renal

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

Garner, M.M., Helmick, K., Ochsenreiter, J., Richman, L.K., Latimer, E., Wise, A.G., Maes, R.K., Kiupel, M., Nordhausen, R.W., Zong, J.C., Hayward, G.S., 2009. Clinico-pathologic features of fatal disease attributed to new variants of endotheliotropic herpesviruses in two Asian elephants (Elephas maximus)
119. Vet. Pathol. 46, 97-104.
Abstract: The first herpesviruses described in association with serious elephant disease were referred to as endotheliotropic herpesviruses (EEHV) because of their ability to infect capillary endothelial cells and cause potentially fatal disease. Two related viruses, EEHV1 and EEHV2, have been described based on genetic composition. This report describes the similarities and differences in clinicopathologic features of 2 cases of fatal endotheliotropic herpesvirus infections in Asian elephants caused by a previously unrecognized virus within the betaherpesvirus subfamily. EEHV3 is markedly divergent from the 2 previously studied fatal probosciviruses, based on polymerase chain reaction sequence analysis of 2 segments of the viral genome. In addition to ascites, widespread visceral edema, petechiae, and capillary damage previously reported, important findings with EEHV3 infection were the presence of grossly visible renal medullary hemorrhage, a tropism for larger veins and arteries in various tissues, relatively high density of renal herpetic inclusions, and involvement of the retinal vessels. These findings indicate a less selective organ tropism, and this may confer a higher degree of virulence for EEHV3

Lozi, H., Goodwin, T.E., Rasmussen, L.E.L., Whitehouse, A.M., Schulte, B.A., 2009. Sexual dimorphism in the performance of chemosensory investigatory behaviours by African elephants (Loxodonta africana). Behaviour 146, 373-392.
Abstract: Sexual dimorphism in morphology can be accompanied by behavioural differences between the sexes. We examined if investigatory behaviour involving the trunk of African elephants showed sexual dimorphism. Males compete and search for females, but they have a lengthy period of development before they are socially viable mates. Receptive females are relatively rare. We hypothesized that males would display higher rates of chemosensory behaviour following puberty than females. Because males disperse, they were hypothesized to be more likely to contact elephants outside their kinship group. We observed the trunk tip, chemosensory behaviours of African elephants at Addo Elephant National Park, South Africa. For 208 elephants, we found no significant differences in state behaviours around waterholes by age or sex. Yet, older elephants were more likely to investigate the environment and elephant excrement than younger animals. Males were more likely to investigate urine and faeces than females. Only post-puberty animals contacted non-family with males investigating both sexes, while investigations by and to females only involved post-puberty males. Overall, the probability of performing chemosensory behaviours depended on age and sex. Male elephants appear more reliant than females on signals in urine and faeces with ensuing inspections of individuals through trunk tip contacts.

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

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

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.

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.

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

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

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

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.

Goodwin, T.E., Eggert, M.S., House, S.J., Weddell, M.E., Schulte, B.A., Rasmussen, L.E., 2006. Insect pheromones and precursors in female African elephant urine
450. Journal of Chemical Ecology 32, 1849-1853.
Abstract: Using automated solid-phase dynamic extraction and gas chromatography-mass spectrometry, our search for urinary chemical signals from ovulatory female African elephants (Loxodonta africana) has revealed the bark beetle aggregation pheromones frontalin, exo-brevicomin, and endo-brevicomin, as well as their precursors and the aphid alarm pheromones (E,E)-alpha-farnesene and (E)-beta-farnesene. Enantiomeric ratios for brevicomins have been determined. Prior discovery of common insect/elephant pheromones in Asian elephants, namely, (Z)-7-dodecenyl acetate and frontalin, suggests that the present findings may yield valuable insights into chemical communication among African elephants

