorally. Pathologic processes should not be discounted when considering cortisol levels in evalauting stress in captive elephants.

Methods and Materials: Six female Asian elephants (Studbook numbers 30, 32, 304, 34, 35, 3) had been managed in a free contact system for many years.  Studbook number 304 was captive born and the others were wild born. Serum was collected intermittently during this management system to bank and for reproductive hormone analysis.  The elephants were placed in lateral recumbency by the handlers and blood collected from the ear vein on the caudal aspect of the down ear.  Reproductively sound animals were bled more frequently than the others.  Serum was frozen at -80°C until analyzed.  In August 2004, the first group of three animals was moved to the new barn and started the new positive-reinforcement, PC management system.  Within 5 wk, all animals had been moved over. All animals had been trunkwashed and were culture negative for Mycobacterium tuberculosis and negative on the newly developed MultiAntigen Print ImmunoAssay (MAPIA) and lateral-flow technology (Rapid Test) developed to detected antigen to M. tuberculosis.  As the caudal aspect of the ear was used for sampling, each elephant was asked to station in a static chute designed to allow training of voluntary ear-presentation for manipulation and blood collection. Handler safety and creating an effective learning environment for the elephants required training each to proceed to the chute solo and station there calmly. General desensitization techniques were applied as session durations were increased. Within the chute,individual elephants had significant room to maneuver. Since no physical restraint or sedation was utilized,animals were trained to cooperate fully and voluntarily allowing for blood sampling and other husbandry procedures. By May 2005, training for voluntary bloods draws was firmly established on all six animals.  The first approximately 20 samples collected under this new system were matched against the samples collected in the previous system.  Samples were selected against if the animal had an active problem or was on therapy for any reason.  Several animals had undergone a drug trial and these samples were selected against as well.  Serum was again stored in -80°C freezer until analyzed at Conservation and Research Center (CRC) Endocrine Research Laboratory, Smithsonian Institution, National Zoological Park, Front Royal, VA.  T-tests were utilized to discern any statistically significant results in the mean serum cortisols collected from animals before and after the implementation of the new husbandry systems.  Results were considered significant at alpha levels <0.05.

Results: The results and simple means of serum cortisols are listed in Table 1. Elephant No. 34 had essentially the same level of cortisol in both systems.  Elephant No. 32 had a reduction in the mean cortisol level of approximately 32% (20.84 versus 14.28 ng/ml) from the FC to the PC system.  Elephant No. 304 had a similar reduction of 37% in the mean cortisol (22.59 versus 14.29 ng/ml).  Statistical analyses results are reported here (means, standard deviations, t-test results).

Discussion: Serum was chosen over salivary and fecal sampling as a means to measure cortisol for several reasons. While fecal and salivary cortisol changes can reflect stresses within a reasonable period after the stressor (approximately 24 hr), serum cortisols is more likely to be reflective of the stressors closer to the moment of sampling.  The methodology is straightforward and less subject to the hazards for sample storage.  Timeliness of the sample result is also a benefit to serum sampling.  Blood sampling is a required husbandry practice in all elephant holding facilities belonging to the American Zoo and Aquarium Association (AZA).  While fecal cortisol samples may be useful to look at over a long term period to evaluate the transition from FC to PC, we choose to additionally look at how one specific task, blood collection, was affected by making this transition.  Fecal cortisols have been used to measure stress in transportation and environmental stress in some species, but are not thought to be reflective of the stress in a diagnostic procedure itself.  For this evaluation, the lag time period between the potential stressor (blood collection) and the means to measure the stressor are same.  Elephants No. 304 and 32 both had significant reductions in the mean serum cortisol levels.  Both are in good health and had no apparent inflammatory problems.  The logical deduction here is that the sampling process itself is less stressful in the PC management than the FC management.  Elephant 34 and 30 had essentially the same level of serum cortisol as measured by the mean in the different management systems.  Elephant 34 has developed significant uterine leiomyomas during the time period measured.  Elephant 30 has recently had clinical bouts of anterior enteritis and is suspected of having a dietary hypersensitivity to wheat.  Even with these two pathologic processes, the serum cortisol did not rise.  Elevations in cortisol are quite often explained as resulting from social, behavioral, or environmental causes and little attention is paid to inflammatory causes.  Associations between infections and elevated cortisols  have been noted in wild animals.  It is reasonable to assume that if these two processes did not exist, these levels would indeed be lower. Based on the other two elephants, a reduction of approximately 30% could be expected. Overall it appears that collecting blood from the elephants at BGT in the PC system is less stressful that the FC system.  As this is an example of how the routine husbandry and medical husbandry is now conducted, it can be expected that the overall net effect is going to be lowered stress in the elephants at BGT. ……………………………………………………………………………

Ball, R.L., Brown, J. Preliminary results of a cabergoline trial in captive elephant with hyperprolactinemia.  2006 Proceedings American Association of Zoo Veterinarians.  174-176. 2006.
Ref Type: Conference Proceeding
Abstract: Introduction: An Asian elephant (Elephas maximus) at Busch Gardens Tampa Bay (BGT) was diagnosed with hyperprolactinemia, with a persistently elevated serum prolactin concentration greater than 15 ng/ml, by the Conservation & Research Center (CRC) laboratory in January 1996.  She also had a number of other problems, including uterine disorders that resulted in consistently elevated progesterone. In March 2002, she was given cabergoline orally at a dose of 1 mg twice weekly p.o. for 6 mo.  Cabergoline is a long-acting dopamine receptor agonist with a high affinity for D2 receptors.  It exerts a direct inhibitory effect on the secretion of prolactin.  Cabergoline (Dostinex®, Pfizer Inc. Kalamazoo, Michigan 49007 USA) was purchased from a local pharmacy.  Serum prolactin concentrations declined almost immediately after treatment initiation, followed about 1 mo later by a drop in progesterone to baseline.  Progesterone secretion remained low until November 2002 when she resumed cycling based on the observation of a normal luteal phase based on serum progesterone profile.  From November 2002 through January 2004 she exhibited four normal estrous cycles.  Prolactin secretion also remained within the normal range for elephants, over 1 yr after treatment withdrawal. This female suffered no adverse effects due to the cabergoline treatment.  There were no behavioral changes noted or changes in appetite. Given the need to increase reproductive rates of African elephants (Loxodonta africana) to prevent captive extinction, it might be efficacious to treat genetically valuable females with cabergoline in the hope it will reinitiate reproductive cyclicity. Nearly 1/3 of African elephants with hormone data are not cycling normally, and in an earlier study 1/3 of these (11 of 30) were found to have increased serum prolactin levels.
 
Methods and Materials:
A clinical trial was undertaken with six captive African elephant females that were identified as good candidates for a cabergoline treatment study (i.e., they are acylic and had mean prolactin concentrations of >15 ng/ml).  The treatment consisted of 1 mg cabergoline given twice weekly p.o. for 6 mo.  Serum was banked and then analyzed at the CRC for progesterone and prolactin. All elephants were thought to be otherwise healthy.  Because prolactin is known to be an inflammatory marker, all candidates were required to have a negative lateral flow immunochromatograpy (Rapid Test) and multiple antigen immunoassay (MAPIA) for Mycobacteria tuberculosis.

Results:
A summary of the results is given in Table 1.  The treatment period is complete for three elephants, all of which showed a decrease in prolactin levels.  Elephant 1 showed a good response while on treatment, but did not cycle and serum prolactin has subsequently risen to pretreatment levels.  Increasing the dose in Elephant 2 and 3 reduced prolactin to baseline levels, but again did not result in a return to ovarian cyclicity.  Elephant 4 was taken off the study after only a few doses due to increased aggressive behaviors.  This is believed to be due to changes in the group social dynamics and not related to the cabergoline, as this behavior has continued after withdrawal of the drug. Based on these findings, the two newest candidates, Elephant 5 and 6, with very high prolactin concentrations have been placed on 2 mg/twice weekly for 1 yr pending continuation of this project.

Discussion:
Normalization of prolactin levels facilitated the return of normal cycles in an Asian elephant, but none of the African elephants have resumed cycling so far.  Thus, while the use of cabergoline shows promise in reducing elevated prolactin levels in both Asian and African elephants, other factors may need to be considered or a longer course at higher doses may be required for treatment to be successful in reinitiating ovarian activity.  The latter suggestion is supported by two of the animals (Elephants 2 and 3) in this limited trial, in which a decline in prolactin occurred after the dose was increased.  Understanding the etiology of hyperprolactinemia in elephants may also help in returning females to normal cycling.Relapse of hyperprolactinemia is more common in humans with micro- or macroprolactinomas.  Chronic estrogen stimulation is also known to increase prolactin levels.  A proposed pathophysiology is that elevated estrogen levels from persistent cycling will lead to elevated prolactin levels and acyclicity.  A difference between the two species in the causes of and potential treatment options for hyperprolactinemia should also be evaluated more closely.

1Busch Gardens Tampa Bay, 3605 Bougainvillea Drive, Tampa, FL 33674 USA;2Smithsonian Institution, National Zoological Park, Conservation & Research Center, Front Royal, VA 22630 USA
ACKNOWLEDGMENTS
We would like to thank the participating zoos for their cooperation and patience during this trial.
LITERATURE CITED
1Brown, J.L., S.L. Walker and T. Moeller. 2004.  Comparative endocrinology of cycling and noncycling Asian (Elephas maximus) and African (Loxodonta africana) elephants.  Gen. Comp. Endocrinol. 136:360-370.
2 Colao, A., A. Di Sarno, P. Cappabianca, C. Di Somma, R. Pivonello, and G. Lombardi. 2003. Withdrawal of long-term cabergoline therapy for tumoral and non-tumoral hyperprolactinemia.  New Engl. J. Med. 349:20232033.
3 Ismail, M.S., G.I. Serour, U. Torsten, H. Weitzel, and H.P. Berlien. 1998. Elevated serum prolactin level with high-dose estrogen contraceptive pills. .Eur. J. Contracept. Reprod. Health Care. 3(1):45-50.
4 Montero, A.M., O.A. Bottasso, M.R.Luraghi, A.G. Giovannoni, and L. Sen. 2001. Association between high serum prolactin and concomitant infections in HIV-infected patients. Human Immunol.62: 191-196.
5 Lyashchenko, K., M. Miller, and W.R. Waters. 2005. Application of multiple antigen print immunoassay and rapid lateral flow technology for tuberculosis testing of elephants. .  Proc. Am. Assoc. Zoo Vet. Annu. Meet. Pp. 64-65

Bertschinger, H., Delsink, A., Kirkpatrick, J.F., Human, A., Grobler, D., van Altena, J.J. Management of elephant populations in private South African game reserves with porcine zona pellucida vaccine.  2006 Proceedings American Association of Zoo Veterinarians.  283-285. 2006.
Ref Type: Conference Proceeding
Abstract: Control of African elephant populations has become an absolute necessity in a number of game reserves in southern Africa.  The two main methods used to control populations so far are culling and translocation. Culling, besides being regarded as inhumane and unacceptable in many quarters, is not suitable for smaller populations.  It requires that whole family units are culled simultaneously which could mean that in reserves with 10 to 50 elephants a considerable portion, if not the entire population, is killed.  As far as translocation is concerned, limited new space is available for elephants. The only alternative to the two above options is to control the rate of reproduction. The porcine zona pellucida (pZP) vaccine has been used to successfully contracept wild horses and other wildlife species.  Work on the contraception of African elephants was initiated in the Kruger National Park in 1995 when the potential for using the porcine zona pellucida (pZP) was investigated. Subsequently the first field trials on wild elephants were carried out in Kruger and the results clearly showed that elephants could be contracepted with the pZP vaccine, although the efficacy achieved was 80%. During these field trials safety and reversibility werecould be demonstrated. In 2000 an elephant contraceptive program was initiated at Makalali Private Game Reserve, RSA, which has become the flagship model for immunocontrol in African elephants. The preliminary findings have been reported in three publications.During the first year, all 18 cows that were individually identified and older than 12 yr of age were treated.  During the next 4 yr the number of cows contracepted increased to 23 as young animals were added to the program. The standard vaccination procedure during the first year consisted of a primary vaccination (600 μg or 400 μg pZP with 0.5 ml Freund's modified complete adjuvant) followed by boosters (200 μg pZP with 0.5 ml Freund's incomplete adjuvant) at 3 to 6-wk intervals. Annual boosters to maintain antibody titers and contraceptive effect followed.  To date, the success rate on cows that have passed reserve-specific intercalving period of 56 mo has been 100%. The population stabilized within 3 yr by which time when all cows that had been pregnant at the time of first vaccination in 2000 had calved. Once again safety during pregnancy (14 cows pregnant at 2-21 mo gestation when first treated gave birth to normal healthy calves) as well as side effects that were limited to occasional lumps at the site of vaccination could be shown. Following ground darting, behavioral patterns returned to pre-darting status within 2 days. During 2003 and 2004 most boosters were administered from a helicopter; whereas, previously they had been done from a vehicle or on foot.  In all cases, drop-out darts were used. Time taken for vaccination from helicopter take-off to landing was about 30 min (1.5 min per cow; 30 min for total time). This required prior knowledge of the locations of family units or that an individual in each unit is radio-collared. Herds settled down much more quickly (1-2 days) than if darted from the ground. Since then we have vaccinated another 107 elephant cows in eight game reserves.  The cow populations have ranged from 4 to 43. In one of the reserves, Mabula, RSA, two of the four cows vaccinated have passed the mean intercalving intervals of the reserve with neither of them producing a calf. Treatment at the remaining reserves was initiated in 2004 or 2005 and it is too early to evaluate results.  The most difficult reserve in terms of the vaccination process was Welgevonden, RSA, (35 000 ha) with 43 cows.  The reserve is mountainous and heavily wooded. None of the elephants were collared and individuals could not be easily identified on the day of primary vaccination.  The total flying time during which individuals were identified and vaccinated was 4.5 hr.  Administration of the first booster took about 2 hr to locate and vaccinate each cow. Between the first and second booster the first rains occurred, followed by the spring flush of the vegetation. By the time the second booster was attempted late in November, the trees all had foliage. Only half the cows were located and darted because the elephants were very difficult to spot under the tree canopies.  The valuable lessons we learned from this were: 1) that helicopter vaccinations should be performed when most trees are bare, and 2) when larger populations are vaccinated repeatedly during the first year, one cow in each family unit should be radio-collared. This makes rapid location of each unit possible and cuts down on the major cost factor that is flying time. Elephant behavior is being monitored in all eight reserves where contraception is being applied. Because most of them have been contracepted recently, only the data from Makalali is available. The elephants at Makalali have been monitored intensively almost on a daily basis. To date, no anomalies in terms of aggressive or indifferent behavior with regards to nursing time, nursing behavior and calf proximity have been noted. No change in the cows' social hierarchy has been noted. Since January 2003, a total of 15 heats were observed in 10 cows (nine in 2003 and six in 2004) with four mating episodes. For the same period, 38 musth occasions were seen in five bulls (26 in 2003 and 12 in 2004). These occasions include musth displayed in the same bull during consecutive days or within the same musth cycle. The greatest occurrence of musth was recorded in the largest, dominant bull. Bulls were not observed harassing or separating cows off from their herds or calves as a result of increased estrous frequency. Thus, the Makalali program demonstrates that pZP does not cause herd fragmentation, harassment by bulls, change in rank and other negative behaviors normally associated with hormonal contraceptives. In conclusion we feel that it is important to emphasize the following points: The pZP vaccine can be used successfully to contracept African elephants The vaccine is safe during pregnancy and has no negative effect on birth or calf raising It has no side effects other than occasional swelling at the site of vaccination It is reversible Other than an increased incidence of heat no behavioral side effects were seen.

Brown, J.L., Somerville, M., Riddle, H.S., Keele, M., Duer, C. Comparative endocrinology of testicular and thyroid function in captive Asian and African elephant bulls. Proceedings International Elephant Conservation & Research Symposium.  58-75. 2006. 2006.
Ref Type: Conference Proceeding

Brown, J.L., Freeman, E., Duce, C. Update on the reproductive status of female Asian and African elephants in the SSP population of North America. Proceedings International Elephant Conservation & Research Symposium.  48-57. 2006. 2006.
Ref Type: Conference Proceeding

Cooper, D.W., Larsen, E., 2006. Immunocontraception of mammalian wildlife: Ecological and immunogenetic issues. Reproduction 132, 821-828.
Abstract: Immunocontraception involves stimulating immune responses against gametes or reproductive hormones thus preventing conception. The method is being developed for the humane control of pest and overabundant populations of mammalian wildlife. This paper examines three fundamental issues associated with its use: (1) the difficulties of obtaining responses to self-antigens, (2) the likely evolution of genetically based non-response to immunocontraceptive agents, and (3) the possible changes in the array of pathogens possessed by the target species after generations of immunocontraception. Our review of the literature demonstrates that the barriers to an effective immunocontraceptive are at present very basic. Should they be overcome, the effects of immunocontraception on the immunogenetic constitution of wildlife populations through the selection for nonresponders must be examined. We suggest that the attempt to use the animal's own immune system to modulate reproduction may be incompatible with the basic biological function of protection against infectious disease. Research programs on mammalian immunocontraception should involve measurement of the heritability of non-response and an assessment of the likely change in the response of the contracepted population to possible pathogens.

Delsink, A.K., van Alten, J.J., Grobler, D., Bertschinger, H., Kirkpatrick, J., Slotow, R., 2006. Regulation of a small, discrete African elephant population through immunocontraception in the Makalali Conservancy, Limpopo, South Africa. South African Journal of Science 102, 403-405.
Abstract: Populations of the African elephant, Loxodonta africana, are growing rapidly in southern Africa, to the extent that population control has become essential. The management option of translocation is no longer realistically available, whilst culling has become ethically unacceptable, especially to the general public. Previous immunocontraception trials on elephants with Porcine Zona Pellucida (PZP) vaccine demonstrated that it is safe, effective, reversible, remotely deliverable, and has had no evident adverse side effects. We demonstrate effective contraceptive management of a discrete, small population of free-roaming elephants in the Makalali Conservancy, Limpopo province, South Africa. Complete reproductive control has been demonstrated in all 18 original targeted females, who have by now passed the population's average intercalving interval of 56 months without giving birth. A zero population growth rate has been maintained within this target group since August 2002. On the basis of this small sample over a short period, immunocontraception should be considered a viable means of population management as an alternative to long-term culling strategies in small populations

Drews, B., Göritz, F., Hermes, R., Streich, J.W., Rich, P., Schmitt, D., Lung, N., Renfree, M.B., Gaeth, A.P., Short, R.V., Hildebrandt, T.B. Morphological and ultrasonographic characterization of the embryonic development in elephants. Proceedings International Elephant Conservation & Research Symposium.  82-83. 2006. 2006.
Ref Type: Conference Proceeding

Druce, H., Pretorius, K., Druce, D., Slotow, R., 2006. The effect of mature elephant bull introductions on resident bull's group size and musth periods: Phinda Private Game Reserve, South Africa. South African Journal of Wildlife Research 36, 133-137.
Abstract: African elephants have been reintroduced into small, enclosed reserves in South Africa,many populations being established with orphans <10 years old.This has resulted in abnormal behaviour in some elephant populations, which was corrected in Pilanesberg National Park by introducing older bulls and culling certain problem elephants.In July 2003, three older bulls (29-41 years old) were introduced into Phinda Private Game Reserve, KwaZulu-Natal, South Africa, in order to normalize the bull age structure and in an attempt to reduce the abnormally long musth period of one particular resident bull. These introduced bulls were monitored intensively after release, as was the resident bull population, both before and after introduction of the older bulls.The introduced bulls all came into musth within eleven months postrelease.The older bulls do not appear to have had any influence on the musth periods of the oldest resident bull (36 years old at introduction). Detailed behavioural studies of the effects of management actions on elephant populations, within small, enclosed reserves provide information and resources for future management decisions.This study demonstrates that old bulls can be successfully introduced to very small areas provided that electrification of the entire perimeter is secure. Further, the introduction has no detectable medium-term (one year) effect on the behaviour of a relatively dense population of resident elephants, and the welfare of the elephants was not greatly affected.

Freeman, E.W., Brown, J.L., Whyte, I. Reproductive success of elephants in Kruger National Park, South Africa (1976-1995). Proceedings International Elephant Conservation & Research Symposium.  87-91. 2006. 2006.
Ref Type: Conference Proceeding

Hildebrandt, T., Goeritz, F., Hermes, R., Schaftenaar, W., Drews, B. Dystocia - an increasing problem in captive breeding programs: Causes and treatment. Proceedings International Elephant Conservation & Research Symposium.  92. 2006. 2006.
Ref Type: Conference Proceeding

Larke, A., Crews, D.E., 2006. Parental investment, late reproduction, and increased reserve capacity are associated with longevity in humans. J Physiol Anthropol 25, 119-131.
Abstract: Throughout the living world trade-offs between reproductive success and longevity have been observed. In general, two extremes of life history patterning are reported, r- and K-selected species. The latter tend toward larger body sizes, few offspring from any one pregnancy, few offspring over the female reproductive span, longer life spans, and greater parental investment (PI: all efforts and expenses associated with the production, gestation, post-natal care, feeding, and protection of young) (e.g., whales, elephants, hominids). r-selected species tend toward smaller body size, multiple births/litters per pregnancy, female production of many gametes and offspring over the life span, and low levels of PI (e.g., most plants, insects, mice). These differences have significant influences on physiological variation among human populations.Across human samples, reproductive success (RS: the number of offspring successfully birthed and reared to reproductive age) has been reported to vary positively, negatively, and not at all with longevity of women. This complexity may be in part due to the fact that both early-life and late-life fecundity are associated with longevity in women, while total parity seems a poor gauge of female longevity in humankind. Large variations in associations of RS with longevity in women suggest that multiple factors may confound this association. One confounding factor is that among women, RS is largely determined not by fecundity, but by the quality of PI available to offspring. Among modern humans, PI is more complex, longer lasting (both relatively and absolutely), and extensive than for any other ammal. This suggests that modern human life history is a reflection of the co-evolution of longevity and extensive PI as part of our species' biocultural evolution. The need for long-term PI has greatly shaped human physiological variation and patterns of longevity.

Oerke, A.-K., Heistermann, M.A., Hodges, K. Duration of pregnancy and its relation to sex of calf and age of cow in the European population of Asian and African elephants. Proceedings International Elephant Conservation & Research Symposium.  125-131. 2006. 2006.
Ref Type: Conference Proceeding

Riley, L.W., 2006. Of mice, men, and elephants: Mycobacterium tuberculosis cell envelope lipids and pathogenesis
454. J. Clin. Invest 116, 1475-1478.
Abstract: Mycolic acids and structures attached to them constitute a major part of the protective envelope of Mycobacterium tuberculosis, and for this reason, their role in tuberculosis pathogenesis has been extensively studied. In this issue of the JCI, Rao et al. examine the effect of trans-cyclopropanation of oxygenated mycolic acids attached to trehalose dimycolate (TDM) on the murine immune response to infection (see the related article beginning on page 1660). Surprisingly, they found that an M. tuberculosis mutant lacking trans-cyclopropane rings was hypervirulent in mice. The recent recognition of a hypervirulence phenotype in mice associated with laboratory and clinical M. tuberculosis strains with altered cell wall components has provided new insights into how M. tuberculosis may establish persistent infection. However, to date, characterization of these bioactive products in pathogenesis has been largely reductionistic; the relationship of their effects observed in mice to the persistent infection and tuberculosis caused by M. tuberculosis observed in humans remains obscure

Sa-Ardrit, M., Saikhun, J., Thongtip, N., Damyang, M., Mahasawangkul, S., Angkawanish, T., Jansittiwate, S., Faisaikarm, T., Kitiyanant, Y., Pavasuthipaisit, K., Pinyopummin, A., 2006. Ultrastructural alterations of frozen-thawed Asian elephant (Elephas maximus) spermatozoa
491. Int. J. Androl 29, 346-352.
Abstract: Intact plasma and acrosome membranes and functional mitochondria following cryopreservation are important attributes for the fertilizing ability of spermatozoa. In the present study, functional and ultrastructural changes of Asian elephant spermatozoa after cryopreservation either in TEST + glycerol or HEPT + dimethyl sulphoxide (DMSO) were evaluated by fluorescent techniques and electron microscopy. Sperm frozen in TEST + glycerol had higher proportion of sperm with intact plasma (49.1 +/- 9.2% vs. 30.9 +/- 3.9%) and acrosomal (53.7 +/- 4.9% vs. 35.8 +/- 6.1%) membranes, as well as active mitochondria (57.0 +/- 7.2% vs. 42.0 +/- 5.0%) than those cryopreserved in HEPT + DMSO. The results obtained from electron microscopy were similar to those obtained by fluorescence microscopy. The percentage of normal spermatozoa was higher when spermatozoa were frozen in TEST + glycerol than those frozen in HEPT + DMSO (31.8 +/- 5.6 vs. 28.5 +/- 6.4). The ultrastructural alterations revealed by transmission electron microscopy could be classified as (i) distension of plasma membrane, while the acrosome was swollen; (ii) disruption or loss of plasma membrane, while acrosome was swollen with distended outer acrosomal membrane; (iii) disruption or loss of plasma and outer acrosomal membrane with leakage of acrosome content; (iv) extensive vesiculation of plasma and outer acrosomal membrane and leakage of acrosome content; (v) a complete loss of both plasma membrane and outer acrosomal membrane; and (vi) swelling of mitochondria. These findings suggest that the freezing and thawing procedure caused structural damage to elephant spermatozoa, especially in the plasma membrane, acrosome and mitochondria. Fluorescence and electron microscopic evaluations are potentially a powerful tool in the analysis of elephant spermatozoa after freezing and thawing

Saikhun, J., Thongtip, N., Kornkaewrat, K., Mahasawangkul, S., Angkawanish, T., Boonprasert, K., Pinyopummin, A. Osmotic stress on motility and membrane integrity of Asian elephant spermatozoa analyzed by computer-assisted semen analysis. Proceedings International Elephant Conservation & Research Symposium.  163. 2006. 2006.
Ref Type: Conference Proceeding

Shannon, G., Page, B.R., Duffy, K.J., Slotow, R., 2006. The role of foraging behaviour in the sexual segregation of the African elephant
419. Oecologia. 150, 344-354.
Abstract: Elephants (Loxodonta africana) exhibit pronounced sexual dimorphism, and in this study we test the prediction that the differences in body size and sociality are significant enough to drive divergent foraging strategies and ultimately sexual segregation. Body size influences the foraging behaviour of herbivores through the differential scaling coefficients of metabolism and gut size, with larger bodied individuals being able to tolerate greater quantities of low-quality, fibrous vegetation, whilst having lower mass-specific energy requirements. We test two distinct theories: the scramble competition hypothesis (SCH) and the forage selection hypothesis (FSH). Comprehensive behavioural data were collected from the Pongola Game Reserve and the Phinda Private Game Reserve in South Africa over a 2.5-year period. The data were analysed using sex as the independent variable. Adult females targeted a wider range of species, adopted a more selective foraging approach and exhibited greater bite rates as predicted by the body size hypothesis and the increased demands of reproductive investment (lactation and pregnancy). Males had longer feeding bouts, displayed significantly more destructive behaviour (31% of observations, 11% for females) and ingested greater quantities of forage during each feeding bout. The independent ranging behaviour of adult males enables them to have longer foraging bouts as they experience fewer social constraints than females. The SCH was rejected as a cause of sexual segregation due to the relative abundance of low quality forage, and the fact that feeding heights were similar for both males and females. However, we conclude that the differences in the foraging strategies of the sexes are sufficient to cause spatial segregation as postulated by the FSH. Sexual dimorphism and the associated behavioural differences have important implications for the management and conservation of elephant and other dimorphic species, with the sexes effectively acting as distinct "ecological species"

Shannon, G., Page, B.R., Duffy, K.J., Slotow, R., 2006. The consequences of body size dimorphism: Are African elephants sexually segregated at the habitat scale? Behaviour 143, 1145-1168.
Abstract: Sexual segregation is a commonly observed phenomenon in dimorphic ungulates, which has been categorised into two distinct components: social segregation and habitat segregation. In this study we investigated whether elephants were sexually segregated at the habitat scale. The locations of 12 family groups and 16 males, in three distinct populations were recorded over a period of 2.5 years. Selection ratios were calculated for each habitat type and a Kendall's coefficient of concordance was used for the analyses. The habitat and foraging preferences were firstly tested for concordance within sex, and then between the sexes. Female habitat preferences showed significant concordance across all reserves and they also exhibited strong concordance in their summer foraging preferences. Their weakest association with habitat and foraging preference was during winter, which may be related to resource scarcity. Males exhibited significant concordance in their habitat preferences in two out of the three reserves. They had their weakest associations in the summer months and this may be linked to avoidance of other males in musth and the abundance of forage. There were no significant differences in habitat preference between males and females and it is likely that individual preferences vary as much within sex as between sexes. Differential habitat utilisation does not appear to be driving sexual segregation in elephants and it is postulated that sociality, divergent reproductive strategies and foraging behaviour at the plant scale play a more significant role. The results of this study highlight the importance of scale in elucidating the mechanisms involved in sexual segregation.

Shrader, A.M., McElveen, M.E., Lee, P.C., Moss, C.J., van Aarde, R.J., 2006. Growth and age determination of African savanna elephants. Journal of Zoology, London 270, 40-48.
Abstract: Understanding the population dynamics of savanna elephants depends on estimating population parameters such as the age at first reproduction, calving interval and age-specific survival rates. The generation of these parameters, however, relies on the ability to accurately determine the age of individuals, but a reliable age estimation technique for free-ranging elephants is presently not available. Shoulder heights of elephants were measured in 10 populations in five countries across southern and eastern Africa. Data included shoulder height measurements from two populations where the age of each individual was known (i.e. Addo Elephant National Park, South Africa and Amboseli National Park, Kenya). From the known-age data, Von Bertalanffy growth functions were constructed for both male and female elephants. Savanna elephants were found to attain similar asymptotic shoulder heights in the 10 populations, while individuals in the two known-age populations grew at the same rate. The Von Bertalanffy growth curves allowed for the accurate age estimation of females up to 15 years of age and males up to 36 years of age. The results indicate that shoulder height can serve as an indicator of chronological age for elephants below 15 years of age for females and 36 years of age for males. Ages derived from these growth curves can then be used to generate age-specific population variables, which will help assess the demographic status of savanna elephant populations across Africa.

Thitaram, C., Pongsopawijit, P., Thongtip, N., Angkavanich, T., Chansittivej, S., Wongkalasin, W., Somgird, C., Suwankong, N., Prachsilpchai, W., Suchit, K., Clausen, B., Boonthong, P., Nimtrakul, K., Niponkit, C., Siritepsongklod, S., Roongsri, R., Mahasavankul, S., 2006. Dystocia following prolonged retention of a dead fetus in an Asian elephant (Elephas maximus)
458. Theriogenology 66, 1284-1291.
Abstract: A 32-year-old nulliparous female Asian elephant (Elephas maximus) showed signs of parturition 8 months later than predicted from the breeding records. However, while serosanguineous fluid, necrotic tissue and pieces of amnion were expelled, second-stage labor did not progress. Since the fetus was not found during an endoscopic examination of the vestibule, it was assumed that the elephant had calved unseen and she was returned to the forest to recuperate. Twelve months later, the elephant showed clear signs of second-stage labor accompanied by a bulge in the perineum and passage of keratinized nail through the vulva. A 35 cm episiotomy incision was made in the perineum just below the anus, via which chains were attached to the forelimbs of the fetus. Traction on the forelimbs alone proved insufficient to achieve delivery because the fetal head kept rotating and impacting in the pelvis. However, traction applied via a hook inserted behind the mandibular symphysis allowed the head to be elevated and extended, and the fetus to be delivered. The episiotomy wound was sutured in two layers and although the skin did not heal during primary closure it subsequently healed uneventfully by second intention. Retrospective evaluation of the elephant's serum progestagens profile demonstrated a fall to baseline at the suspected onset of parturition, supporting the supposition that the fetus was retained in the uterus for 12 months after parturition began. It is suggested that serum progestagens concentrations should be monitored regularly in mated elephant cows to verify the establishment of pregnancy and to better estimate the expected timing, and the onset of calving

Thitaram, C., Thongtip, N., Somgird, C., Colenbrander, B., Van Boxtel, D.C.J., Lenstra, J.A. Molecular tool for genetic management and parentage test to control poaching in Asian elephants. Proceedings International Elephant Conservation & Research Symposium.  205-209. 2006.
Ref Type: Conference Proceeding

Thongtip, N., Mahasawangkul, S., Kornkaewrat, K., Saikhun, J., Angkwanish, T., Jansittiwate, S., Boonprasert, K., Wajjwalku, W., Songsasen, N., Pinyopummintr, T., Pinyopummin, A. Potential factors affecting Asian elephant semen quality in Thailand. Proceedings International Elephant Conservation & Research Symposium.  210. 2006. 2006.
Ref Type: Conference Proceeding

Thongtip, N., Mahasawangkul, S., Thitaram, C., Pongsopavijitr, P., Kornkaewrat, K., Saikhun, J., Angkawanish, T., Jansittiwate, S., Rungsri, R., Boonprasert, K., Somkird, C., Wongkalasinh, W., Wajjwalku, W., Songsasen, N., Pinyopummintr, T., Pinyopummin, A. First report of pregnancy by artificial insemination with chilled semen of an Asian elephant in Thailand. Proceedings International Elephant Conservation & Research Symposium.  273. 2006. 2006.
Ref Type: Conference Proceeding

Vinogradov, I.V., Kochneva, G.V., Shchelkunov, S.N., Riabchikova, E.I., 2006. [Reproduction of cowpox virus strain EP-2 isolated from an elephant in primary fibroblast cultures and chorion-allantoic chick embryos]
451. Vopr. Virusol. 51, 44-48.
Abstract: Electron microscopy was used to study the reproduction of cowpox virus strain EP-2 in the cells of a primary fibroblast cultures (PFC) and chorion-allantoic membrane (CAM) of chick embryos (CE). The sequential stages of viral morphogenesis and the structure of A-type inclusions were described. The parameters of viral reproduction in PFC and CE CAM were compared. The formation of crystalloid tubular structures in PFC, unusual electron dense inclusions in the cells of CE CAN, and different variants of A-type inclusions in the cells of a pock was found. The histological and ultrastructural characteristics of pocks in CE CAM are described

Allen, W.R., Mathias, S., Ford, M., 2005. Placentation in the African elephant, Loxodonta africana. IV. Growth and function of the fetal gonads
550. Reproduction. 130, 713-720.
Abstract: The gonads, both ovaries and testes, of 44 elephant fetuses weighing 0.09-112 kg (6.1-21.3 months gestation) were examined grossly and histologically. As in equids, elephant fetal gonads undergo a phase of marked growth and enlargement during the second half of gestation, which is more pronounced in ovaries than testes due to growth and antrum formation of numerous follicles in the former. Stromal cells undergo hypertrophy and transformation to form zones of interstitial cells that are associated with the enlarged follicles in the ovaries and in which the primitive seminiferous tubules are embedded in the testes. The interstitial cells have the capacity to synthesize 5alpha-dihydroprogesterone and other 5alpha-reduced progestagens from cholesterol and pregnenelone and the hypothesis is raised that these fetal gonadal progestagens may supplement significantly the progestagens secreted by the multiple large corpora lutea of pregnancy in the elephant

Andrews, J., Mecklenborg, A., Bercovitch, F.B., 2005. Milk intake and development in a newborn captive African elephant  (Loxodonta africana). Zoo Biology 24, 275-281.
Abstract: In August 2003 the San Diego Zoo's Wild Animal Park (WAP) and the Lowry Park Zoo, under the auspices of the AZA's Elephant SSP and a USFWS permit, imported 3.8 African elephants (Loxodonta africana ) from the Kingdom of Swaziland. When they were captured, transrectal ultrasound examinations revealed that one nulliparous cow was approximately 10 months pregnant.  At the time of their arrival (August 2003), all of the animals were estimated to be approximately 13 years old and were thought to be nulliparous.  Based on the ultrasound examination results and the average African elephant gestation period, parturition was predicted to be 20 February 2004. In this report, we provide the first detailed data about nursing activity around the clock and newborn calf development, describe maternal and neonatal nighttime activity budgets, and explore maternal weight changes during suckling and lactation. The newborn calf suckled significantly more at night than during the day, but suckled for only about 2 hr per 24-hr period. Regression analysis revealed that through the first 3 months of life the calf gained 0.385 kg/day while it suckled on a regular basis. We compare our findings with published information on wild elephants, and conclude that although the growth rate is reduced compared to hand-reared elephant calves, the suckling patterns are almost identical to those reported for wild calves.

Deem, S.L., Brown, J.L., Eggert, L., Wemmer, C., Htun, W., Nyunt, T., Murray, S., Leimgruber, P. Health and management of working elephants in Myanmar (Burma). Procedings American Association of Zoo Veterinarians.  228-231. 2005.
Ref Type: Conference Proceeding
Abstract: Myanmar has approximately 6,000 working elephants.  Remaining wild elephants are declining, partly because of live-capture for captivity.  Through health and reproductive assessments, genetic analyses and GPS tracking of captive and wild elephants, we are exploring linkages between the two populations and conducting studies to reduce morbidity and mortality of captive elephants. Captive elephants live and work in Myanmar's forests in close proximity and contact to the remaining wild herds. We propose that reducing morbidity and mortality in the captive elephants will decrease the need for live-capture, and the risk of disease transmission, to wild elephants.
Introduction
There are an estimated 6,000 working elephants in Myanmar - half owned by the government operated Myanmar Timber Enterprise (MTE) and half owned privately^.5 This may be one of the largest captive elephant populations in the world and its management will have a significant impact on remaining wild herds in Myanmar.^4,6,8  With mortality rates higher than birth rates, the working population is probably maintained by supplementing it with elephants captured from the wild.⁵ There is evidence that continued harvest of wild elephants may have reduced the remaining wild populations of Myanmar.  Recent surveys of wild populations in two of Myanmar's protected elephant ranges revealed extremely low dung counts, indicative of small and declining herds. Constant contact with captive elephants in Myanmar's forests may exacerbate the threat to Myanmar wild elephants by increasing the transmission of disease between these two groups. For both the above reasons, we believe that the conservation of wild elephants in Myanmar will require significant improvements in the care and management of currently existing captive populations.  
Elephants owned by MTE receive veterinary care from the Burmese veterinarians that work for the timber company and travel extensively throughout the country to sites were the elephants are located.¹ There is a dire need for veterinary supplies and laboratory capabilities in the country. Currently, veterinary practices are based on the extensive field experience of lead MTE veterinarians. However, MTE veterinarians frequently rely on older published work ^3,7 and would benefit significantly from training that incorporates new insights into elephant health and new veterinary techniques. Similarly, because of their close-up experience of elephant health problems in the forests, MTE veterinarians may be able to make important new contributions to the care and management of elephants elsewhere.     
The overall objective of our study is to work jointly with MTE veterinarians to develop long-term captive population management strategies to reduce mortality and increase births in the working timber elephants and stop the continued off-take of animals from the wild to supplement captive herds.
Methods
The health component of this study has five major objectives.  These are to:
1              Conduct a training workshop, in conjunction with MTE veterinarians, on elephant management and veterinary care. 
2              Develop protocols so that the MTE veterinarians can collect samples for reproductive, genetic, and health status assessments.
3              Analyze samples and provide data to MTE veterinarians to improve husbandry, preventive care and disease treatment of working elephants.
4              Develop a comprehensive bibliography of all published information on the health and management of Myanmar elephants.
5              Perform an epidemiologic evaluation of records available on the historic and current working elephant population.
Specific steps to achieve these objectives include: 
1              Determine causes and rates of morbidity and mortality of captive MTE elephants.
2              Determine causes of low rates of reproduction in captivity.
3              Develop a genetic profile of the captive herds.
4              Develop a protocol to assess oozies-Burmese mahout-expertise in parallel with endocrine and health assessments to determine quality of care and potentially related stress.
5              Develop small population viability models to assess how current mortality effects long-term survival of the captive population and what supplementation from the wild is needed for short- and long-term sustainability.
6              Use population viability models to demonstrate how supplementation from the wild will negatively affect that population.
7              Get baseline health parameter data on free-ranging elephants.
8              Quantify habitat/space use using GPS and satellite tracking of captive and wild elephants. 
Results and Discussion
During an initial exploratory visit in November 2004, we learned that the annual mortality rate for MTE working elephants was 2.4% (66) in 2003.  Deaths occurred in all age groups (>18 yr, n = 40; 4 - 17 yr, n = 11; <4 yr, n = 15) and included preventable diseases (i.e., poor nutrition, heat stroke, diarrhea, dystocia, infectious and parasitic agents).  Additionally, we collected samples for performing health, genetic and endocrine analyses of 22 elephants maintained in one of the working camps (results to be presented). A relationship also was established with the veterinary staff at the Yangon Zoo, including follow up donations of veterinary literature and journals to the zoo. We provided medical advice for the care of an orphaned elephant calf and other animals housed at the zoo during our brief visit. We are seeking funds for a training course to be conducted in late 2005 and hope to perform health evaluations on a larger number of zoo and working elephants during that visit.
The National Zoo already has an extensive conservation program for wild elephants in Myanmar.^4,6,8  This program has focused on assessing wild elephant populations in protected areas and satellite-tracking of four wild elephants to learn more about their conservation status and ecology in Myanmar.  Currently this work is being extended to a national elephant survey. Part of this work included collecting fecal samples for genetic and health assessments.
The Smithsonian team of researchers involved in this project includes a veterinarian, reproduction physiologist, geneticist, conservation biologist, and landscape ecologist.  All members of this multidisciplinary team have extensive experience working with elephants and together provide the necessary expertise to study and understand the numerous factors affecting Myanmar's captive elephants and the long-term survival of elephants in Myanmar.  These challenges range from human land use and elephant population fragmentation, human-elephant conflict, poor reproduction and health care of captive elephants and lack of information on the health status of the wild elephants.  A viable conservation initiative for the elephants of Myanmar requires that health issues be addressed as one component of a comprehensive program to address the anthropogenic pressures on both working and wild elephants.²
The elephants of Myanmar are an excellent example of the fine line that exists between captive and wild animals, especially as it relates to health.  Captive and wild elephants are regularly in direct and indirect contact.  The working elephants live with their oozies who may expose them to diseases, such as tuberculosis.  The working elephants in turn may encounter wild elephants at night in the forests where they forage and live during non-working hours. In fact, the majority of captive born calves are said to be sired by wild bulls.  Potentially, the use of working elephants in selectively extracting valuable timber provides new strategies for the conservation of elephants and forests. Most likely, "elephant-logging" is less damaging than machine-operated timbering projects that tend to clear-cut areas and also damage the soil and streams.  However, decreasing the negative impact of such practices (i.e., minimizing off-take of elephants from the wild, decreasing disease risks to the wild elephants) is imperative.  
LITERATURE CITED
1 Aung, T., and T. Nyunt.  2002.  The care and management of the domesticated Asian elephant in Myanmar.  In: Baker, I., and M. Kashio (eds.): Giants on our hands. Proc. Int. Workshop Domesticated Asian Elephant. Dharmasarn Co., Ltd. Bangkok, Thailand. Pp. 89 - 102.
2 Deem, S.L., W.B. Karesh, and W. Weisman.  2001.  Putting theory into practice: wildlife health in conservation.  Conserv. Biol. 5: 1224-1233.
3 Evans, G.H. 1910.  Elephants and Their Diseases.  Government Printing. Rangoon. 323 
4 Kelly, D.S. 2005.  Habitat selection in declining elephant populations of Alaungdaw Kathapa National Park. Masters Thesis.  George Mason University.
5 Lair, R.C. 1997.  Myanmar. In: Gone Astray: The Care and Management of the Asian Elephant in Domesticity. FAO Regional Office for Asia and the Pacific, Thailand.  RAP Publication. Pp. 99-131
6 Leimgruber, P., and C. Wemmer.  2004.  National elephant symposium and workshop. Report to the USFWS and the Myanmar Forest Department.
7 Pfaff, G. 1930.  Reports on Diseases of Elephants.  Government Printing. Rangoon. 91
8 Wemmer, C., P. Leimgruber and D. S. Kelly.  2005.  Managing wild elephants in Alaungdaw Kathapa National Park and Htamanthi Wildlife Sanctuary.  Report to the USFWS and the Myanmar Forest Department.

Delves, P.J., Roitt, I.M., 2005. Vaccines for the control of reproduction--status in mammals, and aspects of comparative interest
592. Dev. Biol. (Basel) 121, 265-273.
Abstract: The objective of producing vaccines which target elements of the reproductive system to control fertility has been pursued for many years. Of the many targets for such vaccines, several sperm-associated antigens have been proposed for antibody-mediated intervention before fertilization but the very abundance of antigen to be neutralized has been a barrier. Zona pellucida antigens associated with the surface of the oocyte have also been targeted and used successfully for control of 'wild' elephant populations but worries concerning immunopathologically-mediated tissue damage have been mooted. Vaccines using human chorionic gonadotropin (hCG) which is required for the implantation and maintenance of the fertilized egg, although of interest for the development of fertility control in human populations, has no relevance in the context of the present conference because external fertilization of fish eggs is independent. The pathways by which gonadotropin-releasing hormone (GnRH) secreted by the hypothalamus promote release of luteinizing (LH) and follicle-stimulating hormone (FSH) which govern the physiological maturation and maintenance of the reproductive organs, provide many targets for immunological intervention. Most consistent success has been reported using GnRH-based vaccines which are immunosterilizing in a variety of mammalian species such as pigs, rodents and white-tailed deer. The fact that the structure of the decapeptide, GnRH, has been maintained over so many years of evolution and been conserved across so many animal species, encourages the view that a strategy for control of sexual maturation in fish based upon stimulation of GnRH antibodies may well prove to be a practical proposition, provided the formulation of an appropriate highly immunogenic vaccine can be achieved

Ganswindt, A., Rasmssen, H.B., Heistermann, M., Hodges, J.K., 2005. The sexually active states of free-ranging male African elephants (Loxodonta africana): defining musth and non-musth using endocrinology, physical signals, and behavior. Horm Behav 47, 83-91.
Abstract: Musth in male African elephants, Loxodonta africana, is associated with increased aggressive behavior, continuous discharge of urine, copious secretions from the swollen temporal glands, and elevated androgen levels. During musth, bulls actively seek out and are preferred by estrous females although sexual activity is not restricted to the musth condition. The present study combines recently established methods of fecal hormone analysis with long-term observations on male-female associations as well as the presence and intensity of physical signals to provide a more detailed picture about the physical, physiological, and behavioral characteristics of different states of sexual activity in free-ranging African elephants. Based on quantitative shifts in individual bull association patterns, the presence of different physical signals, and significant differences in androgen levels, a total of three potential sub-categories for sexually active bulls could be established. The results demonstrate that elevations in androgen levels are only observed in sexually active animals showing temporal gland secretion and/or urine dribbling, but are not related to the age of the individual. Further, none of the sexually active states showed elevated glucocorticoid output indicating that musth does not represent an HPA-mediated stress condition. On the basis of these results, we suggest that the term "musth" should be exclusively used for the competitive state in sexually active male elephants and that the presence of urine dribbling should be the physical signal used for defining this state.

Ganswindt, A., Heistermann, M., Hodges, K., 2005. Physical, physiological, and behavioral correlates of musth in captive African elephants (Loxodonta africana)
594. Physiol Biochem. Zool. 78, 505-514.
Abstract: Although musth in male African elephants (Loxodonta africana) is known to be associated with increased aggressiveness, urine dribbling (UD), temporal gland secretion (TGS), and elevated androgens, the temporal relationship between these changes has not been examined. Here, we describe the pattern of musth-related characteristics in 14 captive elephant bulls by combining long-term observations of physical and behavioral changes with physiological data on testicular and adrenal function. The length of musth periods was highly variable but according to our data set not related to age. Our data also confirm that musth is associated with elevated androgens and, in this respect, show that TGS and UD are downstream effects of this elevation, with TGS responding earlier and to lower androgen levels than UD. Because the majority of musth periods were associated with a decrease in glucocorticoid levels, our data also indicate that musth does not represent a physiological stress mediated by the hypothalamic-pituitary-adrenal axis. Furthermore, we demonstrate that the occurrence of musth is associated with increased aggression and that this is presumably androgen mediated because aggressive males had higher androgen levels. Collectively, the information generated contributes to a better understanding of what characterizes and initiates musth in captive African elephants and provides a basis for further studies designed to examine in more detail the factors regulating the intensity and duration of musth

Ganswindt, A., Rasmussen, H.B., Heistermann, M., Hodges, J.K., 2005. The sexually active states of free-ranging male African elephants (Loxodonta africana): defining musth and non-musth using endocrinology, physical signals, and behavior
652. Horm. Behav. 47, 83-91.
Abstract: Musth in male African elephants, Loxodonta africana, is associated with increased aggressive behavior, continuous discharge of urine, copious secretions from the swollen temporal glands, and elevated androgen levels. During musth, bulls actively seek out and are preferred by estrous females although sexual activity is not restricted to the musth condition. The present study combines recently established methods of fecal hormone analysis with long-term observations on male-female associations as well as the presence and intensity of physical signals to provide a more detailed picture about the physical, physiological, and behavioral characteristics of different states of sexual activity in free-ranging African elephants. Based on quantitative shifts in individual bull association patterns, the presence of different physical signals, and significant differences in androgen levels, a total of three potential sub-categories for sexually active bulls could be established. The results demonstrate that elevations in androgen levels are only observed in sexually active animals showing temporal gland secretion and/or urine dribbling, but are not related to the age of the individual. Further, none of the sexually active states showed elevated glucocorticoid output indicating that musth does not represent an HPA-mediated stress condition. On the basis of these results, we suggest that the term "musth" should be exclusively used for the competitive state in sexually active male elephants and that the presence of urine dribbling should be the physical signal used for defining this state

Garstang, M., 2005. Long-distance, low-frequency elephant communication. J Comp Physiol A Neuroethol Sens Neural Behav Physiol. 191, 299.
Abstract: Erratum: J Comp Physiol A Neuroethol Sens Neural Behav Physiol. 2004; Oct;190(10):791-805. Epub 2004 Sep 2. The production, transmission, and reception of and the behavioral response to long-distance, low-frequency sound by elephants is reviewed. The structure of low-frequency calls generated by elephants is separated into the "source" and the "filter" roles played by the lungs, larynx and vocal track, the composition of the expired air and the ambient air temperature. Implications regarding the size, age, sex, sexual and physical status follow from the call structure and detection. Reception of the signal is discussed in terms of the characteristics of the elephant's ear with particular attention to the determination of the threshold of hearing and the ability to locate the source of low-frequency sounds. Factors which influence the transmission of near infrasound are related to atmospheric structure. The critical role played by the thermal stratification and vertical gradient and magnitude of the wind in determining both the range and the detection of a signal are discussed for open and closed elephant habitats. Infrasound plays a pervasive role in reproduction, resource utilization, avoidance of predation and other social interactions. Current and future technology can be expected to contribute to the detection and interpretation of elephant communication. This will aid in the understanding of behavior and in efforts to sustain the species.

Glickman, S.E., Short, R.V., Renfree, M.B., 2005. Sexual differentiation in three unconventional mammals: Spotted hyenas, elephants and tammar wallabies. Hormones and Behaviour 48, 403-417.
Abstract: The present review explores sexual differentiation in three non-conventional species: the spotted hyena, the elephant and the tammar wallaby, selected because of the natural challenges they present for contemporary understanding of sexual differentiation. According to the prevailing view of mammalian sexual differentiation, originally proposed by Alfred Jost, secretion of androgen and anti-Mullerian hormone (AMH) by the fetal testes during critical stages of development accounts for the full range of sexually dimorphic urogenital traits observed at birth. Jost's concept was subsequently expanded to encompass sexual differentiation of the brain and behavior. Although the central focus of this review involves urogenital development, we assume that the novel mechanisms described in this article have potentially significant implications for sexual differentiation of brain and behavior, a transposition with precedent in the history of this field. Contrary to the ''specific'' requirements of Jost's formulation, female spotted hyenas and elephants initially develop male-type external genitalia prior to gonadal differentiation. In addition, the administration of anti-androgens to pregnant female spotted hyenas does not prevent the formation of a scrotum, pseudoscrotum, penis or penile clitoris in the offspring of treated females, although it is not yet clear whether the creation of masculine genitalia involves other steroids or whether there is a genetic mechanism bypassing a hormonal mediator. Wallabies, where sexual differentiation occurs in the pouch after birth, provide the most conclusive evidence for direct genetic control of sexual dimorphism, with the scrotum developing only in males and the pouch and mammary glands only in females, before differentiation of the gonads. The development of the pouch and mammary gland in females and the scrotum in males is controlled by genes on the X chromosome. In keeping with the ''expanded'' version of Jost's formulation, secretion of androgens by the fetal testes provides the best current account of a broad array of sex differences in reproductive morphology and endocrinology of the spotted hyena, and androgens are essential for development of the prostate and penis of the wallaby. But the essential circulating androgen in the male wallaby is 5α androstanediol, locally converted in target tissues to DHT, while in the pregnant female hyena, androstenedione, secreted by the maternal ovary, is converted by the placenta to testosterone (and estradiol) and transferred to the developing fetus. Testicular testosterone certainly seems to be responsible for the behavioral phenomenon of musth in male elephants. Both spotted hyenas and elephants display matrilineal social organization, and, in both species, female genital morphology requires feminine cooperation for successful copulation. We conclude that not all aspects of sexual differentiation have been delegated to testicular hormones in these mammals. In addition, we suggest that research on urogenital development in these non-traditional species directs attention to processes that may well be operating during the sexual differentiation of morphology and behavior in more common laboratory mammals, albeit in less dramatic fashion.