Wiedner, E., Alleman, R., Isaza, R. Urinalysis in Asian elephants (Elephas maximus).
2006 Proceedings American Association of Zoo Veterinarians. 265. 2006.
Ref Type: Conference Proceeding
Abstract: Elephants have been reported with renal disease, and arthritic elephants are often maintained on long-term administration of non-steroidal anti-inflammatory drugs which have been associated with renal injury in other species. Therefore, establishment of the reference values of normal urine would be a useful tool to elephant health care. Routine urinalysis was performed on urine collected from healthy female adult Asian elephants (Elephas maximus, n=30). Elephants were included in the study if they had normal serum urea nitrogen and creatinine serum concentrations, no history of urinary tract disease, and had not received any medication within the past 3 mo. Urinalysis included gross description of urine color and clarity, measurement of pH and specific gravity, biochemical analysis and sediment evaluation. Presence of protein was assessed by the sulfosalicylic acid tubidimetric test. All urine samples were submitted for aerobic bacterial culture. Initial findings included identification of calcium carbonate crystals in the urine of most of the animals in this study; some (n=5) contained small numbers of struvite crystals (magnesium ammonium phosphates). Most samples were alkaline and clarity ranged from clear to flocculent. Specific gravity tended to be fairly low, and isosthenuria was frequent. Trace bilirubin was detected in some samples. No ketonuria or proteinuria was found. Glucosuria, which has been reported as a transient and normal finding in captive elephants, was not detected in any samples.

Williams, M.F., 2006. Morphological evidence of marine adaptations in human kidneys. Med Hypotheses 66, 247-257.
Abstract: Amongst primates, kidneys normally exhibiting lobulated, multipyramidal, medullas is a unique attribute of the human species. Although, kidneys naturally multipyramidal in their medullary morphology are rare in terrestrial mammals, kidneys with lobulated medullas do occur in: elephants, bears, rhinoceroses, bison, cattle, pigs, and the okapi. However, kidneys characterized with multipyramidal medullas are common in aquatic mammals and are nearly universal in marine mammals. To avoid the deleterious effects of saline water dehydration, marine mammals have adaptively thickened the medullas of their kidneys--which enhances their ability to concentrate excretory salts in the urine. However, the lobulation of the kidney's medullary region in marine mammals appears to be an adaptation to expand the surface area between the medulla and the enveloping outer cortex in order to increase the volume of marine dietary induced hypertonic plasma that can be immediately processed for the excretion of excess salts and nitrogenous waste. A phylogenetic review of freshwater aquatic mammals suggest that most, if not all, nonmarine aquatic mammals inherited the medullary pyramids of their kidneys from ancestors who originally inhabited, or frequented, marine environments. So this suggest that most, if not all, aquatic mammals exhibiting kidneys with lobulated medullas are either marine adapted--or are descended from marine antecedents. Additionally, a phylogenetic review of nonhuman terrestrial mammals possessing kidneys with multipyramidal medullas suggest that bears, elephants and possibly rhinoceroses, also, inherited their lobulated medullas from semiaquatic marine ancestors. The fact that several terrestrial mammalian species of semiaquatic marine ancestry exhibit kidneys with multipyramidal medullas, may suggest that humans could have, also, inherited the lobulated medullas of their kidneys from coastal marine ancestors. And a specialized marine diet in ancient human ancestry could, also, explain the reactivation and enumeration of corporeal eccrine sweat glands and the copious secretion of salt tears. The substantial loss of genetic variation in humans relative to other hominoid primates, combined with the apparent isolation of early Pliocene human ancestors from particular retroviruses that infected all other African primate species, may suggest that such a semiaquatic marine phase, during the emergence of Homo, may have occurred on an island off the coast of Africa during the early Pliocene.

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

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

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

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.

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

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., 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.