Glickman, S.E., Short, R.V., Renfree, M.B., 2005. Sexual differentiation in three unconventional mammals: spotted hyenas, elephants and tammar wallabies
566. Horm. Behav. 48, 403-417.
Abstract: The present review explores sexual differentiation in three non-conventional species: the spotted hyena, the elephant and the tammar wallaby, selected because of the natural challenges they present for contemporary understanding of sexual differentiation. According to the prevailing view of mammalian sexual differentiation, originally proposed by Alfred Jost, secretion of androgen and anti-Mullerian hormone (AMH) by the fetal testes during critical stages of development accounts for the full range of sexually dimorphic urogenital traits observed at birth. Jost's concept was subsequently expanded to encompass sexual differentiation of the brain and behavior. Although the central focus of this review involves urogenital development, we assume that the novel mechanisms described in this article have potentially significant implications for sexual differentiation of brain and behavior, a transposition with precedent in the history of this field. Contrary to the "specific" requirements of Jost's formulation, female spotted hyenas and elephants initially develop male-type external genitalia prior to gonadal differentiation. In addition, the administration of anti-androgens to pregnant female spotted hyenas does not prevent the formation of a scrotum, pseudoscrotum, penis or penile clitoris in the offspring of treated females, although it is not yet clear whether the creation of masculine genitalia involves other steroids or whether there is a genetic mechanism bypassing a hormonal mediator. Wallabies, where sexual differentiation occurs in the pouch after birth, provide the most conclusive evidence for direct genetic control of sexual dimorphism, with the scrotum developing only in males and the pouch and mammary glands only in females, before differentiation of the gonads. The development of the pouch and mammary gland in females and the scrotum in males is controlled by genes on the X chromosome. In keeping with the "expanded" version of Jost's formulation, secretion of androgens by the fetal testes provides the best current account of a broad array of sex differences in reproductive morphology and endocrinology of the spotted hyena, and androgens are essential for development of the prostate and penis of the wallaby. But the essential circulating androgen in the male wallaby is 5alpha androstanediol, locally converted in target tissues to DHT, while in the pregnant female hyena, androstenedione, secreted by the maternal ovary, is converted by the placenta to testosterone (and estradiol) and transferred to the developing fetus. Testicular testosterone certainly seems to be responsible for the behavioral phenomenon of musth in male elephants. Both spotted hyenas and elephants display matrilineal social organization, and, in both species, female genital morphology requires feminine cooperation for successful copulation. We conclude that not all aspects of sexual differentiation have been delegated to testicular hormones in these mammals. In addition, we suggest that research on urogenital development in these non-traditional species directs attention to processes that may well be operating during the sexual differentiation of morphology and behavior in more common laboratory mammals, albeit in less dramatic fashion

Greenwood, D.R., Comeskey, D., Hunt, M.B., Rasmussen, L.E., 2005. Chemical communication: chirality in elephant pheromones
528. Nature 438, 1097-1098.
Abstract: Musth in male elephants is an annual period of heightened sexual activity and aggression that is linked to physical, sexual and social maturation. It is mediated by the release of chemical signals such as the pheromone frontalin, which exists in two chiral forms (molecular mirror images, or enantiomers). Here we show that enantiomers of frontalin are released by Asian elephants in a specific ratio that depends on the animal's age and stage of musth, and that different responses are elicited in male and female conspecifics when the ratio alters. This precise control of communication by molecular chirality offers insight into societal interactions in elephants, and may be useful in implementing new conservation protocols

Lacasse, C., Gamble, K.C., Terio, K., Farina, L.L., Travis, D.A., Miller, M. Mycobacterium szulgai osteoarthritis and pneumonia in an African elephant (Loxodonta Africana). 2005 Proceedings AAZV, AAWV, AZA Nutrition Advisory Group.  170-172. 2005.
Ref Type: Conference Proceeding
Abstract: Tuberculosis, particularly Mycobacterium bovis and M. tuberculosis, is an important health issue in zoological collections.  Zoos are a particular public health concern because of the close contact between tuberculosis-susceptible animals and humans, specifically animal handlers and visitors.¹⁶ Evidence of M. tuberculosis transmission between humans and elephants, confirmed by DNA fingerprinting, has been reported.¹³ Between 1994 and 2001, M. tuberculosis was isolated from trunk washes of captive elephants from 11 herds in the United States.¹⁷  To date, most reported cases of tuberculosis have occurred in captive Asian elephants (Elephas maximus).¹⁴ In 1997, the National Tuberculosis Working Group for Zoo and Wildlife Species partnered with the USDA to formulate the "Guidelines for the Control of Tuberculosis in Elephants." ¹⁵ This document outlines criteria for the testing, surveillance, and treatment of tuberculosis in elephants. The guidelines recommend annual monitoring of elephants by mycobacterial culture of three direct trunk washes collected over 1 wk.  Isolation of Mycobacterium avium and non-tuberculous mycobacteria from elephant trunk wash samples is common, but these organisms have not been associated with clinical disease.^14,18 This case report details clinical disease with fatal complications of an atypical mycobacterial infection in an African elephant (Loxodonta africana). In September 2003, an African elephant presented with acute, severe lameness of the left rear limb with subsequent swelling of the stifle.  Diagnostic procedures included aspiration cytology of the swelling, radiographs, and thermographic imaging.  The exact location of the injury could not be detected, but a lesion to the stifle or coxofemoral articulation was suspected.  After 13 mo of treatment, including pulse therapy with a variety of nonsteroidal anti-inflammatory drugs (NSAIDs), weekly to biweekly injections of polysulfated glycosaminoglycan, and intensive foot care efforts to treat secondary pedal lesions of both rearlimbs, the animal died acutely.  Gross necropsy revealed granulomatous osteomyelitis with necrosis/loss of the femoral head and acetabulum and pulmonary granulomas.  Both of these lesions contained acid-fast bacteria on cytology. While awaiting confirmatory culture results, quarantine procedures were established for the elephant facility and a program was established to screen all zoo personnel in close contact with the elephant or who participated in the necropsy.  All personnel were tested by the Chicago Department of Public Health without documented conversion. Mycobacterium szulgai was ultimately cultured from both coxofemoral and pulmonary lesions. Mycobacterium szulgai is an uncommon nontuberculous mycobacterium that is usually isolated from pathologic lesions in humans.²¹ This bacterial species was first identified in 1972.¹¹ The lungs are the main locality for pathologic manifestation in humans and several cases have been in patients with acquired immunodeficiency syndrome.^9,20,21 Infection due to M. szulgai most frequently produces thin-walled cavities in lungs resembling tuberculosis.⁴ Other documented sites of infection include the skin, bone, and tendon sheath (causing a carpal tunnel syndrome).^{2,9,10,12,19,20}  Intra-operative contamination from ice water has led to M. szulgai keratitis after laser-assisted ophthalmic surgeries^.6 A case of disseminated disease in a previously healthy young human has been reported.⁵  No evidence of human-to-human transmission of this organism has been documented and human cases are believed to originate from environmental sources.¹²  The natural habitat of the organism is unknown, but previous reports suggest an association of the bacteria with water of swimming pools and fish tanks.^1,21 The organism has been cultured from a snail and tropical fish.^1,3 No standard recommendation for the treatment of M. szulgai infection currently exists.  In general, triple antibiotic therapies used in standard mycobacterial treatments are reported with a low rate of relapses and sterilization of sputum cultures within a mean of 3 mo.³ Pulmonary lesions in this elephant were chronic; it was not possible to determine when initial infection occurred. Infection could have occurred in captivity or in the wild prior to captivity. Three trunk washes over the past year had been negative for mycobacterial culture. Osteomyelitis in the hip may have developed secondary to hematogenous spread from the lungs with the acute lameness resulting from a pathologic fracture associated with this infection. Alternatively, though considered less likely, a traumatic fracture of the hip could have occurred, with bacterial inoculation and secondary osteomyelitis as a result of increased blood flow to the site. The source of infection for this elephant remains unknown.  Prevalence of this organism in the natural habitat or captive environment of the elephants has not been previously documented.
LITERATURE CITED
1 Abalain-Colloc, M.L., D. Guillerm, M. Salaun, S. Gouriou, V. Vincent, and B. Picard.  2003.  Mycobacterium szulgai isolated from a patient, a tropical fish, and aquarium water.  Eur. J. Clin. Microbiol. Infect. Dis.  22: 768-769.
2.Cross, G.M., M. Guill, and J.K. Aton.  1985.  Cutaneous Mycobacterium szulgai infection. Arch. Dermatol. 121: 247-249.
3. Davidson, P.T. 1976. Mycobacterium szulgai: a new pathogen causing infection of the lung.  Chest 69: 799- 801.
4. Dylewski, J.S., H.M. Zackon, A.H. Latour, and G.R. Berry.  1987.  Mycobacterium szulgai: an unusual pathogen.  Rev. Infect. Dis.  9: 578-580.
5. Gur, H., S. Porat, H. Haas, Y. Naparstek, and M. Eliakim.  1984.  Disseminated mycobacterial disease caused by Mycobacterium szulgai. Arch. Intern. Med. 144: 1861-1863.
6.Holmes, G.P., G. Bond, R.C. Fader, and S.F. Fulcher.  2002. A cluster of cases of Mycobacterium szulgai keratitis that occurred after laser-assisted in situ keratomileusis.  Clin. Infect. Dis. 34: 1039-1046.
7.Horusitzky, A., X. Puechal, D. Dumont, T. Begue, M. Robineau, and M. Boissier.  2000.  Carpal tunnel syndrome caused by Mycobacterium szulgai. J. Rheumatol 27: 1299-1302.
8.Hurr, H., and T. Sorg.  1998.  Mycobacterium szulgai osteomyelitis.  J. Infect.  37: 191-192.
9.Luque, A.E., D. Kaminski, R. Reichman, and D. Hardy. 1998.  Mycobacterium szulgai osteomyelitis in an AIDS patient. Scand. J. Infect. Dis. 30: 88-91.
10.Maloney, J.M., C.R. Gregg, D.S. Stephens, F.A. Manian, and D. Rimland.  1987.  Infections caused by Mycobacterium szulgai in humans.  Rev. Infect. Dis.  9: 1120-1126.
11.Marks, J., P.A. Jenkins, and M. Tsukamura.  1972.  Mycobacterium szulgai: a new pathogen.  Tubercle 53: 210.
12.Merlet, C., S. Aberrane, F. Chilot, and J. Laroche.  2000.  Carpal tunnel syndrome complicating hand flexor tenosynovitis due to Mycobacterium szulgai. Joint Bone Spine 67: 247-248.
13.Michalak, K., C. Austin, S. Diesel, J.M. Bacon, P. Zimmerman, and J. N. Maslow.  1998. Mycobacterium tuberculosis infection as a zoonotic disease: transmission between humans and elephants. Emerg. Infect. Dis. 4: 283-287.
14.Mikota, S.K., R.S. Larsen, and R.J. Montali.  2000.  Tuberculosis in elephants in North America.  Zoo Biol. 19: 393-403.
15.National Tuberculosis Working Group for Zoo and Wildlife Species. 2000. Guidelines for the control of tuberculosis in elephants.  USDA Animal and Plant Health Inspection Services.
16.Oh, P., R. Granich, J. Scott, B. Sun, M. Joseph, C. Stringfield, S. Thisdell, J. Staley, D. Workman-Malcolm, L. Borenstein, E. Lehnkering, P. Ryan, J. Soukup, A. Nitta, and J. Flood.  2002.  Human exposure following  Mycobacterium tuberculosis infection of multiple animal species in a metropolitan zoo.  Emerg. Infect. Dis. 8: 1290-1293.
17.Payeur, J.B., J.L. Jarnagin, J.G. Marquardt, and D.L. Whipple.  2002.  Mycobacterial isolations in captive elephants in the United States.  Ann. N.Y. Acad. Sci. 969: 256-258.
18.Shojaei, H., J.G. Magee, R. Freeman, M. Yates, N.U. Horadagoda, and M. Goodfellow.  2000. Mycobacterium elephantis sp. nov., a rapidly growing non-chromogenic Mycobacterium isolated from an elephant.  Int. J. Syst. Evol. Microbiol.  50: 1817-1820.
19.Stratton, C.W., D.B. Phelps, and L.B. Reller.  1978.  Tuberculoid tenosynovitis and carpal tunnel syndrome caused by Mycobacterium szulgai.  Am. J. Med.  65: 349-351.
20.Tappe, D., P. Langmann, M. Zilly, H. Klinker, B. Schmausser, and M. Frosch.  2004.  Osteomyelitis and skin ulcers caused by Mycobacterium szulgai in an AIDS patient.  Scand. J. Infect. Dis. 36: 883-885.
21.Tortoli, E., G. Besozzi, C. Lacchini, V. Penati, M.T. Simonetti, and S. Emler.  1998.  Pulmonary infection due to Mycobacterium szulgai, case report and review of the literature.  Eur. Respir. J.  11: 975-977.

Leong, K.M., Burks, K., Rizkalla, C.E., Savage, A., 2005. Effects of reproductive and social context on vocal communication in captive female African elephants (Loxodonta africana). Zoo Biology 24, 331-347.
Abstract: Female African elephants advertise changes in reproductive condition to males through a variety of modalities, including an increase in low-frequency vocalizations, presumed to travel long distances.  Although males respond to these vocalizations, it has been suggested that their proximate function may be to signal to nearby females rather than to distant males. Because elephants live in a female-bonded society, it is likely  that  changes in female reproductive condition also affect close-range  interactions between high- and low-ranking females and that  vocalizations  may mediate these interactions. To examine female-female interactions related to vocal production and the ovulatory cycle, this year-long study monitored behavior, vocalizations and hormonal cycles for a group of six female captive African elephants at Disney's Animal Kingdom. Rates of several types of close-range interactions were observed to change over the phases of the estrous cycle, and rank seemed to affect whether or not low-frequency vocalizations were given in association with these interactions. Results of this study suggest that a female African  elephant's  immediate social context and rank in the social hierarchy interact with  the  hormonal cycle in the production of low-frequency vocalizations, thus  many  of these vocalizations may not function proximately as signals to  distant  males, but may be a result of the changing dynamics among females.

Mobasheri, A., Gent, T.C., Womack, M.D., Carter, S.D., Clegg, P.D., Barrett-Jolley, R., 2005. Quantitative analysis of voltage-gated potassium currents from primary equine (Equus caballus) and elephant (Loxodonta africana) articular chondrocytes
618. Am. J. Physiol Regul. Integr. Comp Physiol 289, R172-R180.
Abstract: In this comparative study, we have established in vitro models of equine and elephant articular chondrocytes, examined their basic morphology, and characterized the biophysical properties of their primary voltage-gated potassium channel (Kv) currents. Using whole cell patch-clamp electrophysiological recording from first-expansion and first-passage cells, we measured a maximum Kv conductance of 0.15 +/- 0.04 pS/pF (n = 10) in equine chondrocytes, whereas that in elephant chondrocytes was significantly larger (0.8 +/- 0.4 pS/pF, n = 4, P </= 0.05). Steady-state activation parameters of elephant chondrocytes (V = -22 +/- 6 mV, k = 11.8 +/- 3 mV, n = 4) were not significantly different from those of horse chondrocytes (V = -12.5 +/- 4.3 mV, k = 12 +/- 2, n = 10). This suggests that there would be slightly more resting Kv activation in elephant chondrocytes than in their equine counterparts. Kinetic analysis revealed that both horse and elephant chondrocyte Kv currents had similar activation and inactivation parameters. Pharmacological investigation of equine chondrocyte Kv currents showed them to be powerfully inhibited by the potassium channel blockers tetraethylammonium and 4-aminopyridine but not by dendrotoxin-I. Immunohistochemical studies using polyclonal antibodies to Kv1.1-Kv1.5 provided evidence for expression of Kv1.4 in equine chondrocytes. This is the first electrophysiological study of equine or elephant chondrocytes. The data support the notion that voltage-gated potassium channels play an important role in regulating the membrane potential of articular chondrocytes and will prove useful in future modeling of electromechanotransduction of fully differentiated articular chondrocytes in these and other species

Murwira, A., Skidmore, A.K., 2005. The response of elephants to the spatial heterogeneity of vegetation in a Southern African agricultural landscape. Landscape Ecology 20, 217-234.
Abstract: Based on the agricultural landscape of the Sebungwe in Zimbabwe, we investigated whether and how the spatial distribution of the African elephant (Loxodonta africana) responded to spatial heterogeneity of vegetation cover based on data of the early 1980s and early 1990s. We also investigated whether and how elephant distribution responded to changes in spatial heterogeneity between the early 1980s and early 1990s.  Vegetation cover was estimated from a normalised difference vegetation index (NDVI).  Spatial heterogeneity was estimated from a new approach based on the intensity (i.e., the maximum variance exhibited when a spatially distributed landscape property such as vegetation cover is measured with a successively increasing window size or scale) and dominant scale (i.e., the scale or window size at which the intensity is displayed). We used a variogram to quantify the dominant scale (i.e., range) and intensity (i.e., sill) of NDVI based congruent windows (i.e., 3.84 km x 3.84 km in a 61 km x 61 km landscape). The results indicated that elephants consistently responded to the dominant scale of spatial heterogeneity in a unimodal fashion with the peak elephant presence occurring in environments with dominant scales of spatial heterogeneity of around 457-734 m. Both the intensity and dominant scale of spatial heterogeneity predicted 65 and 68% of the variance in elephant presence in the early 1980s and in the early 1990s respectively.  Also, changes in the intensity and dominant scale of spatial heterogeneity predicted 61% of the variance in the change in elephant distribution. The results imply that management decisions must take into consideration the influence of the levels of spatial heterogeneity on elephants in order to ensure elephant persistence in agricultural landscapes.

Naz, R.K., Gupta, S.K., Gupta, J.C., Vyas, H.K., Talwar, A.G., 2005. Recent advances in contraceptive vaccine development: a mini-review
577. Hum. Reprod. 20, 3271-3283.
Abstract: Contraceptive vaccines (CV) may provide viable and valuable alternatives to the presently available methods of contraception. The molecules that are being explored for CV development either target gamete production [luteinizing hormone-releasing hormone (LHRH)/GnRH, FSH], gamete function [sperm antigens and oocyte zona pellucida (ZP)], and gamete outcome (HCG). CV targeting gamete production have shown varied degrees of efficacy; however, they either affect sex steroids causing impotency and/or show only a partial rather than a complete effect in inhibiting gametogenesis. However, vaccines based on LHRH/GnRH are being developed by several pharmaceutical companies as substitutes for castration of domestic pets, farm and wild animals, and for therapeutic anticancer purposes such as in prostatic hypertrophy and carcinoma. These vaccines may also find applications in clinical situations that require the inhibition of increased secretions of sex steroids, such as in uterine fibroids, polycystic ovary syndrome, endometriosis and precocious puberty. CV targeting molecules involved in gamete function such as sperm antigens and ZP proteins are exciting choices. Sperm constitute the most promising and exciting target for CV. Several sperm-specific antigens have been delineated in several laboratories and are being actively explored for CV development. Studies are focused on delineating appropriate sperm-specific epitopes, and increasing the immunogenicity (specifically in the local genital tract) and efficacy on the vaccines. Anti-sperm antibody (ASA)-mediated immunoinfertility provides a naturally occurring model to indicate how a vaccine might work in humans. Vaccines based on ZP proteins are quite efficacious in producing contraceptive effects, but may induce oophoritis, affecting sex steroids. They are being successfully tested to control feral populations of dogs, deer, horses and elephants, and populations of several species of zoo animals. The current research for human applicability is focused on delineating infertility-related epitopes (B-cell epitopes) from oophoritis-inducing epitopes (T-cell epitopes). Vaccines targeting gamete outcome primarily focus on the HCG molecule. The HCG vaccine is the first vaccine to undergo Phase I and II clinical trials in humans. Both efficacy and lack of immunopathology have been reasonably well demonstrated for this vaccine. At the present time, studies are focused on increasing the immunogenicity and efficacy of the birth control vaccine, and examining its clinical applications in various HCG-producing cancers. The present article will focus on the current status of the anti-sperm, anti-ZP, anti-LHRH/GnRH and anti-HCG vaccines

Ortolani, A., Leong, K., Graham, L., Savage, A., 2005. Behavioral indices of estrus in a group of captive African elephants (Loxodonta africana). Zoo Biology 24, 311-329.
Abstract: This study investigated behavioral signals of estrus by systematically monitoring the interactions of one male with four female African elephants housed in a naturalistic outdoor enclosure at Disney's Animal Kingdom over a period of 11 months. We measured changes in five spatial behaviors and 22 tactile-contact behaviors, as well as changes in serum progestagen and LH concentrations, across three ovarian cycles for each female. Two females did not cycle during the study. Three different phases of the ovarian cycle were identified: mid luteal, anovulatory follicular, ovulatory follicular.  The male followed more and carried out more genital inspections, flehmen, and trunk-to-mouth behaviors toward cycling females during their ovulatory phase. Genital inspections by the male peaked above baseline levels on  the  day of an LH surge, and up to 9 days before, in both cycling females  and,  thus, might be a useful behavioral index of estrus. The male also carried out more genital inspections, flehmen, and trunk touches to the back leg toward ovulatory cycling than noncycling females. Overall, our results  indicated that: 1) a single subadult African elephant male could  discriminate two females in the ovulatory phase of their cycle (i.e.,  during  the 3 weeks preceding ovulation) from the mid luteal phase; 2) the male  also  discriminated two cycling females in the ovulatory and anovulatory  follicular phases from two noncycling females; 3) two females in the  ovulatory phase of the cycle displayed a greater variety of  tactile-contact  behavior toward the male compared to the other cycle phases.

Perez-Barberia, F.J., Gordon, I.J., 2005. Gregariousness increases brain size in ungulates
586. Oecologia. 145, 41-52.
Abstract: The brain's main function is to organise the physiological and behavioural responses to environmental and social challenges in order to keep the organism alive. Here, we studied the effects that gregariousness (as a measurement of sociality), dietary habits, gestation length and sex have on brain size of extant ungulates. The analysis controlled for the effects of phylogeny and for random variability implicit in the data set. We tested the following groups of hypotheses: (1) Social brain hypothesis-gregarious species are more likely to have larger brains than non-gregarious species because the former are subjected to demanding and complex social interactions; (2) Ecological hypothesis-dietary habits impose challenging cognitive tasks associated with finding and manipulating food (foraging strategy); (3) Developmental hypotheses (a) energy strategy: selection for larger brains operates, primarily, on maternal metabolic turnover (i.e. gestation length) in relation to food quality because the majority of the brain's growth takes place in utero, and finally (b) sex hypothesis: females are expected to have larger brains than males, relative to body size, because of the differential growth rates of the soma and brain between the sexes. We found that, after adjusting for body mass, gregariousness and gestation length explained most of the variation in brain mass across the ungulate species studied. Larger species had larger brains; gregarious species and those with longer gestation lengths, relative to body mass, had larger brains than non-gregarious species and those with shorter gestation lengths. The effect of diet was negligible and subrogated by gestation length, and sex had no significant effect on brain size. The ultimate cause that could have triggered the co-evolution between gestation length and brain size remains unclear

Poole, J.H., Tyack, P.L., Stoeger-Horwath, A.S., Watwood, S., 2005. Animal behaviour: elephants are capable of vocal learning
623. Nature 434, 455-456.
Abstract: There are a few mammalian species that can modify their vocalizations in response to auditory experience--for example, some marine mammals use vocal imitation for reproductive advertisement, as birds sometimes do. Here we describe two examples of vocal imitation by African savannah elephants, Loxodonta africana, a terrestrial mammal that lives in a complex fission-fusion society. Our findings favour a role for vocal imitation that has already been proposed for primates, birds, bats and marine mammals: it is a useful form of acoustic communication that helps to maintain individual-specific bonds within changing social groupings

Rasmussen, L.E.L., Krishamurthy, V., Sakumar, R., 2005. Behavioural and chemical confirmation of the preovulatory pheromone, (Z)-7-dodecenyl acetate, in wild Asian elephants: its relationship to musth. Behaviour 142, 351-396.
Abstract: Mammalian breeding strategies vary depending on particular social contexts and sensory systems emphasized in various species. Among sexually dimorphic non-territorial Asian elephants,  Elephas maximus, a multiplex olfactory chemical signaling system has been implicated in ensuring effective reproduction. This study explores how, using chemosensory mechanisms, widely roaming, wild male elephants locate periovulatory females in matriarchal-led female family units and precisely assess their ovulatory status. In this species, the dual obstacles of separately living sexes and infrequent oestrus are overcome by lengthy female cycles. During an extended preovulatory period captive females release increasing concentrations of the urinary pheromone (Z)-7-dodecenyl acetate, timed to reach a maximum just before ovulation. The current field studies combined chemical identification and quantification of female urinary (Z)-7-dodecenyl acetate with behavioural observations, monitoring the frequencies of chemosensory responses and premating  behaviours by various categories of males. The results suggest the temporal extension of the preovulatory period effectively provides a synchrony between sexes for successful reproduction. Male elephants undergo a two-decade-long maturation process that involves physical, sexual, social, and physiological maturation. Males older than 30 years are generally large, sexually active, socially adept and capable of sustaining long periods of musth, during which they release secretions distinctive of adult musth.  These older adult males in musth demonstrated significantly more chemosensory responses and premating behaviours than their younger or nonmusth counterparts; they apparently are more skilled at detecting the precise ovulatory status of females. Male-male interactions are affected by size, age, and musth; the winners gain greater access to females, as indicated by the high incidence of mate guarding.  The Asian elephant shares some breeding tactics common to other mammals including some primates (e.g. orangutans) and whales, while the musth parameter adds a unique feature. Fusion-fission events are influenced by elephant reproductive strategies, as roving males join female groups while tracking preovulatory pheromone concentrations.

Raubenheimer, E.J., Ngwenya, S.P., 2005. The role of ivory in the survival of the African elephant
510. SADJ. 60, 426, 430.
Abstract: The unique chequered pattern of polished ivory has created a perverted commercial demand for elephant tusks. The morphologic basis of the pattern, which makes ivory a sought after product for the manufacturing of works of art, is discussed. Chemical analyses of ivory holds great potential in tracing the source of illegally harvested tusks and exposing poorly managed elephant sanctuaries. The impact of uncontrolled ivory hunting on the population genetics of the African elephant is briefly reviewed

Saragusty, J., Hildebrandt, T.B., Natan, Y., Hermes, R., Yavin, S., Goeritz, F., Arav, A., 2005. Effect of egg-phosphatidylcholine on the chilling sensitivity and lipid phase transition of Asian elephant (Elephas maximus) spermatozoa. Zoo Biology 24, 233-245.
Abstract: This study was conducted in an effort to improve our understanding of the response of Asian elephant (Elephas maximus, Em) spermatozoa to chilling.  Semen was collected from two elephant bulls by means of the manual rectal stimulation method. Five out of seven semen collections were deemed to be suitable for use based on motility (ranging from 20% to 60%) and membrane integrity. We evaluated the chilling sensitivity by incubating the sperm with a fluorescent dye (5-carboxyfluorescein diacetate (cFDA)) at 16 degrees C, 12 degrees C, 4 degrees C, and 22 degrees C (control). Cells with an intact membrane retained the dye and were identified as viable. The membrane lipid phase transition (LPT) temperature curve was determined with a Fourier transform infrared (FTIR) spectrometer connected to an FTIR microscope.  The LPT center, T-m, was determined by statistical analysis. The LPT and T-m were also assessed in fresh spermatozoa and spermatozoa incubated with egg yolk or egg-phosphatidylcholine (EPC) liposomes at 16 degrees C, 12 degrees C, 4 degrees C, and 26 degrees C (control). The results show that the membrane integrity of spermatozoa incubated at 16 degrees C, 12 degrees C, and 4 degrees C decreased by 39%, 62%, and 67%, respectively, compared to the control. The LPT temperatures were between room temperature (26 degrees C) and 10 degrees C, with Tm at 14-16 degrees C. The T-m for sperm incubated with liposomes or egg-yolk extender was below the measured range (2 degrees C). Chilling sensitivity was found at a wide range of temperatures and transition temperatures, suggesting the presence of a wide variety of fatty acids (FAs) in the membrane with a high ratio of saturated-to-polyunsaturated FAs. Here we show that the protection  afforded  by the presence of egg yolk or liposomes in the extender is  accomplished by  shifting the T. to below the 4 degrees C point at which chilled semen  is  maintained for transport, or the point at which fast freezing begins to  minimize cellular damage.

Steinetz, B.G., Brown, J.L., Roth, T.L., Czekala, N., 2005. Relaxin concentrations in serum and urine of endangered species: correlations with physiologic events and use as a marker of pregnancy
596. Ann. N. Y. Acad. Sci. 1041, 367-378.
Abstract: Many mammalian species are facing extinction due to problems created by human encroachment, agriculture, pollution, and willful slaughter. Among those at risk are the Asian and African elephant, Sumatran rhinoceros, and giant panda. Conservation groups try to save species in the wild by preserving habitat and limiting animal-human conflicts, often with limited success. Another alternative is to preserve the extant gene pool through captive breeding as a hedge against extinction. Measurement of circulating reproductive hormones is impractical for most wildlife species; determination of urinary or fecal hormone metabolites provides a more viable approach. To aid breeding management, one important tool is the ability to diagnose and monitor pregnancy, especially in species with long gestations (e.g., rhinos over 15 mo and elephants over 20 mo). Unfortunately, measuring progestins often is not useful diagnostically, because concentrations are similar during at least part of the pregnancy and the nonpregnant luteal phase in some species (e.g., elephants, rhinoceroses, and giant pandas). As serum relaxin reliably distinguishes between pregnancy and pseudopregnancy in bitches, relaxin measurement might also provide a method for detecting a successful pregnancy in endangered species. Appropriate immunoassay reagents have enabled the estimation of relaxin concentrations in the serum of elephants and rhinos and the determination of pregnancy establishment and the outcome. Relaxin was also detected in panda serum and urine. However, the extreme variability of the time between observed mating and parturition and the confounding factors of delayed implantation, pseudopregnancy, and frequent fetal resorptions made it impossible to use the panda relaxin data as a specific marker of pregnancy

Stetter, M., Grobler, D., Zuba, J.R., Hendrickson, D., Briggs, M., Castro, L., Neiffer, D., Terrell, S., Robbins, P.K., Stetter, K., Ament, B.S., Wheeler, L. Laprascopic reproductive sterilization as a method of population control in free-ranging African elephants (Loxodonta africana). 2005 Proceedings AAZV, AAWV, AZA Nutrition Advisory Group.  199-200. 2005.
Ref Type: Conference Proceeding

Vinogradov, I.V., Kochneva, G.V., Malkova, E.M., Shchelkunov, S.N., Riabchikova, E.I., 2005. [Intranasal infection in mice inoculated with cowpox virus strain EP-2 isolated from the elephant]
579. Vopr. Virusol. 50, 37-42.
Abstract: The specific features of reproduction of EP-2 strain of cowpox virus (CPV) were studied in intranasally infected BALC/C mice by light and electron microscopy. Virus replication was found in the ciliated, intercalary, basal, and goblet cells (the nasal respiratory area), basal and supporting cells (the nasal olfactory area), ciliated, intercalary, goblet cells (the tracheal and bronchial epithelium), and collagen-producing, Schwann's, endothelial, smooth muscle, and adventitial cells. It has been shown that the CPV strain EP-2 locally replicates in the nasal cavity, trachea, and large bronchi and that there is no generalized infection

Walker, C.L., Stewart, E.A., 2005. Uterine fibroids: the elephant in the room. Science 308, 1589-1592.
Abstract: Uterine fibroids (leiomyomas) have historically been viewed as important chiefly as the major indication for hysterectomy. As new therapies are developed, the heterogeneity of this disease becomes therapeutically relevant. An awareness of the role of genetics, the extracellular matrix, and hormones in tumor etiology is key to understanding this disease. 

Wooding, F.B., Stewart, F., Mathias, S., Allen, W.R., 2005. Placentation in the African elephant, Loxodonta africanus: III. Ultrastructural and functional features of the placenta
598. Placenta 26, 449-470.
Abstract: Successful transfer of nutrients to the elephant fetus during pregnancy relies on a variety of placental modifications. Our light and electron microscopical investigations show that the structure is endotheliochorial from implantation to term, with unicellular, never syncytial trophoblast. Light and electron microscope immunocytochemistry shows the restriction of the glucose transporter 1 isoform to the basolateral surfaces of the trophoblast, with the glucose transporter 3 restricted to the apical plasmalemma of the trophoblast. Glucose transport to the fetus therefore requires a sequential use of both isoforms. Light and electron microscope cytochemistry indicate the presence of iron deposits only in the haemophagous zones confirming their iron transport function. No trophoblast areas with high concentrations of Calcium binding protein, specialised for Calcium transport were found. In situ hybridisation demonstrated the presence of IGF-II mRNA in the trophoblast from the earliest stage, with TGFbeta1 and HGF-SF mRNA expressed subsequently but only IGF-II and HGF mRNA present in the second half of pregnancy. The results are briefly discussed in terms of placental growth and function and indicate that the elephant placenta is another example of a unique solution to the variety of problems posed by a resident fetus

 2004. Elephant Husbandry Resource Guide. International Elephant Foundation, Azle. TX.

Agnew, D.W., Munson, L., Ramsay, E.C., 2004. Cystic endometrial hyperplasia in elephants
741. Vet. Pathol. 41, 179-183.
Abstract: Most captive female elephants are nulliparous and aged and many have endometrial disease, factors that may hinder fertility. This study characterized the pathologic features and demographic distribution of endometrial lesions from 27 captive Asian (Elephas maximus) and 13 African elephants (Loxodonta africanus), 12- to 57-years of age. The principal lesion was marked cystic and polypoid endometrial hyperplasia (CEH), present in 67% of Asian and 15% of African elephants ranging from 26 to 57 years. The lower prevalence in African elephants likely reflects their younger age range in this study. Fourteen of 15 affected elephants with breeding information were nulliparous. These results suggest that CEH and polyps are common in aged nulliparous elephants, and the severity of these lesions may impair fertility. These findings will be useful in the interpretation of ultrasonographic findings during reproductive examinations of potential breeding cows. Also, breeding programs should focus on younger animals

Brown, J.L., Olson, D., Keele, M., Freeman, E.W., 2004. Survey of the reproductive cyclicity status of Asian and African elephants in North America. Zoo Biology 23, 309-321.
Abstract: The Asian and African elephant populations in North America are not self sustaining, and reproductive rates remain low. One problem identified from routine progestagen analyses is that some elephant females do not exhibit normal ovarian cycles. To better understand the extent of this problem, the Elephant TAG/SSP conducted a survey to determine the reproductive status of the captive population based on hormone and ultrasound evaluations. The survey response rates for facilities with Asian and African elephants were 81% and 71%, respectively, for the studbook populations, and nearly 100% for the SSP facilities. Of the elephants surveyed, 49% of Asian and 62% of African elephant females were being monitored for ovarian cyclicity via serum or urinary progestagen analyses on a weekly basis. Of these, 14% of Asian and 29% of African elephants either were not cycling at all or exhibited irregular cycles. For both species, ovarian inactivity was more prevalent in the older age categories (>30 years); however, acyclicity was found in all age groups of African elephants. Fewer elephant females (B30%) had been examined by transrectal ultrasound to assess reproductive-tract integrity, and corresponding hormonal data were available for about three-quarters of these females. Within this subset, most (B75%) cycling females had normal reproductive-tract morphologies, whereas at least 70% of noncycling females exhibited some type of ovarian or uterine pathology. In summary, the survey results suggest that ovarian inactivity is a significant reproductive problem for elephants held in zoos, especially African elephants. To increase the fecundity of captive elephants, females should be bred at a young age, before reproductive pathologies occur. However, a significant number of older Asian elephants are still not being reproductively monitored. More significantly, many prime reproductive-age (10-30 years) African females are not being monitored. This lack of information makes it difficult to determine what factors affect the reproductive health of elephants, and to develop mitigating treatments to reinitiate reproductive cyclicity.

Brown, J.L., Walker, S.L., Moeller, T., 2004. Comparative endocrinology of cycling and non-cycling Asian (Elephas maximus) and African (Loxodonta africana) elephants
729. Gen. Comp Endocrinol. 136, 360-370.
Abstract: Up to 14% of Asian and 29% of African elephants in captivity are not cycling normally or exhibit irregular cycles based on progestin profiles. To determine if ovarian acyclicity is related to other disruptions in endocrine activity, serum pituitary, thyroid, adrenal, and ovarian hormones in weekly samples collected for 6-25 months were compared between normal cycling (n=22 each species) and non-cycling (n=6 Asian; n=30 African) elephants. A subset of cycling females (n=4 Asian, 7 African) also were blood sampled daily during the follicular phase to characterize the peri-ovulatory period. In normal cycling females, two leutinizing hormone (LH) surges were observed 3 weeks apart during a normal follicular phase, with the second inducing ovulation (ovLH). Serum FSH concentrations were highest at the beginning of the non-luteal phase, declining to nadir concentrations within 4 days of the ovLH surge. FSH remained low until after the ovLH surge and then increased during the luteal phase. A species difference was noted in prolactin secretion. In the African elephant, prolactin was increased during the follicular phase, but in Asian elephants concentrations remained stable throughout the cycle. Patterns of thyroid hormones (thyroid-stimulating hormone, TSH; free and total thyroxine, T4; free and total triiodothyronine, T3) and cortisol secretion were not affected by estrous cycle stage or season in cycling elephants. In non-cycling elephants, there were no fluctuating patterns of LH, FSH, or prolactin secretion. Overall mean concentrations of all hormones were similar to those in cycling animals, with the exception of FSH, prolactin, and estradiol. Mean serum FSH concentrations were lower due to females not exhibiting normal cyclic increases, whereas serum estradiol was higher overall in most acyclic females. Prolactin concentrations were significantly increased in 11 of 30 non-cycling females, all of which were African elephants. In sum, while there were no consistent endocrine anomalies associated with ovarian acyclicity, hyperprolactinemia may be one cause of ovarian dysfunction. The finding of elevated estrogens in some acyclic females also deserves further investigation, especially determining how it relates to reproductive tract pathologies

Brown, J.L., Goritz, F., Pratt-Hawkes, N., Hermes, R., Galloway, M., Graham, L.H., Gray, C., Walker, S.L., Gomez, A., Moreland, R., Murray, S., Schmitt, D.L., Howard, J., Lehnhardt, J., Beck, B., Bellem, A., Montali, R., Hildebrandt, T.B., 2004. Successful artificial insemination of an Asian elephant at the National Zoological Park. Zoo Biology 23, 45-63.
Abstract: For decades, attempts to breed elephants using artificial insemination (AI) have failed despite considerable efforts and the use of various approaches. However, recent advances in equipment technology and endocrine-monitoring techniques have resulted in 12 elephants conceiving by AI within a 4-year period (19982002). The successful AT technique employs a unique endoscope-guided catheter and transrectal ultrasound to deliver semen into the anterior vagina or cervix, and uses the "double LH surge" (i.e., identifying the anovulatory LH (anLH) surge that predictably occurs 3 weeks before the ovulatory LH (ovLH) surge to time insemination. This study describes the 6-year collaboration between the National Zoological Park (NZP) and the Institute for Zoo Biology and Wildlife Research (IZW), Berlin, Germany, that led to the refinement of this AI technique and subsequent production of an Asian elephant calf. The NZP female was the first elephant to be inseminated using the new AI approach, and was the fifth to conceive. A total of six AI trials were conducted beginning in 1995, and conception occurred in 2000. Semen was collected by manual rectal stimulation from several bulls in North America. Sperm quality among the bulls was variable and was thus a limiting factor for AI. For the successful AI, semen quality was good to excellent (75-90% motile sperm), and sperm was deposited into the anterior vagina on the day before and the day of the ovLH surge. Based on transrectal ultrasound, ovulation occurred the day after the ovLH surge. Pregnancy was monitored by serum and urinary progestagen, and serum prolactin analyses in samples collected weekly. Fetal development was assessed at 12, 20, and 28 weeks of gestation using transrectal ultrasound. Elevated testosterone measured in the maternal circulation after 36 weeks of gestation reliably predicted the calf was a male. Parturition was induced by administration of 40 IU oxytocin 3 days after serum progestagens dropped to undetectable baseline levels. We conclude that AI has potential as a supplement to natural breeding, and will be invaluable for improving the genetic management of elephants, provided that problems associated with inadequate numbers of trained personnel and semen donors are resolved.

Carter, A.M., Enders, A.C., Kunzle, H., Oduor-Okelo, D., Vogel, P., 2004. Placentation in species of phylogenetic importance: The Afrotheria. Animal Reproduction Science 82-83, 35-48.
Abstract: Afrotheria, one of four mammalian superorders, comprises elephants, sea cows, hyraxes, aardvark, elephant shrews, tenrecs and golden moles. Their placentas either form an equatorial band or are discoid in shape. The interhemal region, separating fetal and maternal blood, is endotheliochorial in elephants, aardvark and possibly the sea cows, but hemochorial in the remaining orders. There is a secondary epitheliochorial placenta in elephant shrews while a similar structure in tenrecs erodes maternal tissues. Specialized hemophagous regions are a striking characteristic of some of these placentas yet absent in hyraxes, elephant shrews, and golden moles. It is possible that the common ancestor of the Afrotheria had an endotheliochorial placenta. Establishment of a hemochorial condition, as seen in rock hyraxes, elephant shrews, tenrecs, and golden moles, would be a more recent development. The elephant, manatee, and aardvark all have circumferential placentas. Thus the formation of a discoid placenta with a more or less extensive secondary placenta in elephant shrews and tenrecs would also be a derived state.

Carter, A.M., Enders, A.C., 2004. Comparative aspects of trophoblast development and placentation. Reproductive Biology and Endocrinology 2.
Abstract: Based on the number of tissues separating maternal from fetal blood, placentas are classified as epitheliochorial, endotheliochorial or hemochorial. We review the occurrence of these placental types in the various orders of eutherian mammals within the framework of the four superorders identified by the techniques of molecular phylogenetics. The superorder Afrotheria diversified in ancient Africa and its living representatives include elephants, sea cows, hyraxes, aardvark, elephant shrews and tenrecs. Xenarthra, comprising armadillos, anteaters and sloths, diversified in South America. All placentas examined from members of these two oldest superorders are either endotheliochorial or hemochorial. The superorder Euarchontoglires includes two sister groups, Glires and Euarchonta. The former comprises rodents and lagomorphs, which typically have hemochorial placentas. The most primitive members of Euarchonta, the tree shrews, have endotheliochorial placentation. Flying lemurs and all higher primates have hemochorial placentas. However, the lemurs and lorises are exceptional among primates in having epitheliochorial placentation. Laurasiatheria, the last superorder to arise, includes several orders with epitheliochorial placentation. These comprise whales, camels, pigs, ruminants, horses and pangolins. In contrast, nearly all carnivores have endotheliochorial placentation, whilst bats have endotheliochorial or hemochorial placentas. Also included in Laurasiatheria are a number of insectivores that have many conserved morphological characters; none of these has epitheliochorial placentation. Consideration of placental type in relation to the findings of molecular phylogenetics suggests that the likely path of evolution in Afrotheria was from endotheliochorial to hemochorial placentation. This is also a likely scenario for Xenarthra and the bats. We argue that a definitive epitheliochorial placenta is a secondary specialization and that it evolved twice, once in the Laurasiatheria and once in the lemurs and lorises.

Dahl, N.J., Olson, D., Schmitt, D., Blasko, D.R., Kristipati, R., Roser, J.F., 2004. Development of an enzyme-linked immunosorbent assay (ELISA) for luteinizing hormone (LH) in the elephant (Loxodonta africana and Elephas maximus). Zoo Biology 23, 65-78.
Abstract: A simple, rapid enzyme-linked immunosorbent assay (ELISA) for the measurement of LH in plasma and serum of elephants (Loxodonta africana and Elephas maximus) has been developed, validated, and used for comparative studies. Purified elephant LH (eleLH) diluted in elephant plasma was used as standards (0.78-50 ng/ml). A monoclonal antibody against the beta-subunit of bovine LH (518B(7)) was used as the capture antibody. The second antibody (a polyclonal rabbit anti-human LH antibody), conjugated to horseradish peroxidase, cleaved a substrate (tetramethyl benzidine), resulting in a color change. The total assay time was approximately 21/2 hr, with incubations at room temperature. Sensitivity was found to be 1.56 ng/ml. Cross-reactivities to elephant FSH and TSH were low: 0.9% and 0.15%, respectively. The accuracy of the assay was demonstrated by comparing the ELISA with a validated eleLH radioimmunoassay (RIA), progesterone data, and  ultrasound observations. Blood samples from 18 Asian and African elephant cows were analyzed with the ELISA and RIA, and an additional 11 cows were used to describe endocrine parameters for LH and progesterone using only RIA. No difference was found in LH peak concentrations between the ELISA and RIA. The time from the progesterone decline to the first LH peak, and the time between the two peaks were similar between species. Asian cows had higher LH peaks than African cows. Ultrasound confirmed the time of ovulation occurring with the second LH peak. Three cows were inseminated and confirmed to be pregnant using this ELISA as a timing device. Instrumentation is not always required, as LH peaks approximating 3 ng/ml can be visually observed. In conclusion, this ELISA can be used as a field test to determine time of ovulation for artificial insemination (AI) or natural breeding of both species of elephants, and thus is an important tool for the preservation of captive populations worldwide.

de Oliveira, C.A., West, G.D., Houck, R., Leblanc, M., 2004. Control of musth in an Asian elephant bull (Elephas maximus) using leuprolide acetate
716. J. Zoo. Wildl. Med. 35, 70-76.
Abstract: The results of long-term administration of leuprolide acetate (LA) depot in a 52-yr-old Asian elephant bull (Elephas maximus) for control of musth are presented. Twelve injections were administered for 6 yr during our interpretation of early musth or "premusth." Intervals between musth periods during the study varied from 2 to 34 mo. Blood samples, drawn weekly, were assayed for serum testosterone concentrations; mean levels were 11.78 +/- 1.97 nmol/L throughout the first 26 mo of the study, 7.28 +/- 1.28 nmol/L during the following 21 mo, and 0.45 +/- 0.035 nmol/L in the last 34 mo of this study. Early musth signs ceased within 3 days of drug administration after 10 of 12 injections. The mean serum testosterone concentrations were significantly decreased by the last 34 mo of the study. The results suggest leuprolide is a suitable alternative for controlling or preventing (or both) musth in captive Asian elephants, although permanent reproductive effects may occur. Zoos and wildlife conservation institutions could benefit from the use of LA in Asian elephants to increase the male availability in captivity, consequently ensuring genetic diversity and the perpetuation of the species

deOliveira, C.A., West, G.D., Houck, R., Leblanc, M., 2004. Control of musth in an Asian elephant bull (Elephas maximus) using leuprolide acetate. Journal of Zoo and Wildlife Medicine 35 , 70-76.
Abstract: The results of long-term administration of leuprolide acetate (LA) depot in a 52-yr-old Asian elephant bull (Elephas maximus) for control of musth are presented. Twelve injections were administered for 6 yr during our interpretation of early musth or "premusth". Intervals between musth periods during the study varied from 2 to 34 mo. Blood samples, drawn weekly, were assayed for serum testosterone concentrations; mean levels were 11.78 +/- 1.97 nmol/L throughout the first 26 mo of the study. 7.28 +/- 1.28 nmol/L during the following 21 mo. and 0.45 +/- 0.035 nmol/L in the last 34 mo of this study. Early musth signs ceased within 3 days of drug administration after 10 of 12 injections. The mean serum testosterone concentrations were significantly decreased by the last 34 mo of the study. The results suggest leuprolide is a suitable alternative for controlling or preventing (or both) musth in captive Asian elephants, although permanent reproductive effects may occur. Zoos and wildlife conservation institutions could benefit from the use of LA in Asian elephants to increase the male availability in captivity, consequently ensuring genetic diversity and perpetuation of the species.

Freeman, E.W., Weiss, E., Brown, J.L., 2004. Examination of the interrelationships of behavior, dominance status, and ovarian activity in and African elephants. Zoo Biology 23, 431-448.
Abstract: Ovarian inactivity has been identified in captive African (Loxodonta africana) and Asian (Elephas maximus) elephants and is thought to be mediated in part by social influences. Thus, a survey was conducted to determine how behavior and dominance status relate to each other and to ovarian cyclicity. For both Asian and African elephants, dominance status was positively correlated with relative size, age, temperament, disciplinary nature, and willingness to share novel objects. Relative size and temperament were also related to disciplinary nature toward herdmates. Behavior toward keepers was a good indicator of the willingness of elephants to follow cmmands, whereas sharing novel objects was positively correlated with object curiosity. Finally, dominance status, temperament, and disciplinary nature were all correlated with willingness to share. Comparisons of ovarian cyclicity status with behavior rankings were conducted only for African elephants because of the low number of noncycling Asian elephants surveyed. Overall, social status appeared to be the best predictor of ovarian activity in African elephants. Noncycling African elephant females ranked higher in the dominance hierarchy and gave more discipline to herdmates than cycling cohorts. It remains to be determined whether these are cause or effect relationships, but clearly it is important to understand how physical and social attributes impact physiological processes, such as reproduction. Captive management now needs to focus on optimizing social and environmental conditions to maximize reproductive potential in elephants.

Garstang, M., 2004. Long-distance, low-frequency elephant communication. J Comp Physiol A 190, 791-805.
Abstract: The production, transmission, and reception of and the behavioral response to long-distance, low-frequency sound by elephants is reviewed. The structure of low-frequency calls generated by elephants is separated into the ''source'' and the ''filter'' roles played by the lungs, larynx and vocal track, the composition of the expired air and the ambient air temperature. Implications regarding the size, age, sex, sexual and physical status follow from the call structure and detection. Reception of the signal is discussed in terms of the characteristics of the elephant's ear with particular attention to the determination of the threshold of hearing and the ability to locate the source of low-frequency sounds. Factors which influence the transmission of near infrasound are related to atmospheric structure. The critical role played by the thermal stratification and vertical gradient and magnitude of the wind in determining both the range and the detection of a signal are discussed for open and closed elephant habitats. Infrasound plays a pervasive role in reproduction, resource utilization, avoidance of predation and other social interactions. Current and future technology can be expected to contribute to the detection and interpretation of elephant communication. This will aid in the understanding of behavior and in efforts to sustain the species.

Garstang, M., 2004. Long-distance, low-frequency elephant communication
686. J. Comp Physiol A Neuroethol. Sens. Neural Behav. Physiol 190, 791-805.
Abstract: The production, transmission, and reception of and the behavioral response to long-distance, low-frequency sound by elephants is reviewed. The structure of low-frequency calls generated by elephants is separated into the "source" and the "filter" roles played by the lungs, larynx and vocal track, the composition of the expired air and the ambient air temperature. Implications regarding the size, age, sex, sexual and physical status follow from the call structure and detection. Reception of the signal is discussed in terms of the characteristics of the elephant's ear with particular attention to the determination of the threshold of hearing and the ability to locate the source of low-frequency sounds. Factors which influence the transmission of near infrasound are related to atmospheric structure. The critical role played by the thermal stratification and vertical gradient and magnitude of the wind in determining both the range and the detection of a signal are discussed for open and closed elephant habitats. Infrasound plays a pervasive role in reproduction, resource utilization, avoidance of predation and other social interactions. Current and future technology can be expected to contribute to the detection and interpretation of elephant communication. This will aid in the understanding of behavior and in efforts to sustain the species

Graham, L.H., Bando, J., Gray, C., Buhr, M.M., 2004. Liquid storage of Asian elephant (Elephas maximus) sperm at 4 degrees C
737. Anim Reprod. Sci. 80, 329-340.
Abstract: The Asian elephant (Elephas maximus) population in the wild has been in decline for several decades and breeding in captivity has not been self-sustaining. The use of artificial insemination (AI) can help overcome many of the difficulties associated with breeding elephants in captivity; however, the ability to store semen for extended periods of time is critical to the successful application of AI to elephants. The objective of the present study was to assess the effects of four different semen extenders and the presence of egg yolk on the viability and motility of Asian elephant semen stored at 4 degrees C. High quality ejaculates (n=4) were collected from two Asian elephant bulls by rectal massage. Aliquots of each ejaculate were extended in four different diluents (Beltsville thawing solution (BTS); Tris-citric acid (TCA)/fructose-based; Beltsville F5 (BF5); dextrose-supplemented phosphate-buffered saline (PBS)) with or without egg yolk then cooled and stored at 4 degrees C. The percentages of viable (viability) and motile (motility) sperm were evaluated at 8, 24 and 48 h following collection. The addition of egg yolk significantly reduced the percentage loss in viability from initial collection to 48 h compared to extenders without egg yolk (17.0 +/- 8.2 versus 32.6 +/- 8.9 decline in percent viable sperm in the population, respectively; P<0.05). Extender and egg yolk affected (P<0.005) total motility and percent progressively motile sperm at all evaluation times during incubation. TCA + egg yolk maintained higher (P<0.05) levels of progressive motility compared to other extenders supplemented with egg yolk. These results indicate that Asian elephant semen extended in TCA diluent supplemented with egg yolk can maintain at least 50% viability and motility when stored at 4 degrees C for 48 h.

Hermes, R., Hildebrandt, T.B., Goritz, F., 2004. Reproductive problems directly attributable to long-term captivity--asymmetric reproductive aging
704. Anim Reprod. Sci. 82-83, 49-60.
Abstract: Problems attributable to long-term captivity have been identified and are responsible for the difficulties in establishing successful reproduction in captive populations of wildlife, specifically, elephants and rhinoceroses. Historically, non-reproductive periods of 10-15 years in nulliparous female rhinoceroses and elephants have not been considered problematic. New evidence suggests that prolonged exposure to endogenous sex steroids and that long stretches of non-reproductive periods induce asymmetric reproductive aging in captive animals. The consequences are reduced fertility, shortened reproductive life-span and, eventually, irreversible acyclicity. Although age-related reproductive lesions have also been documented in male rhinoceroses, they continue to maintain a longer reproductive life-span than females. Since human and domestic animal models have already indicated that early pregnancy provides natural protective mechanism against asymmetric reproductive aging processes and premature senescence, it is imperative that appropriate counter measures such as assisted reproductive technologies (ART) be utilized to ensure early pregnancy in captive animals for their preservation and to ensure increased genetic diversity of the captive populations

Hermes, R., Hildebrandt, T.B., Goritz, F., 2004. Reproductive problems directly attributable to long-term captivity-asymmetric reproductive aging. Animal Reproduction Science 82-83, 49-60.
Abstract: Problems attributable to long-term captivity have been identified and are responsible for the difficulties in establishing successful reproduction in captive populations of wildlife, specifically, elephants and rhinoceroses. Historically, non-reproductive periods of 10-15 years in nulliparous female rhinoceroses and elephants have not been considered problematic. New evidence suggests that prolonged exposure to endogenous sex steroids and that long stretches of non-reproductive periods induce asymmetric reproductive aging in captive animals. The consequences are reduced fertility, shortened reproductive life-span and, eventually, irreversible acyclicity. Although age-related reproductive lesions have also been documented in male rhinoceroses, they continue to maintain a longer reproductive life-span than females. Since human and domestic animal models have already indicated that early pregnancy provides natural protective mechanism against asymmetric reproductive aging processes and premature senescence, it is imperative that appropriate counter measures such as assisted reproductive technologies (ART) be utilized to ensure early pregnancy in captive animals for their preservation and to ensure increased genetic diversity of the captive populations.