Lazar, J., Rasmussen, L.E., Greenwood, D.R., Bang, I.S., Prestwich, G.D., 2004. Elephant albumin: a multipurpose pheromone shuttle
691. Chem. Biol. 11, 1093-1100.
Abstract: (Z)-7-dodecenyl acetate (Z7-12:Ac) is present in the urine of female Asian elephants (Elephas maximus) approaching ovulation and functions as a female-to-male sex pheromone. Here we show that a significant fraction of the pheromone in the urine is bound to a protein, elephant serum albumin (ESA), and provide evidence for key physiological functions of urinary ESA. Our biochemical and behavioral experiments suggest a three-fold role of ESA in pheromone signaling: (1) transporting Z7-12:Ac from serum into urine; (2) extending the presence of the pheromone in the environment without hampering detection; and (3) targeting pheromone delivery to chemosensory organs through localized release of the ligand induced by a pH change. The exploitation of albumin in pheromone transport clearly distinguishes the elephant from other mammals studied, and complements the uniqueness of elephant anatomy, physiology, and behavior

Leal, W.S., 2004. Pheromone unwrapping by pH flip-flopping
692. Chem. Biol. 11, 1029-1031.
Abstract: The Asian elephant utilizes the same sex pheromone as a number of moth species, (Z)-7-dodecen-1-yl acetate encapsulated in a serum-derived albumin. The chemical signal is emitted in the urine and received in the mucus of the trunk. The unwrapping of the package is pH mediated

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.

Suarez, R.K., Darveau, C.A., Childress, J.J., 2004. Metabolic scaling: a many-splendoured thing
656. Comp Biochem. Physiol B Biochem. Mol. Biol. 139, 531-541.
Abstract: Animals at rest and during exercise display rates of aerobic metabolism, VO2, that represent mainly the sum of mitochondrial respiration rates in various organs. The relative contributions of these organs change with physiological state such that internal organs such as liver, kidney and brain account for most of the whole-body VO2 at rest, while locomotory muscles account for >90% of the maximum rate, VO2max, during maximal aerobic exercise. Mechanisms that regulate VO2 are complex and the relative importance of each step in a series, estimated by metabolic control analysis, depends upon the level of biological organization under consideration as well as physiological state. Despite this complexity, prominent single-cause models propose that metabolic rates are supply-limited and that the scaling of supply systems provides a sufficient explanation for the allometric scaling of metabolism. We argue that some assumptions, as well as current interpretations of the meaning (or consequences) of these constraints are flawed, i.e., elephants do not have lower mass-specific basal or maximal rates of aerobic metabolism because their mitochondria are more supply-limited than those of shrews. Animals do not violate the laws of physics, and the allometric scaling of supply systems would be expected, to some extent, to be matched by capacities for (and rates of) energy expenditure. But life is not so simple. Animals are so diverse that to do justice to metabolic scaling, it is also necessary to consider the scaling of energy expenditure. It is by doing so that models of metabolic scaling can be consistent with current paradigms in metabolic regulation and accommodate the range of inter- and intraspecific exponents found in nature. The "allometric cascade," a first attempt at such an accounting, was a source of great satisfaction to Peter Hochachka. It was the last door that he helped open to comparative physiologists before he said goodbye

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.

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

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

Pothiwong, W., Kamonrat, P., Uthaichotiwan, P., 2003. A morphological study and diagnotic ultrasonography of Asian elephant kidney. Thai Journal of Veterinary Medicine 33, 79-88.

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

Endo, H., Akihisa, N., Sasaki, M., Yamamoto, M., Arishima, K., 2002. The renal structure in an Asian elephant (Elephas maximus). Anat Histol Embryol 31, 269-272.
Abstract: The renal structure of a female Asian elephant (Elephas maximus) was observed in both macroscopic and light microscopic levels. The left kidney was elongated-ellipse in shape, whereas the right appeared round. The left kidney was 31 cm in cranio-caudal length, 21 cm in medio-lateral length, and 2950 g in weight. The right kidney was 34 cm in cranio-caudal length, 22 cm in medio-lateral length, and 3250 g in weight. The external appearance showed the six separated renal lobes in both sides of the kidney. The four pairs of the lobes were fused in the deepest region in both sides of kidney, so we considered it as an incompletely lobated kidney in this species. We observed the proximal and distal urinary tubules in histological sections. Many renal corpuscles consisted of the glomerulus and Bowman's capsule. Many mesangial cells and some podocytes were confirmed in each glomerulus; however, Bowman's capsules were larger than those in other mammalian species.

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.