Hildebrandt, T.B., Hermes, R., Janssen, D.L., Oosterhuis, J.E., Murphy, D., Göritz, F. Reproductive evaluation in wild African elephants prior to translocation. 2004 PROCEEDINGS AAZV, AAWV, WDA JOINT CONFERENCE.  75-76. 2004.
Ref Type: Conference Proceeding
Abstract: Translocations of wild African (Loxodonta africana) elephants have increased significantly since 1993 after Clem Coetzee developed a new method to move adult elephants in Zimbabwe. Since then the technique have been optimized mainly by the staff of the Kruger National Park (KNP) and over 750 elephants in family units and almost 100 mature bulls have been translocated by the KNP capture team.1 The translocations were mainly performed for reducing the number of elephants in KNP and for stocking other reserves. Few elephants were also moved for overseas export to international zoological institutions. However, each elephant translocation is always a logistic challenge and is extremely costly. Therefore, it is very important to select the right elephants or elephant groups for the future translocation. If the main goal of a translocation is the establishment of a new breeding group, it is especially important to select infertile individuals and highly pregnant females which could have a miscarriage due to the transport stress. The IZW team developed a field applicable portable ultrasound technique which allows the reproductive ev ry Killmar (ZSSD), and Randy Rieches (ZSSD).

LITERATURE CITED
1. Hofmeyr, M. 2003. Translocation as a management tool for control of elephant populations. Managing African Elephant Populations: Act or Let Die. Beekbergen, The Netherlands, 6.-7.Nov., Pp. 38-39.
2. Hildebrandt T. B., F. Göritz, N. C. Pratt, D. L. Schmitt, S. Quandt, J. Raath and R. R. Hofmann. 1998. Reproductive assessment of male elephants (Loxodonta africana and Elephas maximus) by ultrasonography. J. Zoo Wildl. Med. 29: 114-128.

Jones, C.J., Wooding, F.B., Mathias, S.S., Allen, W.R., 2004. Fetomaternal glycosylation of early placentation events in the African elephant Loxodonta africana
740. Placenta 25, 308-320.
Abstract: During implantation in the African elephant (Loxodonta africana), fetal trophoblast displaces the surface uterine epithelium and superficially penetrates the uterine glands. This limited invasion is followed by the upgrowth of blunt fingers of endometrial stroma, covered with trophoblast and containing capillaries that subsequently vascularize the growing placenta. We have used lectin histochemistry to compare the glycosylation of maternal endothelial cells in the endometrium with those growing within the trophoblastic processes of a 2 g embryo (approximately 125 days' gestation), and also examine changes in the endometrial glands associated with trophoblastic invasion. Maternal vessels at the apices of the trophoblast-covered stromal upgrowths showed increased expression of terminal N-acetyl galactosamine, N-acetyl glucosamine oligomers, some sialic acids, and tri/tetra-antennate non-bisected complex N-linked glycan, as indicated by increased lectin staining. The areas of increased staining were also more resistant to neuraminidase digestion. Invaded glands had distended walls composed of flattened epithelial cells, some of which showed heavy lectin staining suggestive of intracellular glycan accumulation. The vascular changes suggest that new maternal capillary growth is accompanied by alterations in surface glycosylation. This may be the result of increased glycosyl transferase activity associated with cell proliferation and may also indicate the expression of significantly increased anti-adhesive molecules preventing blood stasis and egress of maternal immunocompetent cells into the fetal compartment

Meyer, J.M., Walker, S.L., Freeman, E.W., Steinetz, B.G., Brown, J.L., 2004. Species and fetal gender effects on the endocrinology of pregnancy in elephants
682. Gen. Comp Endocrinol. 138, 263-270.
Abstract: Quantitative and temporal progestin profiles vary during gestation in the elephant, sometimes making it difficult to determine if a pregnancy is progressing normally. The aim of the present study was to determine if circulating progestin variability was related to species or fetal gender effects. A similar comparison also was conducted for secretory profiles of prolactin, relaxin, and cortisol. Overall mean progestin concentrations during gestation in Asian (n = 19) and African (n = 8) elephants were similar; however, the temporal profiles differed (P < 0.001). Concentrations were higher in African elephants during the first half of pregnancy, but then declined to levels below those observed in Asian elephants (P < 0.05). There also was a fetal gender effect in Asian, but not African elephants. Progestin concentrations were higher in Asian cows carrying male calves (n = 9) as compared to those carrying females (n = 10) (P < 0.001). Overall prolactin concentrations were higher in Asian than in African elephants between 8 and 15 months of gestation ( P< 0.001). There were no species differences in the secretory patterns of relaxin. Cortisol was relatively stable until the end of gestation when significant surges were observed, mainly between 8 and 11 days before parturition, and again on the day of birth. In sum, a comparison of progestin patterns between Asian and African elephants identified notable differences related to species and fetal gender. A role for cortisol in the initiation of parturition also was inferred from these data. From a practical standpoint, understanding the factors affecting gestational hormone characteristics and recognizing what the species differences are will help ensure that data used in diagnosing and monitoring elephant pregnancies are properly interpreted

Ruedi, D. Collection and handling of semen and insemination in the African elephant (Loxodonta africana) at Basle Zoo.  2004.
Ref Type: Unpublished Work
Abstract: It is urgently necessary that we take measures towards enhancing the efficiency of captive breeding of African and Asian elephants. This might be accomplished in several ways: for instance, by an extensive loan of cows for natural mating and, last but not least, by breeding under artificial conditions. With the third possibility in mind we initiated a programme in 1977 to develop a technique for artificial insemination in the African elephant; it encompassed the collection and handling of semen, oestrus detection and insemination.

Sanchez, C.R., Murray, S., Montali, R.J., Spelman, L.H., 2004. Diagnosis and treatment of presumptive pyelonephritis in an Asian elephant (Elephas maximus). J Zoo Wildl Med 35, 397-399.
Abstract: A 37-yr-old female Asian elephant (Elephas maximus) presented with anorexia, restlessness, and dark-colored urine. Urinalyses showed hematuria, leukocyturia, isosthenuria, proteinuria, granular casts, and no calcium oxalate crystals. Bloodwork revealed azotemia. Urine culture revealed a pure growth of Streptococcus zooepidemicus resistant to sulfamethoxazole-trimethoprim but susceptible to cephalosporins. A presumptive diagnosis of pyelonephritis was made based on bloodwork, urinalysis, and urine culture. The animal was treated with intravenous ceftiofur, and intravenous and per rectum fluids were given for hydration. The elephant's attitude and appetite returned to normal, the abnormal blood parameters resolved, and urinary calcium oxalate crystals reappeared after treatment, supporting presumptive diagnosis. Follow-up ultrasonography revealed an abnormal outline of both kidneys with parenchymal hyperechogenicity and multiple uterine leiomyomas.

Stumpf, P., Welsch, U., 2004. Secretory and defensive functions of the duct system of the lactating mammary gland of the African elephant (Loxodonta africana, Proboscidea). Zoomorphology 123, 155-157.
Abstract: The duct system of the lactating mammary Gland of the African elephant (Loxodonta africana) was investigated with histochemical and immunohistochemical techniques and with the transmission electron microscope in order to detect specific cell biological differentiations in the ductal epithelia of this species, which is marked by an unusually long lactation period. General histology and electron microscopy allow to distinguish several segments in the entire duct system. The apical membranes of the epithelia have binding sites for several lectins [Canavalia ensiformis agglutinin (ConA), Ricinus communis agglutinin (RCA 1), Wisteria floribunda agglutinin (WFA), peanut agglutinin (PNA)] and also stain with alcian blue indicating the presence of a highly differentiated negatively charged glycocalyx forming an effective barrier between lumen and epithelium. Cytokeratins, actin, tubulin and vinculin show different expression intensities in the proximal and distal portion of the duct system. Lysozyme, lactoferrin, the secretory component of IgA and human beta defensin-2 are expressed in the epithelium of the entire duct system. In the distal portion of the ducts the staining intensity is stronger than in the proximal portion. We conclude that the duct system of the elephant mammary gland has specific secretory functions and that the secretory products are part of the defensive mechanisms against invading microorganisms.

Thongtip, N., Saikhun, J., Damyang, M., Mahasawangkul, S., Suthunmapinata, P., Yindee, M., Kongsila, A., Angkawanish, T., Jansittiwate, S., Wongkalasin, W., Wajjwalkul, W., Kitiyanant, Y., Pavasuthipaisit, K., Pinyopummin, A., 2004. Evaluation of post-thaw Asian elephant (Elephas maximus) spermatozoa using flow cytometry: the effects of extender and cryoprotectant
711. Theriogenology 62, 748-760.
Abstract: Although the development of semen cryopreservation in the African elephants (Loxodonta africana) has been accomplished, effective procedures for cryopreservation of Asian elephant (Elephas maximus) spermatozoa have not been established. In the present study, we investigate the freezing methods for conservation of Asian elephant spermatozoa under field conditions and identify the most suitable freezing protocols which provide acceptable post-thaw semen quality. Semen was collected from two Asian elephant bulls (EM1 and EM2, 10 ejaculates from each bull) by manual manipulation and were assessed for volume, pH, sperm cell concentration, and progressive motility. Eight out of 20 ejaculates were of acceptable quality (progressive motility >/= 60%), and were used for cryopreservation studies. Semen were frozen in TEST + glycerol, TEST + DMSO, HEPT + glycerol, or HEPT + DMSO. The post-thaw progressive sperm motilities were assessed, and sperm cells were stained with PI and FITC-PNA for membrane and acrosomal integrity assessment using flow cytometry. Post-thaw progressive motility of spermatozoa (EM1: 42.0 +/- 4.3%; EM2: 26.0 +/- 17.3%) and the percentage of membrane and acrosome intact spermatozoa (EM1: 55.5 +/- 8.1%; EM2: 46.3 +/- 6.4%) cryopreserved in TEST + glycerol were significantly higher than (P < 0.05) those frozen in the other medium investigated choices for cryopreservation of Asian elephant spermatozoa. The data support the use of TEST + glycerol as an acceptable cryopreservation media of Asian elephant semen for the establishment of sperm banks

Visscher, D.R., vanAarde, R.J., Whyte, I., 2004. Environmental and maternal correlates of foetal sex ratios in the African buffalo (Syncerus caffer) and savanna elephant (Loxodonta africana). Journal of Zoology 263 , 111-116.
Abstract:  Many species exhibit skewed sex ratios at birth. Here we investigate the relationships between environmental and maternal variables (as surrogates for maternal condition) and foetal sex in African buffalo Syncerus caffer and elephant Loxodonta africana of the Kruger National Park, South Africa. Using logistic regression no significant effect was found of year, maternal lactational status, maternal age, rainfall or density on foetal sex ratio. Using a subset of our data, it was also concluded that maternal body condition did not affect foetal sex ratio in buffalo. Our analyses failed to support hypotheses predicting that mothers will skew the sex ratios of their offspring in relation to their body condition. In this study, buffalo and elephant produced offspring with a sex ratio close to parity. Our results are discussed in light of the implications for testing such hypotheses in analyses of population level.

 2003. Healthcare, Breeding and Management of Asian Elephants. Project Elephant. Govt. of India, New Delhi.

Allen, W.R., Mathias, S., Wooding, F.B., van Aarde, R.J., 2003. Placentation in the African elephant (Loxodonta africana): II morphological changes in the uterus and placenta throughout gestation. Placenta 24, 598-617.
Abstract: The gross and microscopic development of the zonary endotheliochorial placenta in the African elephant was studied in 22 gravid uteri that ranged in
gestational stage from 0.5 to 20.6 months. The conceptus only ever occupies one horn of the uterus and is associated with 2-5 large corpora lutea that persist in the ipsilateral ovary throughout gestation. Initially, the trophoblast in the equatorial region of the conceptus completely replaces the lumenal epithelium of the endometrium to which it is apposed. Blunt upgrowths of endometrial stroma then develop, each closely invested by trophoblast, and  containing the capillaries that will vascularize this maternal component of the resulting placental band. With advancing gestation the lamellate stromal upgrowths increase markedly in length and become much thinner, thereby bringing the trophoblast into intimate contact with the endothelium of the maternal capillaries. They also become folded or pleated to increase the total area of intimate feto-maternal contact. At the lateral edges of the placental band the lamellae bend over towards the endometrium to form a blind cleft. Leakage of blood into this area creates haemophagous zones in which phenotypically specialized trophoblast cells phagocytose the blood components. The presence of large resorbing blood clots and circumferential scars in the uteri of three post parturient animals initiated the hypothesis that, when the standing elephant gives birth at term, the passage of the 120 kg fetus through the vagina may wrench the placenta off the endometrium by severing its very narrow maternal placental hilus. The resulting intrauterine haemorrhage may then play a role in preventing further conception for around 2 years.Department of Clinical Veterinary Medicine Equine Fertility Unit, University of Cambridge, Mertoun Paddocks,Woodditton Road, Suffolk CB8 9BH, Newmarket, UK. vetart@aht.org.uk

Bhattacharyya, B.K., 2003. Some reproductive traits of elephants. In: Das, D. (Ed.), Healthcare, Breeding and Management of Asian Elephants. Project Elephant. Govt. of India, New Delhi, pp. 37-41.

Czekala, N.M., MacDonald, E.A., Steinman, K., Walker, S., Garrigues, N.W., Olson, D., 2003. Estrogen and LH dynamics during the follicular phase of the estrous cycle in the Asian elephant. Zoo Biology 22, 443-454.
Abstract: Pituitary and corpus luteum hormone patterns throughout the elephant estrous cycle have been well characterized. By contrast, analysis of follicular maturation by measurement of circulating estrogens has been uninformative. This study tested the ability of a urinary estradiol-3-glucuronide radioimmunoassay to noninvasively assess follicular development during the nonluteal phase of the elephant estrous cycle, and to determine the relationship between estrogen production and the "double LH surge." Daily urine and serum samples were collected throughout seven estrous cycles from three Asian elephants, and urine was collected from an additional three females, for a total of 13 cycles. Serum was analyzed for luteinizing hormone (LH), and urine was analyzed for estrogens and progestins. Elephants exhibited a typical LH pattern, with an anovulatory LH (anLH) surge occurring approximately 21 days before the ovulatory LH (ovLH) surge. The urinary estrogen pattern indicated the presence of two follicular waves during the nonluteal phase. The first wave (anovulatory) began 5 days before the anLH surge and reached a maximum concentration the day before the peak. Thereafter, urinary estrogens declined to baseline for 2 weeks before increasing again to peak concentrations on the day of the ovLH surge. Urinary progestins were baseline throughout most of the follicular phase, increasing 2-3 days before the ovLH surge and continuing into the luteal phase. These results support previous ultrasound observations that two waves of follicular growth occur during the nonluteal phase of the elephant estrous cycle. Each wave is associated with an increase in estrogen production that stimulates an LH surge. Thus, in contrast to serum analyses, urinary estrogen monitoring appears to be a reliable method for characterizing follicular activity in the elephant.

Das, D., 2003. Breeding, reproduction and conservation strategies in Asian elephants. In: Das, D. (Ed.), Healthcare, Breeding and Management of Asian Elephants. Project Elephant. Govt. of India, New Delhi, pp. 45-57.

Davis, B.L., Dill, W.M., Hicks, A.R., Goodwin, T.E., Hollister-Smith, J., Alberts, S.C. Use of SPME and GC-MS for chemical analysis of urine from African elephants in musth. Abstracts Of Papers Of The American Chemical Society 225[614-CHED Part 1 MAR 2003]. 2003.
Ref Type: Abstract
Abstract: Addresses: Hendrix Coll, Dept Chem, Conway, AR 72032 USA ;Duke Univ, Dept Biol Sci, Durham, NC 27706 USA

Dehnhard, M., Hatt, J.M., Eulenberger, K., Ochs, A., Strauss, G., 2003. Headspace solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS) for the determination of 5alpha-androst-2-en-17-one and -17beta-ol in the female Asian elephant: application for reproductive monitoring and prediction of parturition. J Steroid Biochem Mol Biol. 84, 383-391.
Abstract: Asian elephants are not self-sustaining in captivity. The main reasons for this phenomenon are a low birth rate, an aging population, and poor calf-rearing. Therefore, it is essential that reproductive rates had to be improved and there is need for rapid quantitative measures to monitor reproductive functions focussing on estrous detection and the prediction of the period of parturition. The objective of this study was to develop a method which combines headspace solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS) for analyses of 5alpha-androst-2-en-17beta-ol and -17-one to prognose estrous and to predict the period of parturition. SPME was carried out with a CTC Combi Pal system.The course of the luteal phase-specific substance 5alpha-androst-2-en-17beta-ol and -17-one followed a cyclic pattern in which the follicular and luteal phases could be clearly distinguished (mean estrous cycle length, 15+/-1.4 weeks). Based on daily urine samples, estrous prognosis might be possibly based on the initial 5alpha-androst-2-en-17beta-o1 increase at the end of the follicular phase. Parturition prognosis was performed in three elephant cows based on the 5alpha-androst-2-en-17beta-o1 drop to baseline levels 5-4 days prior parturition. Experiments revealed that 5alpha-androst-3alpha-ol-17-one and probably 5alpha-androst-3alpha-ol-17beta-ol are generated from sulfate conjugates by a thermal process. Institute for Zoo Biology and Wildlife Research, PF 601103, D-10252 Berlin, Germany. dehnhard@izw-berlin.de

Dill, W.M., Davis, B.L., Hicks, A.R., Goodwin, T.E., Rasmussen, L.E.L., Loizi, H., Schulte, B.A. Chemical analysis of preovulatory female African elephant urine: A search for putative pheromones. Abstracts Of Papers Of The American Chemical Society 225[409-CHED Part I Mar 2003]. 2003.
Ref Type: Abstract

Gage, L., Schmitt, D. Dystocia in an African elephant (Loxodonta africana). Proc Amer Assoc Zoo Vet.  88. 2003.
Ref Type: Conference Proceeding
Abstract: A 24-yr-old African elephant (Loxodonta africana) was inseminated on 12/2/00 and again on 12/4/00.  Pregnancy was confirmed on 1/21/01 with an ultrasound evaluation, and elevated progesterone levels of over a twelve to sixteen week period supported the diagnosis. The pregnancy progressed without complication.  Ultrasounds confirmed calf growth and movement throughout the pregnancy.  On 10/5/02, 673 days post-insemination, the cow passed the mucous plug in the evening.  The chorio-allantois ruptured shortly after that.  There were a few modest contractions over a period of about three hours, which then ceased.  Fetal front feet could be palpated in the birth canal, just into the cranial pelvis. Fetal circulation was confirmed using color doppler ultrasound of the fetal extremities.  Oxytocin was administered on 10/6/02.  The first two doses resulted in a few good contractions.  Fetal feet entered further into the pelvis, and fetal viability was confirmed again with color doppler ultrasound. Oxytocin was administered three more times resulting in a few small contractions and some stretching.  One dose of 110 IU Oxytocin resulted in one large contraction, subsequent doses of oxytocin did not yield any appreciable additional efforts.  That evening while walking the elephant, the amniotic sac broke releasing several liters of fluid. Blood was drawn and serum chemistries were within normal range. On 10/7/02 the uterus did not respond to oxytocin. Color doppler ultrasound of the fetal legs revealed no discernable fetal circulation.  Antibiotics and supportive care were initiated.  For the next week the feet were easily palpated within the pelvis.  For the several days the elephant was stiff and moderately depressed. Her appetite was good and she was drinking water. The elephant was given another dose of oxytocin on 10/15/02, which resulted in a few moderate contractions, but no progress was made in delivering the calf.  The next day the elephant seemed more depressed and uncomfortable. Her appetite and water intake were markedly reduced and within two days she exhibited what appeared to be abdominal pain. The elephant was treated aggressively with antibiotics and fluids for several weeks.  Severe peritonitis was confirmed during a laparoscopic procedure and the elephant was euthanized. The post-mortem examination revealed a tear in the uterus and a normal-sized fetus in normal presentation in the birth canal. The fetus however was abnormal and had arthrogryposis affecting all four limbs to varying degrees.  Both rear limbs of the fetus were particularly affected and the joints of those limbs were bent in an abnormal way, anchoring the fetus inside the dam. The arthrogryposis of the calf was likely the cause of the dystocia, and was likely the contributing factor to the tear in the uterus.

Ganswindt, A., Palme, R., Heistermann, M., Borragan, S., Hodges, J.K., 2003. Non-invasive assessment of adrenocortical function in the male African elephant (Loxodonta africana) and its relation to musth. Gen Comp Endocrinol 134, 156-166.
Abstract: German Primate Centre, Department of Reproductive Biology, Kellnerweg 4, 37077 Gottingen, Germany. ganswindt@www.dpz.gdwg.de

Adult male elephants periodically show the phenomenon of musth, a condition associated with increased aggressiveness, restlessness, significant weight reduction and markedly elevated androgen levels. It has been suggested that musth-related behaviours are costly and that therefore musth may represent a form of physiological stress. In order to provide data on this largely unanswered question, the first aim of this study was to evaluate different assays for non-invasive assessment of adrenocortical function in the male African elephant by (i) characterizing the metabolism and excretion of [3H]cortisol (3H-C) and [14C]testosterone (14C-T) and (ii) using this information to evaluate the specificity of four antibodies for determination of excreted cortisol metabolites, particularly with respect to possible cross-reactions with androgen metabolites, and to assess their biological validity using an ACTH challenge test. Based on the methodology established, the second objective was to provide data on fecal cortisol metabolite concentrations in bulls during the musth and non-musth condition. 3H-C (1 mCi) and 14C-T (100 microCi) were injected simultaneously into a 16 year old male and all urine and feces collected for 30 and 86 h, respectively. The majority (82%) of cortisol metabolites was excreted into the urine, whereas testosterone metabolites were mainly (57%) excreted into the feces. Almost all radioactive metabolites recovered from urine were conjugated (86% 3H-C and 97% 14C-T). In contrast, 86% and >99% of the 3H-C and 14C-T metabolites recovered from feces consisted of unconjugated forms. HPLC separations indicated the presence of various metabolites of cortisol in both urine and feces, with cortisol being abundant in hydrolysed urine, but virtually absent in feces. Although all antibodies measured substantial amounts of immunoreactivity after HPLC separation of peak radioactive samples and detected an increase in glucocorticoid output following the ACTH challenge, only two (in feces against 3alpha,11-oxo-cortisol metabolites, measured by an 11-oxo-etiocholanolone-EIA and in urine against cortisol, measured by a cortisol-EIA) did not show substantial cross-reactivity with excreted 14C-T metabolites and could provide an acceptable degree of specificity for reliable assessment of glucocorticoid output from urine and feces. Based on these findings, concentrations of immunoreactive 3alpha,11-oxo-cortisol metabolites were determined in weekly fecal samples collected from four adult bulls over periods of 11-20 months to examine whether musth is associated with increased adrenal activity. Results showed that in each male levels of these cortisol metabolites were not elevated during periods of musth, suggesting that in the African elephant musth is generally not associated with marked elevations in glucocorticoid output. Given the complex nature of musth and the variety of factors that are likely to influence its manifestation, it is clear, however, that further studies, particularly on free-ranging animals, are needed before a possible relationship between musth and adrenal function can be resolved. This study also clearly illustrates the potential problems associated with cross-reacting metabolites of gonadal steroids in EIAs measuring glucocorticoid metabolites. This has to be taken into account when selecting assays and interpreting results of glucocorticoid metabolite analysis, not only for studies in the elephant but also in other species.

Hama, N., Yamada, A., Noda, A., Murata, K., Shimada, Y., Ashida, M., Ishikawa, K., Matsuo, Y., Okuno, K., 2003. Serum hormonal changes in a female Asian elephant (Elephas maximus) with stillbirth. Japanese Journal of Zoo and Wildlife Medicine 8, 109-113.
Abstract: A case of stillbirth by an 11-year-old Asian elephant on January 11, 2002 is reported. Swelling of the abdominal region as well as the nipples in 2000 and 2001 had been previously observed. Serum progesterone (P), estradiol (E2) and prolactin (PRL) were measured monthly from May 2000 to January 2002 using radioimmunoassay (RIA). And P and E2 were additionally measured weekly from September 4, 1999 to January 12, 2002 using enzyme immunoassay (EIA). On the basis of EIA, elevated P levels were observed during early April 2000, and remained high until the end of 2001. Serum P concentrations gradually began to decrease by December 2001; at 6 days before stillbirth, there was a sudden drop in the level of blood P. On the day that the stillbirth occurred, the P value decreased to the level first recorded prior to April 2000. E2, based on results of EIA, did not display the regular cyclic trend prior to the elevation of serum P for pregnancy. The recorded values of serum P by both EIA and RIA were positively correlated (Pearson's correlative coefficient: 0.763, P < 0.01). On the basis of RIA, serum E2 and PRL were almost below detectable levels (E2: 8.0 pg/ml, PRL:0.5 ng/ml). The period of pregnancy estimated from the change of serum P concentration by EIA was 640 days.

Hermes, R., Arav, A., Saragusty, J., Goeritz, F., Pettit, M., Blottner, S., Flach, E., Eshkar, G., Boardman, W., Hildebrandt, T.B. Cryopreservation of Asian elephant spermatozoa using directional freezing. Proc.Amer Assoc of Zoo Veterinarians.  264. 2003.
Ref Type: Conference Proceeding
Abstract: Male infertility and absence of males in a facility are contributing factors to the limited reproduction of Asian elephants in captivity. Subsequent transport for breeding purposes increase social stress, risks of disease transmission and management costs. Recent success in artificial insemination eliminated these obstacles only transporting the semen. However, the transport of fresh semen involves logistical<bold> </bold>difficulties: access to semen donors, consistent semen quality and preservation of the spermatozoa during transport. The use of cryo-preserved sperm for AI can partially overcome these problems and can additionally be used for the establishment of Genome Resource Banks. However, to date, attempts to cryo-preserve Asian elephant spermatozoa have failed due to its sensitivity to freezing. Aims of this study were to identify the temperature range during which spermatozoa is most sensitive to chilling injury, and to use directional freezing (DF) to reduce cell damage during the freezing process. Semen was collected from two Asian elephants by manual stimulation. DF was used for freezing sperm samples. In contrast to conventional freezing methods DF facilitated a fast cooling rate, controlled ice crystal formation and cryopreservation of large volumes. Samples extended with a variety of DMSO extenders showed post thaw motility of 30-40%. DF was able to cryo-preserve Asian Elephant spermatozoa for the first time. As DF seems to reduce cryo injury it may become of interest to optimize existing cryopreservation protocols of other endangered species, or to make cryopreservation even possible in species with cryo-sensitive spermatozoa.

Hildebrandt, T.B., Strike, T., Flach, E., Sambrook, B.S., Dodds, J., Lindsay, N., Goeritz, F., Hermes, R., McGowan, M. Fetotomy in the elephant. Proc Amer Assoc Zoo Vet.  89-92. 2003.
Ref Type: Conference Proceeding
Abstract: There were several reports about dystocia and its treatment in elephants since they were kept in captivity in western zoos and safari parks. ^{1, 2, 3, 4, 5, 6, 7, 8, 9, 10}  Especially in the last two years there has been a dramatic accumulation of reports about birth associated problems (Tab. 1). Causes for this development are probably the intensified captive elephant breeding programs and the involvement of more older nulliparous cows.

Over the last 100 years, there were more dystocia cases in captive Asian elephants than in African elephants. ⁷  However, this difference seems to be abolished with enhanced number of pregnancies in African elephants by now. The fact of an increased percentage of dystocia cases in both species can have two consequences for captive management. Firstly, older nulliparous cows will be strictly excluded from breeding in the future or secondly, the birth management will be improved.