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

Singh, V.N., 2002. Symptomatic study of haemorrhagic septicaemia in elephant in Mudumalai Wildlife Sanctuary, Tamil Nadu. Indian Forester 128, 1089-1100.
Abstract: Symptoms of haemorrhagic septicaemia, a dreaded disease in elephant is recorded in this study which reveals systematic spread of Oedema from jowl to throat, neck, brisket, abdomen and perenical regions. It also records the changes in character/colour of dung, urine, eye, tongue, trunk, body temperature, feeding habit and body condition along with treatment given to cure the disease.

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.

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.

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

Rasmussen, L.E.L., Krishnamurthy, V., 2001. Urinary, temporal gland and breath odors from Asian elephants of Mudumalai National Park. Gajah 20, 1-7.

Schmid, J., Heistermann, M., Ganslosser, U., Hodges, J.K., 2001. Introduction of foreign female Asian elephants (Elephas maximus) into an existing group: behavioural reactions and changes in cortisol levels. Animal-Welfare 10, 357-372.
Abstract: The present study examined the extent to which the introduction of three female Asian elephants (aged 3, 11 and 27 years) into a group of 5 (1 male, 4 female) elephants at Munster zoo, Germany, affects the behaviour and urinary cortisol levels of the animals involved. At Munster, only the females were monitored , while the bull was mainly kept separate. Behavioural observations were carried out before transfer and during the six-month period following transfer. Urine samples were collected regularly from each elephant during the whole observation period. All the elephants showed behavioural changes to the process of introduction. The transferred animals increased their social behaviour after arrival in the foreign zoo. Two of them showed an increase in stereotypies and one a reduction in stereotypes. The elephants at Munster reacted with decreased frequencies of stereotypies and increased frequencies of social behaviour and manipulation/exploration behaviour. Six months after transfer, three of the four elephants at Munster and one of the three transferred elephants showed nearly the same behavioural activity pattern as before transfer. One female still showed elevated stereotypic behaviour. From the four elephants in which cortisol measurements could be reliably performed (two of the transferred elephants and two elephants at Munster), only one individual at Munster responded to the process of introduction with a short-term elevation in urinary cortisol levels. One elephant showed a negative correlation between locomotion and cortisol levels and one a positive correlation between stereotypies and cortisol levels. Taken together, the results suggest that transfer and introduction caused some stress responses in the elephants, but that stress was neither prolonged nor severe. Serious welfare problems may have been prevented through individual behavioural coping mechanisms and former experience with stressful situations.

Singer, M.A., 2001. Of mice and men and elephants: metabolic rate sets glomerular filtration rate. Am J Kidney Dis 37, 164-178.
Abstract: Allometric scaling deals with the functional consequences of changes in size or scale among geometrically dissimilar animals (ie, animals differing in proportions). For adult mammals ranging in size and proportion from mouse to elephant, the data describe an interdependent set of functions consisting of metabolism (measured as metabolic rate), glomerular filtration rate (GFR), effective renal plasma flow, excretion of nitrogenous waste products, cardiac output, and pulmonary function-related variables. Within this set of functions, evidence indicates that metabolic rate is the primary process. One important design feature is given by the ratio of GFR to metabolic rate. Because this ratio is independent of size, it can be generalized to all mammals in this series. The numeric value of this ratio gives the optimal GFR for each unit of metabolic rate. A simple hypothesis is proposed: metabolic rate, the primary process, sets GFR. This relationship is unidirectional. A decrease in GFR, for example, caused by nephron loss, should not lead to a change in metabolic rate. This hypothesis was tested in four natural experiments: human growth and development, thyroid dysfunction, chronic renal failure, and hibernation. The results are consistent with this hypothesis.

Bielecki, W., Sobczak-Filipiak, M., Pietrak, M., Goslawski, J., Blaszczak, B., 2000. A case of nephritis in an elephant. Zycie-Weterynaryjne 75, 197-199.

Lazar, J., Prestwich, G.D., Rasmussen, L.E.L., 2000. Urinary and trunk mucus protein carriers of (Z)-dodecenyl acetate, the sex pheromone of the Asian elephant. Chemical Senses 25, 603.

Malik, M.R., Shrivastav, A.B., Jain, N.K., Vaish, R., 2000. Morphometry of kidney of elephant. Indian Journal of Veterinary Anatomy 12 , 101-102.

Malik, M.R., Shrivastava, A.B., Jain, V.K., Rakhi, V., 2000. Lobar pattern of kidneys of elephant. Indian Journal of Veterinary Anatomy 12, 18-22.

Singer, M.A., Morton, A.R., 2000. Mouse to elephant: biological scaling and Kt/V. Am J Kidney Dis 36, 306-309.
Abstract: The construct Kt/V is used by the nephrology community in prescribing dialysis dose. The concerns that have been raised as to what value of V to use in the calculation of Kt/V touch on the more central question of whether filtration rate should be normalized by a parameter other than V. Within the animal kingdom, a number of physiological variables scale to body size according to an equation of the form Y = YoMb, where Yo is a constant, M is body mass, and b is a scaling exponent. Glomerular filtration rate (GFR) in mammals weighing from 30 g to 503 kg scales to body weight with an exponent of 0.77. Hence, GFR per unit body weight (or Kt/V) decreases significantly with increasing body size. Metabolic rate also scales to body size in a wide range of mammals according to the same general equation and with a scaling exponent of 0.75. Because GFR and metabolic rate scale to body mass with virtually the same exponent, a ratio of the two yields a constant independent of body size. We propose that the ratio (filtration rate/metabolic rate) replace Kt/V. Such a ratio would underscore the linkage between filtration rate (and dialysis therapy) and the metabolic demands of the body.

1999. Equine Medicine and Surgery. Mosby, St. Louis MO USA.

Fies, M., Heistermann, M., Hodges, J.K., 1999. Patterns of urinary and fecal steroid excretion during the ovarian cycle and pregnancy in the African elephant (Loxodonta africana). Gen Comp Endocrinol 115, 76-89.
Abstract: The aims of the present study were to (I) determine the relative abundance of the 5alpha-reduced progestins 5alpha-pregnane-3-ol-20-one (5alpha-P-3OH) and 5alpha-dihydroprogesterone (5alpha-DHP) and progesterone (P4) in African elephant feces and to establish improved fecal progestin assays for monitoring ovarian function; and (ii) describe longitudinal profiles of urinary and fecal progestin and estrogen metabolites during pregnancy. Matched urine and fecal samples were collected weekly from six adult females throughout 18 nonfertile cycles and two complete pregnancies (89 and 93 weeks duration). Fecal samples were lyophilized and extracted with 80% methanol in water and immunoreactive 5alpha-P-3OH, 5alpha-DHP, and P4 and (for pregnant females only) estrone (E1) and estradiol (E2) determined by enzyme immunoassay. Urine samples were hydrolyzed, ether-extracted, and assayed for 5alpha-P-3OH, E1, and E2. HPLC cochromatography of fecal extracts with various radioactive progestin tracers confirmed the presence of large amounts of both 5-reduced progestins (5alpha-P-3OH > 5alpha-DHP) but not of P4. 5-Reduced progestins (but not P4) were excreted in a cyclic pattern and levels were significantly correlated with urinary 5alpha-P-3OH. Fecal 5alpha-P-3OH showed the more pronounced and consistent luteal-phase elevation and a better correspondence to urine with respect to timing of the luteal-phase rise. Fecal and urinary 5-reduced progestins increased gradually during early pregnancy to maximum values around week 40-45. Levels gradually declined during the second half of pregnancy, reaching baseline values 2 days before parturition. Urinary estrogens did not show any cyclic pattern during the preconception period and levels remained low during the first 30 weeks of gestation. Thereafter, there was a rapid 10- to 20-fold increase to maximum values at mid-pregnancy, followed by a gradual decline to birth. There was no mid-pregnancy elevation in fecal estrogens, but there was a modest increase in E1 during the second half of gestation.