Three (Table 1, Em 4, 5; La 3) of 10 cows died from the consequences of dystocia in western zoological institutions over the last two years. Totally, there were eleven cases of fatal dystocia cases listed in the literature since 1972 (Table 2). ^{5, 6, 7}  The seven cesarian performed (Table 2) as the ultimate intervention to treat the dystocia in elephants ended all with euthanasia or death of the females. The following presentation will described a potential new way of birth management in the elephant, the fetotomy. The authors believe that the fetotomy is prospective tool instead of the unsuccessful cesarian for saving the life of the dam with severe dystocia.  This method is recommended by the authors as an ultimate tool besides all the other important preparations in pregnant cows for an upcoming birth. Most important classical preparations are following: (I) intensive physical exercise, (ii) optimization of the body weight with a pregnancy associated diet, (iii) training for safe handling in case of necessary intervention in free or protected contact.  The fetotomy was never applied before in elephants because of the limited exploration field and the high degree of skeleton calcification in comparison to domestic and exotic hoofstock. 

In this particular case it was firstly tried to remove the dead fetus by episiotomy. ^{2, 3, 4, 7, 9, 10} However, the episiotomy attempt failed due to he inability to extract a dead oversized or malpositioned fetus.  From the two options to leave the dead fetus inside the uterus despite the surgically opened genital tract or to cut out parts of the fetus it was decided to perform a fetotomy. The initial hesitation to perform such drastic and complicated procedure led to the complication of an emphysemic fetus and progressive peritonitis in the female even before the procedure started. Fetus was successfully removed in six parts using an extra long and durable carthorse embryotom (Thygesen's type). Unfortunately, the patient died 3 days later because of the treatment resistant peritonitis.

In conclusion, it was demonstrated that fetotomy can successfully be applied in elephants. The authors suggested in order to achieve a positive outcome for the dam to decide on such ultimate intervention earlier in the dystocia process. Active elephant birth management should always be prepared for dystocia treatment ranging from (I) the administration of oxytocin, (ii) rectal massage of the caudal birth channel, (iii) episiotomy to the last option (iv) of fetotomy to safe the life of the mother.  Based on the complex preparation for an elephant birth and the knowledge about the huge range of potential complications it is advised to consult veterinary elephant specialists well in advance to ensure maximum expertise and experience.

Acknowledgements
The authors like to thank Charlie Gray who helped us to collect the relevant data from the dystocia cases and the elephant staff from Whipsnade Wild Animal Park for their active support. We also like to acknowledge professor Peter Glatzel, DVM and Andrea Krause for helping with the logistic preparation.

Literature cited

1.Lang, E.M. 1963. Geburtshilfe bei einem Indischen Elefanten. Acta Trop. 20, 87-114.
2.Merkt, H., D., Ahlers, H., Bader, H.-P., Brandt, M., Boer and L. Dittrich. 1985a. Bildbericht über den Auszug eines toten Elefantenfetus (Elephas maximus) am 645. Tag p.c. und 65 Stunden nach Geburtsbeginn via Damschnitt (vorläufige Mitteilung). Praktischer Tierarzt 5: 377-378.
3.Merkt, H., D., Ahlers, H., Bader, H.-P., Brandt, M., Boer and L. Dittrich. 1985b. Der Damschnitt, eine geburtshilfliche Interventionsmöglichkeit bei einer Elefantenkuh. Deutsche tieraerztliche Wochenschrift 92: 428-432.
4.Merkt, H., D., Ahlers, H., Bader, D., Rath, H.-P., Brandt, M., Boer and L. Dittrich. 1986. Nachbehandlung und Heilungsverlauf bei einer Elefantenkuh nach Geburtshilfe durch Damschnitt. Berliner Münchner Tierärztliche Wschrift 99: 329-333.
5.Furley, C.W. 1993. A caesarean section in an elephant (The first in europe). Help-Newsletter,15: 2931.
6. Foerner, J.J. 1998. Dystokia in the Elephant. In: Fowler, M. E. & E., Miller (eds.): Zoo & Wild Animal Medicine. 4. Ed. W. B. Saunders Company, 522-525.
7.Lange, A., T.B., Hildebrandt, G., Strauss, O., Czupalla, F., Goeritz and W. Schaftenaar. 1999. Möglichkeiten und Grenzen der Geburtshilfe bei Elefanten. Verhandlungsbericht Erkrankungen der Zootiere 39: 47-58.
8.Fluegger, M., F. Goeritz, R. Hermes, E. Isenbuegel, A. Klarenbeek, W. Schaftenaar, K. Schaller and G. Strauss. 2001. Evaluation of physiological data and veterinary medical experience in 31 Asian elephant (Elephas maximus) births in six European zoos. Verhandlungsbericht Erkrankungen der Zootiere, 40: 123-134.
9.Schaftenaar, W. 1996. Vaginal Vestibulotomy in an Asian Elephant (Elephas maximus). 1996 Proceedings American Association Zoo Veterinarians,  434-439.
10. Schaftenaar, W., T.B., Hildebrandt, M., Fluegger, F., Goeritz, D.J., Schmidt, and G. West. 2001. Guidelines for veterinary assistance during the reproductive process in female elephants. Proceedings American Association Zoo Veterinarians, 348-355.

Note: See source for Table 1: Dystocia cases in captive elephants in the last 2 yr and Table 2: Known dystocia cases with a fatal oucome for the dam

Hildebrandt, T.B., Strike, T., Flach, E., Sambrook, L., Dodds, J., Lindsay, N., Furley, C.F., Glatzel, P.S., McGowan, M., Wisser, J.ed., Hofer, H.e., Frolich, K. Fetotomy in the elephant. Erkrankungen der Zootiere.  315-318. 2003.
Ref Type: Conference Proceeding

Hunt, K.E., Wasser, S.K., 2003. Effect of long-term preservation methods on fecal glucocorticoid concentrations of grizzly bear and african elephant. Physiol Biochem Zool. 76, 918-928.
Abstract: Center for Conservation Biology, Department of Biology, University of Washington, Seattle, WA, 98195-1800, USA.

Leong, K.M., Ortolani, A., Graham, L.H., Savage, A., 2003. The use of low-frequency vocalizations in African elephant (Loxodonta africana) reproductive strategies. Horm Behav. 43, 433-443.
Abstract: Fertility-advertisement calls in females are predicted to occur in nonmonogamous species where males and females are widely separated in space. In African elephants, low-frequency vocalizations have thus been suggested as a reproductive strategy used by fertile females to attract mates. This study examined the use of low-frequency vocalizations with respect to different phases of the estrous cycle in African elephants by simultaneously monitoring vocalizations, behavior, and hormonal profiles. Subjects were one male and six female African elephants housed at Disney's Animal Kingdom. No acoustically distinct vocalizations were restricted to the ovulatory follicular phase. However, overall rate of low-frequency vocalization as well as the rate of one acoustically distinct vocalization changed over the estrous cycle, with highest rates of calling related to the first period of follicular growth, or anovulatory follicular phase. Elevated rates of vocalization thus were not restricted to behavioral estrus and occurred much earlier in the estrous cycle than in most species that produce fertility-advertisement calls. Both herd composition and elephant identity also affected rates of vocalization. Vocalizations therefore may not be reliable signals of actual fertility. However, the increase in vocalizations in advance of estrus may attract males to the herd prior to ovulation, facilitating both male-male competition and female choice. Once present in the herd, males may then switch strategies to use more reliable chemical and visual cues to detect ovulating females. Disney's Animal Kingdom, Lake Buena Vista, FL 32830, USA. Kristen.Leong@disney.com

Malhotra, A.K., Kumar, M., 2003. Management of musth Indian elephant at National Zoological Park, New Delhi. Zoos' Print Journal 18, 10.

Nath, K.C., 2003. Oestrous cycle and pregnancy in elephant. In: Das, D. (Ed.), Healthcare, Breeding and Management of Asian Elephants. Project Elephant. Govt. of India, New Delhi, pp. 42-44.

Pucher, H.E., Stremme, C., Schwarzenberger, F., 2003. Priapism in a semiwild Asian elephant (Elephas maximus) in Vietnam. Vet Rec 153, 717-718.

Rajaram, A., Krishnamurthy, V., 2003. Elephant temporal gland ultrastructure and androgen secretion during musth. Current Science 85, 1467-1471.
Abstract: We have investigated the ultrastructure of the temporal gland of the Asian elephant (Elephas maximus) in the musth condition. We find that the organelles are highly evolved for the production of the androgen, testosterone which is reported to be very high in the Asian male elephant in full musth. The mitochondria bear cristae which are profuse and tubular, and occur along with many Golgi bodies. There is hypertrophy of smooth endoplasmic reticulum. All the structures involved in the production of androgen, as in the Leydig cell or the cells of the adrenal cortex, are thus found in abundance. Cellular structures also seem singularly evolved for the secretion of androgen and its degradation products.

Rasmussen, B., 2003. Why musth elephants use pheromones? Biologist 50, 195-196.

Rasmussen, L.E., Greenwood, D.R., 2003. Frontalin: a chemical message of musth in Asian elephants (Elephas maximus). Chemical Senses 28, 433-446.
Abstract: Musth is an important male phenomenon affecting many aspects of elephant society including reproduction. During musth, the temporal gland secretions (as well as the urine and breath) of adult male Asian elephants (Elephas maximus) discharge a variety of malodorous compounds together with the bicyclic ketal, frontalin. In contrast, teenage male elephants in musth release a sweet-smelling exudate from their facial temporal gland. We recently demonstrated that the concentration of frontalin becomes increasingly evident as male elephants mature. In the present study, we demonstrate that behaviors exhibited towards frontalin are consistent and dependent on the sex, developmental stage and physiological status of the responding conspecific individual. To examine whether frontalin functions as a chemical signal, perhaps even a pheromone, we bioassayed older and younger adult males, and luteal- and follicular-phase and pregnant females for their chemosensory and behavioral responses to frontalin. Adult males were mostly indifferent to frontalin, whereas subadult males were highly reactive, often exhibiting repulsion or avoidance. Female chemosensory responses to frontalin varied with hormonal state. Females in the luteal phase demonstrated low frequencies of responses, whereas pregnant females responded significantly more frequently, with varied types of responses including those to the palatal pits. Females in the follicular phase were the most responsive and often demonstrated mating-related behaviors subsequent to high chemosensory responses to frontalin. Our evidence strongly suggests that frontalin, a well-studied pheromone in insects, also functions as a pheromone in the Asian elephant: it exhibits all of the determinants that define a pheromone and evidently conveys some of the messages underlying the phenomenon of musth. Department of Biochemistry and Molecular Biology, OGI School of Science and Engineering, Oregon Health & Science University, Beaverton, OR 97006-8921, USA. betsr@bmb.ogi.edu

Rees, P.A., 2003. The welfare and conservation of Asian elephants – a reply to Sukumar. Oryx 37, 25.
Abstract: Since my summary of the global fate of Asian elephants in zoos (this issue) was written Clubb & Mason (2002) have published a review of the welfare of zoo elephants in Europe, commissioned by the Royal Society for the Prevention of Cruelty to Animals in the UK. In an attempt to collect data on behaviour, reproduction, group composition, welfare and other aspects of husbandry, they sent questionnaires to the directors of the 18 zoos in the UK that hold elephants. Professor Sukumar doubts my contention that zoo directors lack the commitment necessary to manage the zoo elephant population as viable breeding units. Why then did none of the zoos contacted by Clubb & Mason reply?

Sarma, K.K., 2003. Managing troublesome bulls with special reference to musth in captive Asian elephants. In: Das, D. (Ed.), Healthcare, Breeding and Management of Asian Elephants. Project Elephant. Govt. of India, New Delhi, pp. 58-66.

Schmitt, D.L., 2003. Proboscidea (Elephants). In: Fowler, M.E., Miller, R.E. (Eds.), Zoo and Wild Animal Medicine. Elsevier Science USA, pp. 541-550.

Slade, B.E., Schulte, B.A., Rasmussen, L.E.L., 2003. Oestrous state dynamics in chemical communication by captive female Asian elephants. Animal Behaviour 65, 813-819.
Abstract: In many mammals, reproductive status is revealed through chemical cues in urine. The reproductive status of receivers may influence their interest in such signals. For social mammals that live in matrilineal groups, females may benefit by detecting the reproductive condition of herdmates. Responses to urine during oestrous cycles of senders and receivers are potential indicators of signal functions. We examined the chemosensory responses, first by four captive female Asian elephants, Elephas maximus, over their oestrous cycles to familiar follicular and luteal phase urine and second by 14 different female Asian elephants to unfamiliar conspecific follicular and luteal phase urine. We asked whether females could distinguish the reproductive state of another female as measured by their differential response to luteal-and follicular-phase urine. We further examined the influence of the receiver's reproductive status on response levels. Females responded more with specific tactolfactory trunk behaviours to follicular- than to luteal-phase urine, but only when the receiving female was in her follicular phase. Like their male conspecifics, Asian elephant females can detect changes in the reproductive state of conspecifics. The functional significance of this ability has yet to be determined but may be related more to the resource holding power of females in follicular phase than to a means for females to synchronize oestrous cycles. Such female-female communication may have important effects on social group dynamics.

Sleeman, J.M., Clyde, V.L., Finnegan, M.V., Ramsay, E.C., Shires, M.G., 2003. Mammary botryomycosis and mastectomy in an African elephant (Loxodonta africana). Vet Rec 152, 54-55.

Teng, M.S., Yang, X.L., Wu, D.H., 2003. Characteristics of reproductive biology of Asian elephants. Chinese Journal of Zoology 38, 86-90.
Abstract: The biological characteristics of 1 female Asian elephant that had produced 2 fetuses consecutively were studied through whole-day observation and recording of oestrus, mating, gestation, parturition, postpartum behaviour, and the behaviour of the 2 young elephants (China). Daily frequencies of the young elephant's sucking the breast of the mother reduced day by day. There were differences in the characteristics of different fetuses, including gestation omen, gestation period, defaecation of the young elephant, etc.

West, J.B., Fu, Z., Gaeth, A.P., Short, R.V., 2003. Fetal lung development in the elephant reflects the adaptations required for snorkeling in adult life. Respir Physiol Neurobiol 138, 325-333.
Abstract: The adult elephant is unique among mammals in that the pleural membranes are thickened and the pleural cavity is obliterated by connective tissue. It has been suggested that this peculiar anatomy developed because the animal can snorkel at depth, and this behavior subjects the microvessels in the parietal pleura to a very large transmural pressure. To investigate the development of the parietal pleura, the thickness of the endothoracic fascia (ET) was measured in four fetal African elephants of approximate gestational age 111-130 days, and the appearances were compared with those in human, rabbit, rat and mouse fetuses of approximately the same stage of lung organogenesis. The mean thicknesses of ET in the elephant, human, rabbit, rat and mouse were 403, 53, 29, 27 and 37 microm, respectively. This very early development of a thick parietal pleura in the elephant fetus is consistent with the hypothesis of a long history of snorkeling in the elephant's putative aquatic ancestors. Department of Medicine, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0623, USA. jwest@ucsd.edu

Whitehouse, A.M., Schoeman, D.S., 2003. Ranging behaviour of elephants within a small, fenced area in Addo Elephant National Park, South Africa. African Zoology 38, 95-108.
Abstract: The elephant population (n = 324, December 2000) in Addo Elephant National Park (AENP), South Africa, is restricted to an area (103 km(2)) considerably smaller than most elephant ranges. The Addo elephants' ranging behaviour was studied in order to determine whether natural patterns of male and female ranging behaviour can be maintained within a confined area. Radio-tracking was used to facilitate measurements of hourly distances travelled, larger scale directional movements, home range sizes, centres of activity and associations. Female home ranges overlap, and interactions between family groups are frequently observed. Females and non-musth males travel similar distances and speeds on an hourly basis, but female core range sizes are larger, resumably because bigger areas are needed to satisfy the nutritional requirements of their young. contrary to expectations, Addo males do not travel further and faster on an hourly basis when they are in musth in comparison to when they are not in musth. However, the movement of males in musth is more directional, so that they cover a greater area of the park. Males associate more frequently with females when in musth than when non-musth. Unlike in other populations, non-musth 'male retirement areas' in AENP are not exclusive, as there is spatial and temporal overlap with female ranges, and hence with the ranges of males in musth. It is suggested that the resulting increased frequency of contact between musth and non-musth males may result in elevated mate competition, and this could contribute to the high levels of aggression observed between Addo bulls.

Wilson, J.D., Leihy, M.W., Shaw, G., Renfree, M.B., 2003. Androgen physiology: unsolved problems at the millennium. Molecular and Cellular Endocrinology 198, 1-5.
Abstract: Androgen physiology differs from that of other steroid hormones in two major regards. First, testosterone, the predominant circulating testicular androgen, is both an active hormone and a prohormone for the formation of a more active androgen, the 5alpha-reduced steroid dihydrotestosterone. Genetic evidence indicates that testosterone and dihydrotestosterone work via a common intracellular receptor, and studies involving in vitro reporter gene assays and intact mice in which both steroid 5alpha-reductase isoenzymes have been disrupted by homologous recombination indicate that dihydrotestosterone acts during embryonic life to amplify hormonal signals that can be mediated by testosterone at higher concentrations. However, in post-embryonic life dihydrotestosterone plays unique roles that have not been elucidated. Studies of other 5alpha-reduced steroids, including the plant hormone brassinolide, the hog pheromones androstanol and androstenol, and 5alpha-dihydroprogesterone (in horses and elephants) indicate that this reaction serves different functions in different systems. Second, during embryonic life androgen causes the formation of the male urogenital tract and hence is responsible for development of the tissues that serve as the major sites of androgen action in postnatal life. It has been generally assumed that androgens virilize the male fetus by the same mechanisms as in the adult, namely by the conversion of circulating testosterone to dihydrotestosterone in target tissues. However, in marsupial mammals there is no sexual dimorphism in the levels of testosterone or dihydrotestosterone at the time the male phenotype forms, and in the pouch young of one marsupial, the tammar wallaby, the testes secrete another 5alpha-reduced steroid, 5alpha-androstane-3alpha, 17beta-diol (5alpha-adiol), into plasma. The administration of 5alpha-adiol to female pouch young causes profound virilization of the urogenital sinus and external genitalia, but within target tissues 5alpha-adiol appears to work after oxidation to dihydrotestosterone. Thus, two separate mechanisms evolved for the formation of dihydrotestosterone in target tissues. 5alpha-adiol is the predominant androgen in neonatal testes in several placental mammals, but it is unclear whether it plays a similar role in other mammalian species.

Wingfield, J.C., Sapolsky, R.M., 2003. Reproduction and resistance to stress: When and how. Journal of Neuroendocrinology 15, 711-724.

Zuba, J.R., Stetter, M.D., Dover, S.R., Briggs, M. Development of rigid laparoscopy techniques in elephants and rhinoceros. Proc Amer Assoc Zoo Vet.  223-227. 2003.
Ref Type: Conference Proceeding
Abstract: Diagnostic and surgical laparoscopy has become a routine procedure in human and veterinary medicine and has similar potential uses in zoological medicine. Surgical telescopes and fiberoptic cables allow the veterinarian to look inside body cavities of the patient and specialized instruments provide the ability to perform a wide variety of diagnostic and surgical procedures.  Rigid laparoscopy is considered minimally invasive surgery and is associated with a more rapid post-operative recovery rate and an improved prognosis as compared to conventional surgical techniques.³  Laparoscopic surgery is commonly being utilized in horses and other zoo animals for a variety of abdominal surgical procedures including tubal ligation and ovariectomy.^7,8,12  Recent advances in technology now provide the ability to perform laparoscopy in the largest mammalian species maintained in zoological collections.¹¹

A variety of disease problems and reproductive disorders have been documented in the rhinoceros and elephant.  Due to their size and anatomy, many of the standard diagnostic tests available in human and veterinary medicine are not routine in these animals (radiography, advanced imaging techniques, liver biopsy, abdominal tap, etc.) at this time.  Although conventional abdominal surgery has been performed in rhinoceros and elephants^1,5,10 survival rates have been extremely low.  Laparoscopy can provide a variety of additional diagnostic options, and may also provide an avenue for performing surgical procedures that would otherwise be impossible in these animals.

A multi-institutional collaboration has been organized to address common goals with regard to enhancing diagnostic capabilities and improving surgical techniques in elephants and rhinoceros.  Specifically, we are aiming to: Develop laparoscopic techniques and equipment for use in rhinoceros and elephants, which can be used to significantly expand our diagnostic and treatment capabilities in these large mammals. Improve international conservation efforts in both rhinoceros and elephants by helping improve animal health and welfare of these species in captive and free ranging situations.

This collaborative effort has been organized to critically review current equipment, techniques and uses with the ultimate goal of overcoming some of the inherent difficulties with laparoscopy in these large vertebrates.  This includes further development and modification of equipment, investigation of surgical techniques, and expansion of clinical applications.

Cooperation in the development of innovative surgical equipment for the anatomic variety of our zoological species is necessary for the advancement of zoological medicine.  Karl Storz Veterinary Endoscopy of America (KSVEA, Goleta, California 93117, USA) was instrumental in the initial development of this specialized equipment at the San Diego Wild Animal Park and by generously providing research and development for our continuing investigations.  The availability of appropriately sized equipment has limited the application of laparoscopic techniques to animals of a size consistent with the intended species of its production.  The largest laparoscopy equipment commercially available is marketed for use in equine medicine (57 cm telescope and accessories).  This system has been used with limited success in rhinoceros¹¹ and elephants.  In many cases, to adequately visualize and manipulate visceral organs, a longer telescope and associated instrumentation is required.  We continue to work closely with KSVEA on the production of specialized equipment (Table 1).  Standard light source, fiberoptic light cable and electronic insufflator units manufactured for use in domestic large animals have been used successfully in megavertebrates.  Due to its compact size and versatility, the authors suggest the use of a portable, battery operated laparoscopy kit (Techno Pack, KSVEA) (monitor, light source, camera and digital recording device) especially under field conditions.

To date we have evaluated the utility of specially designed laparoscopic equipment in approximately seven elephants and four rhinoceros.  Experiences from these limited cases have identified several technical and procedural challenges, which need to be overcome if megavertebrate laparoscopy is to be successfully performed.¹¹  Some of these challenges are summarized below:

Technical challenges:
Equine laparoscopic equipment too fragile and/or short for certain surgical applications in rhinoceros and elephant
Size and disposition of megavertebrates (> 1000 kg)
Thick, non-pliable, pachydermatous skin puts unusual pressure (and risk of damage) on equipment
Insufflation and illumination of large abdominal cavity
Great depth to visualize and physically reach/manipulate large organs
Thick, redundant, fibro-elastic peritoneum which is difficult to puncture during surgical attempts to enter the abdominal cavity
Influence of patient positioning:  unable to use conventional laparoscopic positioning techniques in these species
Both rhinoceros and elephants are hindgut fermentors and have very large and extensive lower intestinal tracts.  Gas dilation of these bowel loops combined with limited positioning options, can make laparoscopic visualization of certain organs a problem

Procedural challenges:
Cost of developing new and specialized laparoscopic equipment is high; duplicate equipment is not available at this time
Charismatic nature of megavertebrates makes it difficult to perform surgery on such important and high exposure species 
Limited clinical cases for testing equipment due to the relatively low number of megavertebrates in zoological facilities
Use of laparoscopy in zoo mammals may be out of the comfort zone for many veterinarians and curators due to lack of experience with this instrumentation
Lack of published surgical procedures in these species
Inability to perform laparoscopic procedure in "surgical suite" as with most other species
Risks associated with megavertebrate anesthesia and sedation including difficulty in providing safe anesthetic procedures, proper restraint, and safety of personnel

A multidisciplinary, systematic approach has been initiated to critically review current instrumentation and procedures with the objective of overcoming these technical difficulties. 

The initial phase of our project has been to develop laparoscopic techniques and equipment (Table 1) for use in rhinoceros and elephants by utilizing individuals that may have died of natural causes, or live animals with medical conditions that warrant abdominal surgery.  We have already had the opportunity to perform laparoscopic surgery on both live and deceased white rhinoceros, black rhinoceros, African and Asian elephants.  From our initial studies, it is apparent that laparoscopy in these megavertebrates is possible and may greatly enhance our ability to care for and manage these animals in captive and free ranging situations.  The authors have received some funding and institutional support that allows us to travel to institutions which may benefit from the use of this equipment and/or our experience.  We are also interested in those zoological institutions which may find themselves dealing with a terminal case in an elephant or rhinoceros, to please contact us directly and to consider allowing laparoscopy to be conducted on the animal prior to a post-mortem examination.

Ultimately, we envision the results of our studies on captive animals to be applicable to the management and conservation of elephants and rhinoceros in the wild.  A variety of medical disorders are commonly reported in black and white rhinoceros.  Many of these medical conditions are difficult to diagnose, monitor and treat.  Furthermore, there is a paucity of information on the incidence of these disease conditions in free ranging populations.  The use of minimally invasive laparoscopic techniques will greatly enhance our diagnostic abilities in this species and would be extremely valuable to the understanding of medical conditions of captive and free ranging rhinoceros and to conservation efforts overall.

Throughout many parts of Africa, wildlife professionals are seriously concerned about the negative effects large elephant herds are having on the native flora and fauna within parks and reserves.^2,6,9  The historical rangelands of the elephant have become interrupted by national borders and artificial barriers.  Habitats surrounding wildlife parks are increasingly being converted to agricultural lands. The encroachment of human populations has caused a dramatic increase in the number and severity of human-elephant conflicts.^2,9  These conflicts are commonplace in many parts of East and Southern Africa.  Although there have been a variety of plans to reduce human-elephant and elephant environment impacts, little overall success has been achieved in most countries.⁹

Historically elephant population control has primarily been limited to culling and translocation of small groups.^2,6  Immunocontraception has been attempted with a small population of elephants but is not currently realistic in many situations.^4,6  In wildlife parks where large herds of elephants exist there is currently no effective, humane method of population control.

One of our long-range goals is to develop laparoscopic techniques, such as ovariectomy and tubal ligation, in free ranging African elephants that can be used to sterilize reproductive females.  Once these techniques have been developed, it is our intention to train local wildlife veterinarians and health professionals to perform laparoscopic sterilization of elephants in the field and thus provide local wildlife officials with a tool to help manage elephant populations.  It is our hope this will improve conservation efforts across Africa by reducing human-elephant conflicts and helping to save critical ecosystems.

Acknowledgements
This ongoing project is possible due to the generous contributions of the following individuals:  Hans Lunneman, Christopher Chambliss, Michele McCutcheon, Lynn Richardson, Dean Hendrickson, Robin Radcliffe, Rolf Radcliffe, Laurie Gage, Larry Galuppo, Bill Lindsay, John Olsen, Genny Dumonceaux, and the veterinary staffs at the San Diego Wild Animal Park and Disney's Animal Programs.

LITERATURE CITED
1.Byron, H., J. Olsen, M. Schmidt, J. Copeland, and L. Byron. 1985. Abdominal surgery in three adult male Asian elephants. J Am Vet Med Assoc. 187:1236-1237.
2.Chalfota J. and Owen-Smith N. 1996. Options for the management of elephants in northern Botswana. Pachyderm. 22:67-73.
3.Cook, R.A., and D.R. Stoloff. 1999. The application of minimally invasive surgery for the diagnosis and treatment of captive wildlife. In: Fowler, M., Miller, E. (Eds): Zoo and Wild Animal Medicine: Current Therapy 4. W.B. Saunders, Philadelphia, PA. Pp 30-40.
4.Delsink, A.K., van Altena, J.J., Kirkpatrick, J. Grobler, D., and Fayrer-Hosken, R.A. 2002. Field applications of immunocontraception in African elephants (Loxodonta africana).  J Soc.Reprod. Fert. 60: 117-124.
5.Fowler, M.E., and R. Hart. 1973. Castration of an Asian elephant using etorphine anesthesia. J Am Vet Med Assoc. 163: 539-543.
6.Garai, M.E.  2001.  Managing elephants on private reserves in South Africa.  Proceedings of the International Elephant and Rhino Research Symposium. Vienna, Austria. 259-261.
7.Hendrickson, D.A. 2002. New techniques for performing equine laparoscopic ovariectomy. DVM Best Practices Magazine. Oct. 2002.
8.Hendrickson, D.A., and D.G. Wilson. 1996. Instrumentation and techniques for laparoscopic and thoracoscopic surgery in the horse. Vet. Clin. N.A. Equine Pract.12; 2: 235.
9.Hoare, R., Update on the study and management of human-elephant conflict in Africa. Pachyderm. 33: 91-92.
10.Olsen, J., and H. Byron. 1993. Castration of the elephant. In: Fowler, M. (Ed): Zoo and Wild Animal Medicine: Current Therapy 3, 3^rd ed. W.B. Saunders, Philadelphia, PA. Pp. 441-444.
11.Radcliffe R.M., D.A. Hendrickson, G.L. Richardson., J.R. Zuba, and R.W. Radcliffe. 2000. Standing laparoscopicguided uterine biopsy in a southern white rhinoceros (Ceratotherium simum simum). J. Zoo Wildl. Med. 31:201207.
12.Rogerson, D., M. Brown, B. Watt, C. Keoughan, and M. Hanrath. 2002. Hand-assisted laparoscopic technique for removal of ovarian tumors in standing mares. J. Am. Vet. Med. Assoc. 220(10):1503-1507.

See source for Table 1.  Current laparoscopy equipment specifically manufactured for use in megavertebrates.^a
a All equipment was specially manufactured by Karl Storz Veterinary Endoscopy of America (KSVEA, Goleta, California, USA) and is not commercially available at this time.

Journal of Indian Veterinary Assocaition Kerala. Journal of Indian Veterinary Association Kerala 7[3], 1-64. 2002.
Ref Type: Journal (Full)

Alex, P.C., 2002. The Musth, the vicious and the rogue elephants - a review. Journal of Indian Veterinary Association Kerala 7, 26-27.

Allen, W.R., Mathias, S.S., Wooding, F.B., Skidmore, J.A., van Aarde, R.J., 2002. Placentation in the African elephant, Loxodonta africana. I. Endocrinological aspects. Reprod Suppl 60, 105-116.
Abstract: Placental and fetal tissues were recovered from the uteri of 59 pregnant elephant that ranged in estimated age from day 18 to month 21 of gestation. Incubation of placenta and fetal gonad, alone or in combination, with tritium-labelled cholesterol, pregnenolone and androstenedione failed to yield any labelled progestagens or oestrogens from placenta, but did produce small amounts of labelled progesterone and 5alpha-dihydroprogesterone from fetal gonad. Immunochemical staining of tissues with four antisera specific for enzymes involved in the steroidogenic pathway revealed no staining in sections of placenta but positive labelling for P450 side chain cleavage enzyme (SCC450) and 3beta-hydroxysteroid dehydrogenase (3beta-HSD) of the interstitial cells that comprise the bulk of the enlarged fetal gonads during the second half of gestation. Saline extracts of placental tissue showed no activity in three different gonadotrophin assays. In view of this endocrinological inactivity in the zonary elephant placenta and the probable reliance on maternal luteal sources of progestagens for maintenance of the pregnant state, the argument is advanced that uncomplicated abortion would probably follow a single administration of a PGF analogue given at any stage of pregnancy. If so, the treatment might constitute an efficacious method for controlling population increases in elephants maintained in enclosed game parks in Africa.

Chandrasekharan, K., 2002. Elephant - an overview. Journal of Indian Veterinary Association Kerala 7, 8-11.

Cheeran, J.V., Radhakrishnan, K., Chandrasekharan, K., 2002. Musth. Journal of Indian Veterinary Association Kerala 7, 28-30.

Cheeran, J.V., Chandrasekharan, K., Radhakrishnan, K., 2002. Tranquilization and translocation of elephants. Journal of Indian Veterinary Association Kerala 7, 42-46.

Cheeran, J.V., 2002. Elephant facts. Journal of Indian Veterinary Association Kerala 7, 12-14.

Davis, S.A., Pech, R.P., 2002. Dependence of population response to fertility control on the survival of sterile animals and their role in regulation. Reprod Suppl 60, 89-103.
Abstract: The species for which fertility control is presently used, or for which it is being developed, range from small mammal pests, such as the house mouse (Mus domesticus), to large mammals, such as the African elephant (Loxodonta africana). However, the possibility of a population response other than a reduction in abundance proportional to the fraction of animals rendered infertile has been shown in field trials. For example, when intermediate levels of sterility were imposed on wild populations of European rabbits (Oryctolagus cuniculus), there was an increase in their abundance, on an annual basis, due to enhanced survival of juveniles and adult females. In this article, we relate intraspecific regulatory processes to the response of populations to fertility control using a set of density-dependent structured-population models. In each of the models, the population is exposed periodically to a fertility control agent that renders a fraction of fertile females sterile.  Although our intention is not to predict the population response of any one particular species, the results of the models are illustrated using parameter values that are representative of populations of the European fox (Vulpes vulpes) in south-eastern Australia. When populations were regulated by density-dependent mechanisms in which sterile females did not participate, such as competition for resources among young animals or competition among fertile females for breeding sites or territories, then populations could increase in abundance for low and intermediate levels of imposed sterility. For other intraspecific regulatory mechanisms, such as competition for resources between all individuals, all levels of sterility were observed to reduce abundance. The population response was sensitive to (i) whether the survival of sterile adults was higher than that of fertile adults, (ii) whether animals could be sterilized before sexual maturity, and (iii) whether density dependence was modelled as a threshold process.

Delsink, A.K., van Altena, J.J., Kirkpatrick, J., Grobler, D., Fayrer-Hosken, R.A., 2002. Field applications of immunocontraception in African elephants (Loxodonta africana). Reprod Suppl 60, 117-124.
Abstract: The primary aim of the Makalali elephant immunocontraception programme is to test the efficacy of porcine zona pellucida (PZP) vaccine for practical population control of elephants in small, enclosed reserves, with the goal of stabilizing the current growth rate and reducing it to the 5-10% per annum displayed currently in the Kruger National Park. A secondary aim is to test the hypothesis that PZP treatment does not affect patterns of elephant social behaviour. Eighteen sexually mature cows (age > 12 years) were vaccinated in May 2000 using remote darts. Behavioural observations before, during and after vaccination included noting the activity of individual animals every minute for 15 min. No changes in general behaviour patterns have been noted to date although the animals' spatial use of the reserve was erratic during the period of vaccination, indicating irregular or disturbed patterns associated with vaccination. Normality was resumed on completion of the vaccinations. No aggressive or indifferent behaviour related to nursing, calf proximity or bull-cow interactions have been noted. Ten of the females were in various stages of pregnancy when treated. Subsequently, seven of them gave birth to healthy calves and the other three females are expected to calve shortly. It is too early in the study to draw conclusions about stabilization of growth rates.

Duer, C., Carden, M., Schmitt, D., Tomasi, T., 2002. Utility of maternal serum total testosterone analysis for fetal gender determination in Asian elephants (Elephas maximus). Anim Reprod Sci 69, 47-52.
Abstract: It has been shown in some species that fetal testes produce testosterone early in gestation. This study investigated the possibility that fetal testosterone may be reflected in maternal serum levels in the Asian elephant (Elephas maximus). Weekly serum samples were collected from seventeen pregnant captive Asian elephants and analyzed via radioimmunoassay (RIA) for total testosterone levels. Nine of the cows carried male fetuses and eight carried female fetuses. A non-random pattern over time (P<0.01) was observed in cows carrying either a male or female fetus. Mean maternal serum total testosterone was significantly higher in cows carrying male versus female fetuses (P<0.01). Mean trimester values indicate that first trimester values are not significantly different among male versus female groups. The second and third trimester values of cows carrying male fetuses were higher than cows carrying female fetuses, (P<0.01 and <0.05, respectively). The results of this study show that it is possible via RIA of maternal serum for total testosterone to determine the gender of calves during gestation.

Forsyth, I.A., Wallis, M., 2002. Growth hormone and prolactin--molecular and functional evolution. J Mammary Gland Biol Neoplasia 7, 291-312.
Abstract: Growth hormone, prolactin, the fish hormone, somatolactin, and related mammalian placental hormones, including placental lactogen, form a family of polypeptide hormones that share a common tertiary structure. They produce their biological effects by interacting with and dimerizing specific single transmembrane-domain receptors. The receptors belong to a superfamily of cytokine receptors with no intrinsic tyrosine kinase, which use the Jak-Stat cascade as a major signalling pathway. Hormones and receptors are thought to have arisen as a result of gene duplication and subsequent divergence early in vertebrate evolution. Mammalian growth hormone and prolactin show a slow basal evolutionary rate of change, but with episodes of accelerated evolution. These occurred for growth hormone during the evolution of the primates and artiodactyls and for prolactin in lineages leading to rodents, elephants, ruminants, and man. Placental lactogen has probably evolved independently on three occasions, from prolactin in rodents and ruminants and from growth hormone in man. Receptor sequences also show variable rates of evolution, corresponding partly, but not completely, with changes in the ligand. A principal biological role of growth hormone, the control of postnatal growth, has remained quite consistent throughout vertebrate evolution and is largely mediated by insulin-like growth factors. Prolactin has many and diverse roles. In relation to lactation, the relative roles of growth hormone and prolactin vary between species. Correlation between the molecular and functional evolution of these hormones is very incomplete, and it is likely that many important functional adaptations involved changes in regulatory elements, for example, altering tissue of origin or posttranscriptional processing, rather than change of the structures of the proteins themselves. The Babraham Institute, Babraham, Cambridge, United Kingdom. isabel.forsyth@bbsrc.ac.uk

Graham, L.H., Bolling, J., Miller, G., Pratt-Hawkes, N., Joseph, S., 2002. Enzyme-immunoassay for the measurement of luteinizing hormone in the serum of African elephants (Loxodonta africana). Zoo Biology 21, 403-408.
Abstract: Circulating patterns of progesterone and luteinizing hormone (LH) in the elephant have been well characterized, and routine monitoring of these hormones is now viewed as a valuable tool for making informed decisions about the reproductive management of elephants in captivity. Currently LH monitoring in elephants is done with radio-immunoassays (RIAs); unfortunately, the use of radioactive materials in RIAs limits their application to institutions with laboratory facilities equipped for the storage and disposal of radioactive waste. Enzyme-immunoassays (EIAs)offer an inexpensive and more zoo-friendly alternative to RIA. This work reports on an EIA capable of quantifying circulating LH in African elephants. The EIA employs a biotin label and microtiter plates coated with goat anti-mouse gamma globulin. LH surges in African elephants (n=3) increased fivefold over baseline concentrations (1.00±0.1 ng/ml vs. 0.2±0.1 ng/ml) and occurred 19.3±0.2 days apart. Ovulatory LH surges were associated with an increase in serum progestogens from 4.8±0.4 ng/ml to 11.7±0.4 ng/ml. The ability to quantify reproductive hormones in elephants via EIA is an important step in the process of making endocrine monitoring more accessible to zoos housing these species.

Grandy, J.W., Rutberg, A.T., 2002. An animal welfare view of wildlife contraception. Reprod Suppl 60, 1-7.
Abstract: Although there is some dissent, the animal protection community generally supports the concept of wildlife contraception. However, some contraceptive agents, delivery mechanisms and specific applications will be opposed by animal welfare advocates on environmental, humane or other ethical grounds, and some animal rights advocates may oppose wildlife contraception entirely. The Humane Society of the United States (HSUS) has supported and conducted wildlife contraception studies for more than 10 years. In general, we have invested in contraceptive agents (such as porcine zona pellucida) that we believe will prove environmentally, physiologically and behaviourally benign, and in delivery mechanisms that are narrowly targeted. As we consider contraception to be a major intervention into natural processes, we believe that wildlife contraception should be applied judiciously, locally and in a manner that is sensitive to the needs of animals, humans and ecosystem function.

Khawnual, P., Clarke, B., 2002. General care and reproductive management of pregnant and infant elephants at the Ayutthaya Elephant Camp. In: Baker, I., Kashio, M. (Eds.), Giants on Our Hands: Proceedings of the International Workshop on the Domesticated Asian Elephant, Bangkok, Thailand, 5-10 February 2001. FAO Regional Office for Asia and the Pacific (RAPA), Bangkok; Thailand, pp. 249-256.
Abstract: The management practices used at the Ayutthaya Elephant Camp (Thailand), where animals are kept for ecotourism purposes, are described. The aspects covered include feeding, provision of water, waste management and veterinary care (including health monitoring and disease control). Detailed descriptions are given of the procedures for management of pregnant females. The topics covered include mating, pregnancy diagnosis, parturition and postpartum management of the female and her offspring. These practices had resulted in 4 successful births in 2000.

Kumar, G.A., Ghosh, K.N.A., Sreekumaran, T., Chandrasekharan, K., 2002. Reproduction in elephants. Journal of Indian Veterinary Association Kerala 7, 38-40, 48-48.

Rasmussen, L.E., Riddle, H.S., Krishnamurthy, V., 2002. Mellifluous matures to malodorous in musth. Nature 415, 975-976.
Abstract: Male Asian elephants in musth--an annual period of heightened sexual activity and intensified aggression--broadcast odoriferous, behaviourally influential messages from secretions of the temporal gland. From our observations in the wild, together with instantaneous chemical sampling and captive-elephant playback experiments, we have discovered that young, socially immature males in musth signal their naivety by releasing honey-like odors to avoid conflict with adult males, whereas older musth males broadcast malodorous combinations to deter young males, facilitating the smooth functioning of male society. As elephant--human conflicts can upset this equilibrium, chemically modulating male behaviour may be one way to help the conservation of wild elephants.

Rasmussen, L.E., Wittemyer, G., 2002. Chemosignalling of musth by individual wild African elephants (Loxodonta africana): implications for conservation and management. Proc R Soc Lond B Biol Sci 269, 853-860.
Abstract: Elephants have extraordinary olfactory receptive equipment, yet this sensory system has been only minimally investigated in wild elephants. We present an in-depth study of urinary chemical signals emitted by individual, behaviourally characterized, wild male African elephants, investigating whether these compounds were the same, accentuated, or diminished in comparison with captive individuals. Remarkably, most emitted chemicals were similar in captive and wild elephants with an exception traced to drought-induced dietary cyanates among wild males. We observed developmental changes predominated by the transition from acids and esters emitted by young males to alcohols and ketones released by older males. We determined that the ketones (2-butanone, acetone and 2-pentanone, and 2-nonanone) were considerably elevated during early musth, musth and late musth, respectively, suggesting that males communicate their condition via these compounds. The similarity to compounds released during musth by Asian male elephants that evoke conspecific bioresponses suggests the existence of species-free 'musth' signals. Our innovative techniques, which allow the recognition of precise sexual and musth states of individual elephants, can be helpful to managers of both wild and captive elephants. Such sampling may allow the more accurate categorization of the social and reproductive status of individual male elephants.

Rasmussen, L.E.L., Riddle, H.S., Krishnamurthy, V., 2002. Mellifluous matures to malodorous in musth; Mood-altering secretions by excited male elephants smooth out social interactions. Nature 415, 975-976.

Sanchez, C.R., Murray, S.Z., Montali, R.J., Spelman, L.H. Medical Management of Acute Pylelonephritis in an Asian Elephant. Baer, C. K. American Association of Zoo Veterinarians Annual Conference.  162-164. 2002. 2002.
Ref Type: Conference Proceeding

Stokke, S., duToit, J.G., 2002. Sexual segregation in habitat use by elephants in Chobe National Park, Botswana. African Journal of Ecology 40, 360-371.
Abstract: We report on a study conducted on free-ranging African elephants in the woodlands of northern Botswana.We compared bull groups and family units with regard to (1) their patterns of habitat use and (2) their ranging distances from perennial water sources. During the dry season, adult males frequented more habitat types than family units, whereas family units used a wider diversity of habitats than bulls during the wet season. Bulls roamed widely (>10 km) from perennial drinking water in the dry season, when family units congregated within 3.5 km of the rivers. During the wet season,when ephemeral pans were abundant, all elephant groups were found at intermediate distances (5 km) from the rivers.The spacing of elephants in the dry season is consistent with sexual segregation but we reject the hypothesis that this is an outcome of indirect competition for food, because our concurrent studies on elephant feeding ecology found no evidence for intraspecific competition. Instead, we propose that most adult male elephants space themselves to avoid conflict with musth bulls and roam widely in the dry season between discretely distributed feeding 'hotspots'.The small proportion of males that are in musth remain close to family units to maximize mating opportunities, and family units are unable to range far from water in the dry season. This is due to (1) comparatively high rates of water turn-over among juveniles and lactating cows and (2) the reduced mobility of neonates.

Suedmeyer, W.K. Transabdominal ultrasonic Gestational monitoring in an African elephant (Loxodonta africana). Baer, C. K. American Association of Zoo Veterinarians Annual Conference.  219-220. 2002. 2002.
Ref Type: Conference Proceeding

Welsch, U., Unterberger, P., Hofter, E., Cuttitta, F., Martinez, A., 2002. Adrenomedullin in mammalian and human skin glands including the mammary gland. Acta Histochem 104, 65-72.
Abstract: Adrenomedullin is a peptide that has been ascribed numerous functions. In the present paper, adrenomedullin has been localized immunhistochemically in a variety of skin glands of humans, elephants and impalas: apocrine scent glands, eccrine sweat glands, holocrine glands and mammary glands. In the apocrine glands expression of adrenomedullin varied with respect to staining intensity and intracellular localization. In general, glands which appeared to be actively secreting were more strongly stained than quiescent glands. However, within a single glandular tubule, individual cells differed considerably in the staining intensity of adrenomedullin. Adrenomedullin was present in both non-lactating and lactating mammary secretory epithelia, both ducts and alveoli reacted positively. In human mammary glands displaying apocrine metaplasia, the apical protrusions were strongly positive. Furthermore, positive immunostaining was found in endothelium and often in smooth muscle cells of small arteries and veins and in mast cells as well. Many of the adrenomedullin-positive epithelial cells were most strongly stained in the area of the Golgi apparatus, the cellular apex and particularly close to the basal side of the cell membrane.This pattern suggests packaging of adrenomedullin into secretory granules and secretion both at the apex of cells and at their basis. The first form of secretion suggests exocrine secretion, the latter form endocrine secretion of adrenomedullin. A possible hormonal function is in line with basally located electron dense small secretory granules, which have been found by electron microscopy in the glandular epithelia studied.

Barber, M.R., Lee, S.M., Steffens, W.L., Ard, M., Fayrer-Hosken, R.A., 2001. Immunolocalization of zona pellucida antigens in the ovarian follicle of dogs, cats, horses and elephants. Theriogenology 55, 1705-1717.
Abstract: A comparative evaluation of the location of immunoreactive porcine zona pellucida (pZP) glycoproteins was performed with polyclonal rabbit anti-pZP antibodies on ovarian sections of the dog, cat, horse and elephant. For this, formalin (light microscopy) and glutaraldehyde (transmission electron microscopy [TEM]) fixed ovarian sections were incubated with antibodies raised against highly purified pZP. Staining patterns were determined with diaminobenzidine (DAB) at the light level. The dog ZP had a distinct staining distribution that is characterized by intense staining around the periphery of the ZP and the oolemma and less dense staining throughout the width of the ZP. In dog follicles that contained multiple oocytes, there were oocytes of identical and dissimilar stages. Cat ovarian sections showed uniform staining of the ZP. Horse results showed uniform staining of ZP and ooplasm, and granulosa cells (GC). Elephant sections showed staining of the ZP with dense staining at the oolemma, as well as staining of the ooplasm. In all species the staining of the ZP was not evident until GC differentiation. In all cases there was no staining of ovarian tissue with control normal rabbit serum. Specific staining patterns of ZP were evaluated by TEM and immunogold staining. The immunogold-linked anti-pZP antibodies stained the ZP matrix in all species. There was staining of ooplasm organelles suggesting that ZP secretion originates from the oocyte of the dog and cat. In addition, follicular and ZP measurements were taken that allowed accurate characterization of follicle stage. These findings suggest that in all 4 species the ZP is recognized by anti-pZP antibodies and there is also evidence to suggest the possible origins of ZP glycoproteins.

Campbell, M.M., 2001. Pachyderms, primates, plants and population. Reprod Fertil Dev 13, 697-703.
Abstract: In the past, growth in human population has often been associated with species loss. Current rates of population growth, both globally (1 million more births than deaths every 103 hours) and regionally, pose a threat of additional ecological damage. There is a well-documented unmet demand for family planning in nearly all high-fertility countries. Improved family planning and safe abortion services will improve the health of women and their families, accelerate fertility decline, and help preserve the environment. Many ecologically vulnerable areas are especially poorly served by family planning services. Examples are given here of improving family planning services through private health providers near the Kakamega Forest in western Kenya, and of adding family planning choices to a reforestation project run by the Jane Goodall Institute near the Gombe National Park, Tanzania. Wildlife biologists can play a critical role in identifying local professionals and institutions with the potential to improve family planning.

Dastig, B. Birth and Reproduction Rate in a Herd of Captive Asian Elephants at the Pinnawela Elephant Orphanage. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  19-23. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding
Abstract: The majority of Asian elephants currently or previously held in zoos and circuses in Europe and North America came into captivity as young animals between two and five years of age. This was possible due to the low transportation costs. These young animals, either captured or born to working elephants, were merely a by-product and thus sold at a low price. Today, Asian elephants destined for zoos come from working elephants or from jungle camps. These animals are all orphans and have never lived in a natural social structure. Circuses and zoos usually keep only female elephants, which are then deprived of a social structure including an alpha cow, aunts, sisters and their young. This results in the animals' rarely giving birth in captivity. This is why gathering data and conducting observation in this field is particularly difficult. It is for this reason that the Pinnawela Elephant Orphanage offers an ideal location for learning more about reproduction and social structure in Asian elephants. The orphanage is located in Sri Lanka halfway between the capitol, Colombo, and Kandy, the ancient royal city. The orphanage represents the largest herd of Asian elephants in human care. In 1997 the herd consisted of 56 elephants; in 1998 the population had grown to include 63 elephants.

Dehnhard, M., Hildebrand, T., Rohleder, M., Strauss, G., Meyer, H.H.D., Goritz, F., 2001. Application of an enzyme-immunoassay (EIA) for rapid screening of 5alpha-pregnane-3,20-dione (DHP) in blood plasma of the Asian elephant, Elephas maximus. Berliner und Munchener Tierarztliche Wochenschrift. 114, 161-165.
Abstract: Populations of African (Loxodonta africana) and Asian elephants (Elephas maximus) in zoos and safari parks are at risk due to their low reproductive success. To extend the limited knowledge of their reproductive physiology, easy and practical methods for the analysis of relevant reproductive hormones must be developed to support assisted reproduction, for instance. Blood samples from 2 nonpregnant and 2 pregnant Asian elephants were used in the following study. For the measurement of 5alpha-pregnane-3,20-dione (DHP), the predominant ovarian gestagen in both species, an enzyme-immunoassay (EIA) based on commercial reagents was applied. Advantages of this EIA are the small volume of plasma needed for evaluation (5 µl) and the possibility of direct processing without an extraction stage. The lower limit of detection was 0.16 ng/ml, mean recovery was 101% and the mean coefficients of variation were 7.3 (intra-assay) and 9.9% (inter-assay). In the Asian elephants, DHP levels reached 15 ng/ml during the luteal phase and 21 ng/ml during pregnancy. Oestrous cycle lengths based on the lowest DHP concentrations varied from 12 to 20 weeks (mean of 15.4±2.3). In two Asian elephant cows, a calf was stillborn. Thereafter, ovarian activity in the animals resumed after approximately 8 and 13 weeks, respectively. In one animal, estradiol implants for hormonal contraception caused a down regulation of ovarian function as demonstrated by an irregular pattern of DHP secretion over a period of 48 weeks. It is proposed that the direct DHP-EIA is a suitable method for reproductive monitoring in elephants, as it can be easily established in laboratories.

Dehnhard, M., Hesitermann, M., Goritz, F., Hermes, R., Hildebrand, T., Haber, H., 2001. Demonstration of 2-unsaturated C19-steroids in the urine of female Asian elephants, Elephas maximus, and their dependence on ovarian activity. Reproduction-Cambridge 121, 475-484.
Abstract: An oestrous-related pheromone of the female Asian elephant (Elephas maximus) is known to induce behavioural responses in elephant bulls. Additional data revealed that timing of oestrus in females with close social relationships tends to be synchronized. Therefore, urine from female Asian elephants might be expected to contain luteal phase-dependent volatile substances, which may function as additional chemical signals in this species. The aim of the present study was to identify such compounds and to investigate their pattern of excretion throughout the ovarian cycle. Urine samples were collected 3 times a week during the follicular phase and 1 to 3 times a week during the luteal phase from 5 adult female Asian elephants from a total of 13 non-conception cycles and one conception cycle, including the first 72 weeks of pregnancy. A simple headspace solid-phase microextraction method has been developed for quantification of urinary volatile substances and analysis was performed by gas chromatography. The comparison of urine collected during the follicular and the luteal phase indicated the presence of two luteal phase-dependent substances. Mass spectrometry was used to identify one substance as 5alpha-androst-2-en-17-one and a second substance as the corresponding alcoholic compound 5alpha-androst-2-en-17beta-ol. The 5alpha-androst-2-en-17beta-ol and -17-one profiles reflected cyclic ovarian activity with clear (10-20-fold) luteal phase increases. Furthermore, measurements of both compounds were correlated positively with the concentration of urinary pregnanetriol and indicated cycle duration (15.1±1.2 weeks) similar to that obtained from pregnanetriol measurements (15.2±1.6 weeks). The results demonstrated the presence of 2 luteal phase-specific steroidal volatile compounds in elephant urine. One of the substances, 5alpha-androst-2-en-17-one, has been demonstrated in human axillary bacterial isolates. The measurement of both volatile substances in elephant urine can be used for rapid detection of the stage of the ovarian cycle, as the analysis can be completed within 2 h.

Delsink, A.K., van Altena, J.J., Kirkpatrick, J., Grobler, D., Fayrer-Hosken, R. Field applications of immunocontraception in African elephants (Loxodonta africana). Proceedings of the Fifth International Symposium on Fertility Control in Wildlife.  2001.  Society for Reproduction and Fertility; Cambridge; UK. 2001.
Ref Type: Conference Proceeding
Abstract: The primary aim of the Makalali elephant immunocontraception programme is to test the efficacy of porcine zona pellucida (PZP) vaccine for practical population control of elephants in small, enclosed reserves, with the goal of stabilizing the current growth rate and reducing it to the 5-10% per annum displayed currently in the Kruger National Park. A secondary aim is to test the hypothesis that PZP treatment does not affect patterns of elephant social behaviour. Eighteen sexually mature cows (age > 12 years) were vaccinated in May 2000 using remote darts. Behavioural observations before, during and after vaccination included noting the activity of individual animals every minute for 15 min. No changes in general behaviour patterns have been noted to date although the animals' spatial use of the reserve was erratic during the period of vaccination, indicating irregular or disturbed patterns associated with vaccination. Normality was resumed on completion of the vaccinations. No aggressive or indifferent behaviour related to nursing, calf proximity or bull-cow interactions have been noted. Ten of the females were in various stages of pregnancy when treated. Subsequently, seven of them gave birth to healthy calves and the other three females are expected to calve shortly. It is too early in the study to draw conclusions about stabilization of growth rates.

Doi, O., Yamada, T., Terazono, M., Wada, S., 2001. Macroscopic changes in the mucous membrane of vaginal vestibule during the estrous cycle of female Asian elephant. Japanese Journal of Zoo and Wildlife Medicine 6, 55-59.