Gaeth, A.P., Short, R.V., Renfree, M.B., 1999. The developing renal, reproductive, and respiratory systems of the African elephant suggest an aquatic ancestry. Proc Natl Acad Sci U S A 96, 5555-5558.
Abstract: The early embryology of the elephant has never been studied before. We have obtained a rare series of African elephant (Loxodonta africana) embryos and fetuses ranging in weight from 0.04 to 18.5 g, estimated gestational ages 58-166 days (duration of gestation is approximately 660 days). Nephrostomes, a feature of aquatic vertebrates, were found in the mesonephric kidneys at all stages of development whereas they have never been recorded in the mesonephric kidneys of other viviparous mammals. The trunk was well developed even in the earliest fetus. The testes were intra-abdominal, and there was no evidence of a gubernaculum, pampiniform plexus, processus vaginalis, or a scrotum, confirming that the elephant, like the dugong, is one of the few primary testicond mammals. The paleontological evidence suggests that the elephant's ancestors were aquatic, and recent immunological and molecular evidence shows an extremely close affinity between present-day elephants and the aquatic Sirenia (dugong and manatees). The evidence from our embryological study of the elephant also suggests that it evolved from an aquatic mammal.

Gual-Sill, F., Pickard, A.R., Holt, W.V., Green, D. Preliminary Results of Non-Invasive Monitoring of the Estrous Cycle in Female Asian Elephants (Elephas maximus) through Fecal Steroid Analysis. 1999 Proceedings American Association of Zoo Veterinarians. 87-92. 1999. 1999.
Ref Type: Conference Proceeding
Abstract: For a number of years, estrous cycle monitoring and pregnancy detection in the Asian elephant has been performed using urinary steroid hormone metabolite analysis; this technique presents some practical problems. Monitoring the reproductive status through fecal steroid analysis is possible in this and many other species. The steroid metabolite profiles of female Asian elephants were monitored by enzyme-linked immunosorbent assay (ELISA), to provide detailed information about the estrous cycle and pregnancy in this species, and to investigate causes of reproductive failure. Fecal and matched urine samples were non-invasively collected regularly for 6 mo from captive female Asian elephants. (n = 4 cyclic; n = 1 acyclic). The samples were frozen at -20ºC. Gas Chromatography and Mass Spectroscopy (GC-MS) procedures were used to investigate the steroid hormone metabolite profile and to identify the major excretory metabolites; no steroid metabolites were found in the concentrated extracted feces of this species using the currently available methodology. The fecal pregnanetriol profile observed in three of the cyclic females showed a clear relation with their matched urinary pregnanetriol profile and a cyclic pattern was demonstrated. Fecal pregnanetriol values increased from an overall mean of 94.67 ng/g of dry feces (+/- 13.24, range 31.5 - 219.12 ng/g) during the inter-luteal period to a luteal phase mean of 334.61 ng/g dry feces (+/- 43.48, range 34.35 - 1035.1 ng/g). All the data collected from the fecal and urinary analysis of pregnanetriol in all five individuals investigated demonstrated a significant relationship between urinary and fecal pregnanetriol. The acyclic individual showed a mean fecal pregnanetriol concentration of 84.91 ng/g (+/- 13.06) and values ranged from 33.17 ng/g to 211.42 ng/g. Fecal steroid hormone metabolite analysis for monitoring estrous cycles in Asian elephants may be used in the future to monitor free-roaming, wild or semi-wild individuals as well as those in captivity to assist reproductive and conservation programs of this highly endangered species.