Flugger, M., Goritz, F., Hermes, E., Isenbugel, A., Klarenbeek, W., Schaftenaar, W., Schaller, K., Strauss, G. Evaluation of physiological data and veterinary medical experiences in 31 Asian elephant (Elephas maximus) births in six European zoos. Verh ber Erkrg Zootiere (Proc. 40th Intl Symp  Zoo and Wild Anim  Med).  123-133. 2001. Rotterdam, Netherlands. 2001.
Ref Type: Conference Proceeding

Foley, C.A.H., Papageorge, S., Wasser, S.K., 2001. Noninvasive stress and reproductive measures of social and ecological pressures in free-ranging African elephants. Conserv Biol 15, 1134-1142.

Fritsch, G., Hermes, R., Maltzan, J. New Aspects of Sexual Maturation in Male Elephants. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  25. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Ganslober, U. Behavioural Ecology, Social Relationships, Life History and Evolutionary Constraints in Megaherbivores. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  26-31. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Ganswindt, A., Heistermann, M., Borragan, S., Hodges, J.K., 2001. Assessment of testicular endocrine function in captive African elephants by measurement of urinary and fecal androgens. Zoo Biology 21,  27-36.

Ganswindt, A., Heistermann, M., Hodges, J.K. Faecal Glucocorticoid and Androgen Metabolite Excretion in Male African Elephants (Loxodonta africana). A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  258. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Goeritz, F., Hildebrandt, T.B., Hermes, R., Quandt, S., Jewgenow, K., Hofmann, R.R., Hofer, H., Meyer, H.H.D. Results of Hormonal Contraception in Free-Ranging African Elephants. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  262. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Graham, L., Schwarzenberger, F., Möstl, E., Galama, W., Savage, A., 2001. A versatile ezyme immunoassay for the determination of progestogens in feces and serum. Zoo Biology 20, 227-236.
Abstract: The ability of zoos to monitor the reproductive status of their animals can vastly improve the effectiveness of husbandry/management practices, and noninvasive methods such as fecal steroid analysis are the easiest to apply in a zoo setting. Furthermore, enzyme immunoassay (EIA) is preferred to radioimmunoassay (RIA) as the method of quantifying hormones because EIAs do not involve the use, storage, and disposal of radioactive materials. However, progesterone is excreted in the feces as predominantly unconjugated metabolites (progestogens) and, until recently, antibodies able to cross-react with a variety of progestogens were used primarily in RIAs. An EIA using a broad-spectrum progestogen antibody is described and applied to serum and/or fecal samples from female African elephants, black rhinoceros, white rhinoceros, okapi, and hippopotami. The clear progestogen profiles generated in these species suggest that the described EIA would be as versatile as the RIA using the same antibody and could be a practical and economical alternative to RIAs for monitoring gonadal function via progestogen analysis in zoo species.

Hanks, J., 2001. Conservation strategies for Africa's large mammals. Reprod Fertil Dev 13, 459-468.
Abstract: Africa's large mammals are conserved for their aesthetic, scientific and economic values. Many of these species face a gloomy future precipitated by a combination of factors directly and indirectly influenced by the activities of man, including habitat loss, overexploitation, poor management of designated protected areas, and the vulnerability of small isolated populations. Africa's designated protected areas and biodiversity hotspots are also under threat, highlighting the importance of embracing community participation to address accelerating poverty and malnutrition. Innovative strategies are required for the conservation of Africa's mammals, such as the integration of a wide range of species in the production landscape, including the farming community. Transfrontier conservation areas (TFCAs) have been established with the combined objectives of conserving biodiversity, creating new jobs in the tourism and wildlife industry, and promoting a culture of peace. These areas extend far beyond traditional national parks, providing opportunities for integrating large mammals into sustainable land-use practices, at the same time as addressing some of the continent's more pressing socioeconomic needs. Research on African mammals will inevitably have to change direction to accommodate the growing threats and changed circumstances. Priorities will include the identification of corridors associated with TFCA establishment, the determination of the economic value of certain species in consumptive use programmes, research on contraception as a management option in restricted areas, and further work on the indirect use value of species. There will also be worthwhile opportunities to be pursued with ex situ conservation programmes, but these need to be focussed more efficiently.

Hildebrandt, T.B., Hermes, R., Pratt, N.C., Brown, J.L., Schwammer, H., Schmitt, D., Jewgenow, K., Olson, D., Lehnhardt, J.L., Goritz, F. Results of Artificial Insemination Programmes in Asian and African Elephants Kept Under Different Management Systems. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  52. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Kirkman, S., ., Wallace, E.D., van Aarde, R.J., Potgieter, H.C., 2001. Steroidogenic correlates of pregnancy in the rock hyrax (Procavia capensis). Life Sciences 68, 2061-2072.
Abstract: In pregnant rock hyraxes (P. capensis) isolated leucocytes metabolize both [3H]pregnenolone and [3H]progesterone while whole blood, erythrocytes and an erythrocyte/leucocyte mixture only metabolized [3H]progesterone. Plasma displayed no tendency to metabolically convert any one of these two steroids. In whole blood, [3H]progesterone appears to be converted to 5alpha-pregnane-3,20-dione and a compound with chromatographic properties similar to that of 5alpha-pregnan-3alpha-ol-20-one. 5alpha-Pregnane-3,20-dione exhibited a high relative binding affinity for the uterine progesterone receptor (94%), but 5alpha-pregnan-3alpha-ol-20-one displayed very little affinity for the same receptor (0.4%). 5alpha-Pregnane-3,20-dione may therefore aid in the maintenance of pregnancy. Corpora lutea metabolized progesterone to 17alpha-hydroxyprogesterone, a compound exhibiting no progestational function because of its low relative binding affinity for the uterine progesterone receptor (2%). Progesterone appears to be the main product of the corpus luteum. However, 5alpha-pregnane-3,20-dione circulated at concentrations approximately 8.5 times higher than progesterone, probably due to the metabolic conversion of progesterone to 5alpha-pregnane-3,20-dione by the blood. We conclude that in the hyrax, progesterone, produced by the corpora lutea, enters the circulation, where it is reduced to 5alpha-pregnanes. 5alpha-Pregane-3,20-dione may then be transported to the uterus where it binds to the progesterone receptor to assist in the maintenance of pregnancy. This mechanism appears to be analogous to that of the African elephant (Loxodonta africana) which is phylogenetically related to the hyrax, except that in the elephant the 5alpha-reduced metabolites are produced by luteal tissue and not the blood.

Kurt, F., Touma, C. Musth in Wild - Living and Captive Asian Elephants in Sri Lanka. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  64-69. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Lehnhardt, J., Bolling, J., Pratt, N., Joseph, S., Miller, G., Graham, L., Miller, M., Neiffer, D., Hildebrandt, T., Goeritz, F. Elephant Artificial Insemination (AI) in Protected Contact. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  70. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Miller, D.L., Dougherty, M.M., Decker, S.J., Bossart, G.D., 2001. Ultrastructure of the spermatozoa from a Florida manatee (Trichechus manatus latirostris). Anat Histol Embryol 30, 253-256.
Abstract: Semen was opportunistically collected from a free-ranging, 10-year-old, 275 cm (total length) Florida manatee (Trichechus manatus latirostris) during rehabilitation treatments. Ultrastructure of the spermatozoa was examined by scanning and transmission electron microscopy and differed slightly from that described for other mammals. Comparisons to the manatee's closest phylogenetic relatives, the elephant and hyrax, were made. The manatee spermatozoa had a similar acrosome but a distinct annulus and lacked the dense bodies observed in the neck of the elephant spermatozoa. Additionally, manatee spermatozoa lacked the lateral vacuoles observed in the nuclear chromatin from of the hyrax spermatozoa. These data add to our understanding of manatees and allow for comparative studies with other species that may be useful in phylogenetic and reproductive studies.

Moss, C.J., 2001. The demography of an African elephant (Loxodonta africana) population in Amboseli, Kenya. J. Zool. , Lond. 255, 145-156.
Abstract: This paper presents basic demographic parameters of African elephants (Loxodonta africana) living in and around Amboseli National Park, Kenya. The study was conducted from 1972 to the present and results are based on the histories of 1778 individually known elephants. From 1972 to 1978, the Amboseli elephant population declined and then increased steadily from 1979 to the present. Births occurred throughout the year but over 80% occurred between November and May. Birth rate varied from year to year with a pattern of peaks and troughs at 4- to 5-year intervals. The birth sex ratio did not differ significantly from 1:1. Mean age at first birth was 14.1 years, determined from a sample of 546 known-age females. Mean birth interval (n = 732) was 4.5 years for 255 females. Fecundity and calf survival varied by age of the females. Mortality fluctuated from year to year. Sex-specific mortality rates were consistently higher for males than females at all ages.

Oerke, A., Heistermann, M., Hodges, K. Reproductive Characteristics of the European Elephant Population: Long-Term Cycle and Pregnancy Data Based on Non-Invasive Methodology. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  103. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Pimm, S.L., van Aarde, R., 2001. African elephants and contraception. Nature London 411, 766.

Rasmussen, L.E.L., Riddle, H.S. Musth in Teenage Male Asian Elephants (Elephas maximus): The What & the Why of their Chemical Signals. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  110. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Rasmussen, L.E.L., 2001. Source and cyclic release pattern of (Z)-7-dodecenyl acetate, the pre-ovulatory pheromone of the female Asian elephant. Chemical Senses 26, 611-623.
Abstract: Female Asian elephants (Elephas maximus) release a pre-ovulatory urinary pheromone, (Z)-7-dodecenyl acetate (Z7-12:Ac), to signal males of their readiness to mate. Z7-12:Ac is quantitatively elevated during the follicular stage of oestrus, reaching maximum concentrations just prior to ovulation, as demonstrated by two complementary headspace techniques: (1) evacuated canister capture followed by cryogenic trapping and (2) solid phase microextraction (SPME) used prior to gas chromatography/mass spectrometry (GC/MS). These patterns were coincident with observed male behaviours and were consistent with biochemical and binding properties of the active ligand, including optimal binding pH. To release maximum amounts of Z7-12:Ac for quantitation, serum and urine samples from three mature female Asian elephants in their luteal and follicular stages of several oestrous cycles were subjected to heat and pH changes and were then treated with protease prior to SPME-GC/MS analyses. When the post-luteal serum progesterone concentrations declined to baseline levels, Z7-12:Ac became detectable in the female urine. Throughout the follicular stage, pheromone concentrations increased linearly with no apparent relationship to the two serum luteinizing hormone peaks. Pre-ovulatory urine also contained related compounds, including (Z)-7-12-dodecenol. The relative amount of this alcohol increased relative to acetate during long-term storage, with a proportional reduction in bioactivity. Z7-12:Ac was not detected in mucus samples from the urogenital tract. A potential precursor of Z7-12:Ac was identified in liver homogenates from female elephants in the follicular stage.  Erratum in: Chem Senses 2001 Sep;26(7):935

Sarma, K.K., 2001. Musth in Asian Elephant. Central Zoo Authority, New Delhi, India.

Schaftenaar, W., Hildebrandt, T.B., Flugger, M., Goritz, F., Schmitt, D., West, G. Guidelines for veterinary assistance during the reproduction process in female elephants. Proceedings American Association of Zoo Veterinarians, American Association of Wildlife Veterinarians, Association of Reptilian and Amphibian Veterinarians, and the National Association of Zoo and Wildlife Veterinarians Joint Conference.  348-355. 2001. USA.
Ref Type: Conference Proceeding
Abstract: In February 2000, a group of European zoo veterinarians met at Tierpark Hagenbeck, Hamburg to evaluate a questionnaire about 31 parturitions in Asian elephants.  The results were presented at the 40th International Symposium on Diseases of Zoo and Wild Animals.  The results were combined with the experiences of some North-American zoo veterinarians, which resulted in the protocol presented in this paper.  The protocol may serve as a guideline for institutions that wish to breed elephants. The proper application of the recommendations given in these guidelines should increase the reproductive success in elephants.  It is the moral obligation of everyone who is responsible for the  management and breeding of elephants to consider utilizing the guidelines as they may apply to their situation and to collect data that may help increase our knowledge. The breeding process in elephants requires monitoring of several parameters in both males and females. The most crucial parameters are the determination of the estrous cycle through progesterone and, perhaps, LH assay, evaluation of the genital tract in both sexes, determination of the number of fetuses and finally, parturition.  The first part of the paper will mention briefly the tools that can be used in female elephants to achieve these goals.  The second part describes a protocol for veterinary intervention in elephant parturition.

Schmitt, D. Riddles's Elephant amd Wildlife Sanctuary Elephant Birth Protocol.  2001.
Ref Type: Internet Communication

Schmitt, D., Krywko, R., Reichard, T.A., Shellabarger, W., Bailey, K., Short, J. Surgical approach to artificial insemination in elephants. Proceedings American Association of Zoo Veterinarians, American Association of Wildlife Veterinarians, Association of Reptilian and Amphibian Veterinarians, and the National Association of Zoo and Wildlife Veterinarians Joint Conference.  338. 2001. USA. 1.
Ref Type: Conference Proceeding
Abstract: Artificial insemination is a recent development for assisted reproduction in elephants.  Non-surgical insemination requires a cooperative elephant,   well-habituated to the various procedures.  In addition, a well-trained and equipped insemination team is needed to successfully complete the procedure.  A surgical approach for artificial insemination in elephants reduces both the technology needed for success and the level of cooperation needed from the elephant to be inseminated.  The first successful, surgical, artificial insemination was accomplished by making a 3-cm incision into the urogenital canal just below the anus.  The vestibulotomy incision was guided by the placement of 8-cm PVC tube with a 3-cm opening at the upper end, up through the vulva to a level just below the anus.  The opening in the PVC tube was used as a guide by palpating the opening through the skin.  The incision was made following injection of a local anesthetic above the proposed incision site.  The PVC guide prevents incision into the opposite wall of the urogenital tract.  After the incision is complete a sterile disposable vaginal speculum is introduced into the urogenital canal.  The intact hymen or cervix can be visualized directly with a flashlight or, for documentation of the procedure, a short endoscope can be utilized.  Placement of semen into the vagina can be accomplished with little difficulty using sterile disposable horse insemination pipettes.  Multiple inseminations are possible through the incision for the 2-3 days of estrus.  Following the last insemination, a local anesthetic is administered and the edges of the incision are freshened and four to six simple interrupted sutures are placed to close the incision.  Healing of the incision requires 4 to 6 weeks with good aftercare.

Schmitt, D.L., Krywko, R., Reichard, T.A., Shellabarger, W. Surgical approach to artificial insemination in elephants. Kirk Baer, C. and Wilmette, M. W. Proceedings American Association of Zoo Veterinarians, American Association of Wildlife Veterinarians, Association of Reptilian and Amphibian Veterinarians and the National Association of Zoo and Wildlife Veterinarians Joint Conference  2001.  338. 2001.  American Association of Zoo Veterinarians. 9-18-2001.
Ref Type: Conference Proceeding

Schmitt, D.L., Krywko, R.L., Reichhardt, T., Shellabarger, R.W., Bailey, K.M., Short, J.N. Surgical approach to artificial insemination in elephants. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  129-131. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding
Abstract: Artificial insemination in elephants is a recent development for assisted reproduction in elephants. Non-surgical insemination requires both a cooperative and well-habituated elephant to the various procedures. In addition a well-trained and equipped insemination team is needed to successfully complete the procedure. A surgical approach for artificial insemination reduces the technology needed for success and the level of cooperation needed from the elephant to be inseminated. The first successful surgical artificial insemination was accomplished by making a 3cm incision into the urogenital canal just below the anus. The vestibulotomy incision was guided by placement of an 8 cm diameter PCV tube, with a 3cm opening near the upper end, through the vulva up to the level just below the anus. The opening in the PCV tube was used as a guide by palpating above the proposed incision site. This guide prevents incision into the opposite wall of the urogenital tract. After the incision is complete a sterile disposable vaginal speculum is introduced into the urogenital canal. The intact hymen or cervix can be visualized directly with a flashlight or, for documentation of the procedure, a short endoscope can be utilized. Placement of semen into the vagina or hymen can be accomplished with little difficulty through the incision for the two to three days of estrus. Following the last insemination, a local anesthetic is administered and the edges of the incision are freshened and four to six simple interrupted sutures are placed to close the incision. Healing of the incision requires four to six weeks with good aftercare.

Schulte, B.A. Examining Ideas on the Evolution of Musth. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  287. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Schulte, B.A., Slade, B.E., Rasmussen, L.E.L. The Trunk and Tail of Elephant Communication: Studies on Captive Asian Elephants. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  286. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Schwammer, H. From Artificial Insemination to Birth. A Case Study on African Elephants (Loxodonta africana). A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  289-292. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Suedmeyer, W.K. Serum hydrocortisone levels in a manually restrained African elephant (Loxodonta africana)  pre- and post- semen collection. Kirk Baer, C. and Wilmette, M. W. Proceedings American Association of Zoo Veterinarians, American Association of Wildlife Veterinarians, Association of Reptilian and Amphibian Veterinarians and the National Association of Zoo and Wildlife Veterinarians Joint Conference.  388-389. 2001.   American Association of Zoo Veterinarians. 9-18-2001.
Ref Type: Conference Proceeding

Wingate, L., Lasley, B. Is Musth a Reproductive Event: An Examination of Arguments For and Against this View. A Research Update on Elephants and Rhinos; Proceedings of the International Elephant and Rhino Research Symposium, Vienna, June 7-11, 2001.  150-156. 2001. Vienna, Austria, Schuling Verlag. 2001.
Ref Type: Conference Proceeding

Wisser, J., Pilaski, J., Strauss, G., Meyer, H., Burck, G., Truyen, U., Rudolph, M., Frolich, K., 2001. Cowpox virus infection causing stillbirth in an Asian elephant (Elephas maximus). Veterinary Record 149, 244-246.

Wyatt, J. Elephant breeding soundness examination forms and database. Kirk Baer, C. and Wilmette, M. W. Proceedings American Association of Zoo Veterinarians, American Association of Wildlife Veterinarians, Association of Reptilian and Amphibian Veterinarians, and the National Association of Zoo and Wildlife Veterinarians Joint Conference 2001.  396-400. 2001. USA, AAZV. 1.
Ref Type: Conference Proceeding
Abstract: Routine use of standardized soundness examination forms for male and female elephants provides data for an inter-relational database.  Through queries we may answer population-based reproductive questions essential for promoting self-sustaining populations. This poster presentation demonstrates breeding soundness examination forms and accompanying database used in a pilot project to evaluate 25 elephants.

Agnew, D.W., Munson, L., Gage, L.J., Fowler, M.E., Ramsay, E. Cystic Endometrial Hyperplasia in Nulliparous Asian Elephants. 2000 Proceedings AAZV and IAAAM Joint Conference.  442. 2000. 2000.
Ref Type: Conference Proceeding
Abstract: Maintaining self-sustaining populations of elephants through captive breeding is a new goal of the Elephant Species Survival Plan. Most elephants available for breeding in U.S. zoos are nulliparous and aged, and their fertility is unknown. Endometrial hyperplasia has been noted in aged elephants, and this condition may affect their fertility. The purpose of this study was to better characterize the gross and histopathologic features of these lesions and assess the demographic distribution. Clinical histories, necropsy reports, and endometrial samples from Asian elephants (Elephas maximus) at least 20 yr of age that died from 1985 to 1999 were reviewed. Gross pathologic findings in all cases were similar and consisted of a multifocal to diffuse distribution of 1-2 cm diameter cysts in the endometrium. Pedunculated edematous endometrial polyps up to 10 cm in length also were present, projecting from the endometrial surface or free within the lumen. Some polyps were necrotic. Histologically, the endometrium was characterized by varying sized cystic endometrial glands lined by cuboidal to tall columnar epithelium. Other glands were present in small clusters and lined by hyperplastic endometrium. The endometrial polyps consisted of a core of edematous stroma containing clusters of cystic glands. Tissues expelled from the urogenital tract of another aged, nulliparous cow were also reviewed. These fragments consisted of necrotic tissue with ghost-like remnants of glands similar to endometrial glands. These fragments may represent expelled pedunculated endometrial polyps, which had become necrotic and sloughed. These results indicate that aged nulliparous Asian elephants commonly develop cystic endometrial hyperplasia and that the pedunculated polyps may represent a more advanced form of this disease. Sloughing of these pedunculated polyps may be noted clinically and may offer information about the condition of a cow's endometrium. The effect of endometrial hyperplasia on fertility in elephants is unknown, but in other species large numbers of cysts can interfere with implantation. The prevalence of these lesions in aged elephants suggests that younger animals would be better candidates for breeding and that efforts should be made to clinically evaluate potential breeding cows for endometrial health.

Ananth, D., 2000. Musth in elephants. Zoos' Print Journal 15, 259-262.

Barber, M.R., Fayrer-Hosken, R.A., 2000. Possible mechanisms of mammalian immunocontraception. J Reprod Immunol 46, 103-124.
Abstract: Ecological and conservation programs in ecosystems around the world have experienced varied success in population management. One of the greatest problems is that human expansion has led to the shrinking of wildlife habitat and, as a result, the overpopulation of many different species has occurred. The pressures exerted by the increased number of animals has caused environmental damage. The humane and practical control of these populations has solicited the scientific community to arrive at a safe, effective, and cost-efficient means of population control. Immunocontraception using zona pellucida antigens, specifically porcine zona pellucida (pZP), has become one of the most promising population control tools in the world today, with notable successes in horses and elephants. A conundrum has risen where pZP, a single vaccine, successfully induces an immunocontraceptive effect in multiple species of mammals. This review describes the most current data pertaining to the mammalian zona pellucida and immunocontraception, and from these studies, we suggest several potential mechanisms of immunocontraception.

Brown, J.L. Zoo Biology. Special Issue on elephant biology 19[5], 1-184. 2000.
Ref Type: Journal (Full)
Abstract: This issue focuses on elephant biology and includes the following topics: ultrasonography of the urogenital tract in elephants Loxodonta africana and Elephas maximus (an important tool for assessing female and male reproductive function); reproductive endocrine monitoring of elephants (an essential tool for assisting captive management); ultrasonography of the oestrous cycle in female African elephants; review of a newly recognized disease of elephants caused by endotheliotropic herpesviruses; tuberculosis in elephants in North America; how chemical signals integrate Asian elephant society; elephant communication; social structure and helping behaviour in captive elephants; a postcryogenic comparison of membrane fatty acids of elephant spermatozoa; and first disclosure and preliminary investigation of a liquid released from the ears of African elephants.

Brown, J.L., 2000. Reproductive endocrine monitoring of elephants: an essential tool for assisting captive management. Zoo Biology 19, 347-367.
Abstract: Considerable information now is available about the basic reproductive biology of elephants, especially females. However, as important as this knowledge is, it no longer is enough to simply compile it into a database. The potential exists for using endocrine monitoring techniques to solve real problems. This review summarizes our current knowledge of elephant endocrinology and offers suggestions on how to use the technology to maximize reproductive potential. The oestrous cycle can be monitored through the analysis of serum progestogens, primarily 5alpha-reduced compounds, and consists of an 8- to 12-week luteal phase and a 4- to 6-week inter-luteal period. Proof of ovarian cyclicity currently is mandatory before Species Survival Plan breeding recommendations are approved. However, because many adult females are not cycling normally, the reproductive monitoring of all cows throughout their life span is now encouraged. Complete endocrine evaluations in conjunction with ultrasound examinations and behavioral assessments are needed to identify causes of reproductive failure and develop mitigating treatments. Progestogen analyses also are effective for monitoring pregnancy, but only if longitudinal samples are collected. Alternatively, pregnancy can be diagnosed in occasional samples using serum prolactin or possibly relaxin measurements after 20 weeks of gestation. Parturition can be predicted on the basis of the rapid decrease in progestogens that occurs about 2-5 days before birth. An updated model of ovarian dynamics during the oestrous cycle suggests that two waves of follicular development occur 3 weeks apart during the non-luteal phase, possibly under the control of follicle-stimulating hormone. Each follicular wave culminates in a luteinizing hormone (LH) surge, with the second surge inducing ovulation and corpus luteum formation. The functional significance of the first, anovulatory LH surge is under investigation, but from a practical perspective it can be used to schedule breeding (by artificial insemination or natural mating) to coincide with the ovulatory LH surge. Less is known about the reproductive biology of bulls, aside from the fact that musth is associated with dramatic changes in androgen secretion. Studies are needed to determine whether poor libido and inadequate semen quality observed in some mature elephants are due to testicular steroidogenic dysfunction. When blood samples cannot be collected for routine hormone analysis, gonadal activity can be monitored non-invasively through the measurement of excreted steroid metabolites (males: androgens; females: estrogens, progestogens) in urine and faeces. Lastly, suggestions for future research priorities are provided.

Doi, O., Komatsumoto, M., Terazono, M., Wada, S., 2000. Exfoliative cytology in vaginal vestibule of female Asian elephants: relation to circulating progesterone concentrations. Zoological Science 17, 1303-1309.
Abstract: The oestrous cycle of the Asian elephant (Elephas maximus) was monitored by analysis of exfoliative cytology in the vaginal vestibule and serum progesterone concentrations. Appearance frequency of each 5 exfoliative cells; parabasal, intermediate, superficial anuclear and nuclear cells and leukocytes; on the smear collected from two elephants was calculated, and serum progesterone concentrations were measured by radioimmunoassay. Serum progesterone concentrations changed regularly with the cycle between 14 and 17 weeks. Using spectrum analysis (Yule-Walker method) to appearance frequency of exfoliative cells, it was found that the time when a superficial cell markedly appeared in vaginal vestibule corresponded to the time when serum progesterone concentration was almost negligible. It is suggested that the time when numbers of two kinds of superficial (anuclear and nuclear) cells and parabasal and intermediate cells increase to the smear of the elephant, means the period from pro-oestrus to oestrus and from metoestrus to dioestrus, respectively.

Emanuelson, K.A., Kinzley, C.E. Salmonellosis and subsequent abortion in two African elephants. Proc. AAZV and IAAAM Joint Conf.  269-274. 2000.
Ref Type: Conference Proceeding

Fayrer-Hosken, R.A., Grobler, D., van Altena, J.J., Bertschinger, H., Kirkpatrick, J.F., 2000. Immunocontraception of African elephants. Nature London 407, 6801.
Abstract: Sum: Based on a South African trial of 41 adult females, it is argued that pZP immunocontraception is a humane method to control elephant populations without behavioural side effects.

Fritsch, G., Göritz, F., Hermes, R., Jewgenow, K., Maltzan, J., Hildebrandt, T.B., 2000. Physiology of sexual maturity in male elephants. Reprod Dom Anim 35, 26.

Hermes, R., Olson, D., Goritz, F., Brown, J.L., Schmitt, D.L., Hagan, D., Peterson, J.S., Fritsch, G., Hildebrandt, T.B., 2000. Ultrasonography of the estrous cycle in female African elephants (Loxodonta africana). Zoo Biology 19, 369-382.
Abstract: The endocrinology of the elephant oestrous cycle has been well characterized, but little emphasis has been placed on evaluating corresponding changes in the reproductive tract. Ultrasound was used to document changes in reproductive tract morphology throughout the oestrous cycle in four cycling female African elephants. During a 7-month period, frequent ultrasound examinations (n=190) during the luteal and non-luteal phase were compared with serum progesterone and luteinizing hormone (LH) concentrations during a 7-month period. Ultrasonographic images documented vaginal and cervical oedema and changes in mucus consistency during the non-luteal