Rasmussen, L.E.L., 1999. Evolution of chemical signals in the Asian elephant, Elephas maximus: behavioural and ecological influences. Journal of Biosciences 24, 241-251.
Abstract: In antiquity, the Asian elephant, Elephas maximus, gradually spread southward and eastward to become a successfully surviving, ecologically dominant megaherbivore in the tropical environment of south-east Asia. The changing physical environment forced dynamic fluxes in its social structure and altered its metabolism. Such events shaped the production and ultimately the stability of certain chemicals released by body effluvia. Some of these chemicals took on significance as chemical signals and/or pheromones. This article demonstrates by experimental and observational evidence, and hypothesizes based on speculative reasoning, how and why specific chemical signals evolved in the modern Asian elephant. Evidence, including the functional criteria required by elephant social structure and ecology, is presented for the hypothesis that the recently identified female-emitted, male-received sex pheromone, (Z)-7-dodecenyl acetate evolved first as a chemical signal. Subsequently, the cohesiveness and harmony of small, matriarchally-led female groups were strengthened by a female-to-female chemical signal, recently defined behaviourally. The looser societal structure of freer, roaming males also became bounded by chemical signals; for the males, breath and temporal gland emissions, as well as urinary ones function in chemical signaling. Basic knowledge about elephant chemical signals is now linking chemical information to behaviour and beginning to demonstrate how these signals affect elephant social structure and enable the species to cope with environmental changes.

Rasmussen, L.E.L., Perrin, T.E., 1999. Physiological correlates of musth: lipid metabolites and chemical composition of exudates. Physiology and Behavior 67, 539-549.
Abstract: Physiological changes related to lipid metabolism, behaviour and chemicals released in body exudates were studied during musth in the Asian elephant (Elephas maximus) as a case study. During musth, changes in serum testosterone and triglyceride concentrations followed similar patterns, with the former increasing sooner than the latter. Deviant behaviour increased during changing androgen levels. The observed high concentrations of testosterone were positively and significantly correlated with increased triglycerides. Lipase activity elevated significantly immediately before and after musth. Blood pH increased significantly in alkalinity. Urine and temporal gland secretions released variable amounts of compounds, some of which may be chemical signals. During musth, temporal gland and urinary exudates demonstrated increased acetone and other ketones indicative of lipid metabolic alterations. Large quantities of nonmethane hydrocarbons, especially 2-butanone, were released from the seemingly dry orifice of the temporal gland before the start of over musth and before maximum blood elevations were observed; isoprene release was similar. However, maximal acetone levels occurred simultaneously in blood, temporal gland secretions, and urine. Metabolically, musth is a series of interwoven, changing stages of increasing and decreasing hormones and lipid-related constituents. Released chemicals can be quantitatively related to these internal physiological events; some observed behaviours appear to result from altered chemical signals.

Rasmussen, L.E.L., Schulte, B.A., 1999. Ecological and biochemical constraints on pheromonal signaling systems in Asian elephants and their evolutionary implications. In: Johnston, R.E., Muller-Schwarze, D., Sorenson, P.W. (Eds.), Advances in Chemical Communication in Vertebrates 8. Kluwer/Academic/ Plenum Press, pp. 49-62.
Abstract: The Asian elephant is an unusual example of how intraspecies chemical communication helps maintain societal cohesiveness within familial and herd units. The amount of multi-directional chemical communication is surprising, because long-lived elephants have a highly organized society, are capable of trans-generational passage of information, possess a sophisticated vocalization system, and are capable of complex learning and tool use. This paper discusses the ecological, behavioral, and biochemical aspects of chemical signals in elephants from an evolutionary perspective. Diverse bodily emissions are utilized as intraspecies chemical signals (including pheromones), often with imposed biochemical constraints. In this chapter, chemosignals released from the temporal gland secretions and breath of male Asian elephants in musth and a urinary female-to-male preovulatory pheromone are utilized as examples of these concepts. Furthermore, specific behavioral and biochemical studies with (Z)-7-dodecenyl acetate (a female-to-male urinary sex pheromone) demonstrate that social context significantly influences responsivity (demonstrated by field studies in Myanmar) and that additional biochemical requirements, perhaps lipocalin-like proteins, may be required for full bioactivity. The remarkable convergent evolution of (Z)-7-dodecenyl acetate, both structurally and functionally, in elephants and Lepidoptera, allows the use in elephant studies of effective biochemical tools developed for insect investigations. This convergence of chemical signaling systems of elephants and insects has several interesting implications.

Schulte, B.A., Rasmussen, L.E., 1999. Signal-receiver interplay in the communication of male condition by Asian elephants. Animal Behaviour 57, 1265-1274.
Abstract: Signal design and meaning are dependent on the condition of the sender and receiver as well as the response of the receiver. This study examined (1) whether female Asian elephants, Elephas maximus, can distinguish between a conspecific male in musth and nonmusth states using urinary signals, (2) how the oestrous condition of the female affects discrimination, and (3) correlation of female responses with the testosterone level of the male. Musth is a rut-like state displayed by healthy adult male elephants. Males in musth dominate nonmusth males and may be preferred by females as mates. Urine was collected from two captive male Asian elephants during nonmusth periods and from one of these males during times of musth. Samples of musth and nonmusth urine and control liquids were placed in an elephant enclosure weekly for 16 weeks, the length of a female oestrous cycle. Primary response behaviours were approach and four trunk-tip motions, namely sniff, check, place and flehmen. Musth urine consistently elicited greater responses than nonmusth and control samples. Females were more responsive during their follicular (sexually receptive) than luteal (unreceptive) stages of oestrus. Furthermore, females appeared to be sensitive to the degree of musth as responses increased with rising serum testosterone levels of the male donor. Chemical signals from males are a likely source of honest signals related to status and reproductive condition. Female elephants appear capable of detecting differences in a male based upon urinary chemosignals.

Wrench, A., Kock, N.D., Gordon, S.J.G., Turner, A.D., Fries, D.S., 1999. Investigation of selected plants as possible causes of flaccid trunk paralysis in free-ranging elephants (Loxodonta africana) in Zimbabwe. Zimbabwe Veterinary Journal 30, 93-97.
Abstract: Extracts from four plants collected from the southern shore of Lake Kariba were injected into Balb/c laboratory mice in an attempt to establish the cause of flaccid paralysis of the trunk in elephants. Behavioural and neurological changes were noted in some instances after injection. Histopathological changes similar to those found in affected elephants, including renal tubular karyomegaly and myocardial degeneration and necrosis, were found in mice exposed to Boerhavia diffusa. Patchy gliosis and cerebral oedema were present in mice treated with Blumea geriepina and Geigeria schnizii. The study indicates the need for further investigation into the role of plant intoxication as a possible cause of trunk paralysis in elephants.

Hodges, J.K., 1998. Endocrinology of the ovarian cycle and pregnancy in the Asian (Elephas maximus) and African (Loxodonta africana) elephant. Animal Reproduction Science 53, 3-18.
Abstract: In reviewing the literature, this paper assesses the current level of understanding of the hormonal control of the ovarian cycle and pregnancy in the
elephant. Data are compiled from two genera, Elephas maximus and Loxodonta africana, recognizing differences where known to exist. Measurements of various steroid and peptide hormones, but most notably immunoreactive progesterone (iP), indicate an overall cycle length of 15-16 weeks, comprising an 8-11-week luteal phase and a shorter inter-luteal (follicular) period of 4-6 weeks. Oestrogen related phenomena suggest the occurrence of two (possibly more) waves of follicular development with a 3-week periodicity, although oestrogen levels are low and fluctuate without clear pattern. The inter-luteal period is characterized by two discrete LH peaks also 3 weeks apart, but only the second is associated with an iP rise. Ovulation rate is variable and additional luteal structures lacking ovulation points are probably formed each cycle. The post-ovulatory period is associated with elevated progestogen levels, mainly 5alpha-reduced compounds, while progesterone itself is a minor secretory product. A revised model for the elephant ovarian cycle is presented. Detailed information on the endocrinology of pregnancy is lacking. Elevated progestins beyond the normal luteal phase indicate that conception extends the functional lifespan of the CL, with maximum steroidogenic activity between 3-15 months. The nature of the luteotrophic support is not known and placental gonadotrophins have not been conclusively demonstrated. Progestins fluctuate at or slightly above luteal phase values throughout pregnancy. There is a marked increase in prolactin levels after 16-24 weeks and a relationship with oestrogen secretion may exist since circulating and urinary levels show a progressive increase from 6-8 months. A placental contribution to progestin secretion is likely, although direct evidence is lacking. Considerable enlargement of foetal gonads during the second half of pregnancy in African elephants suggests a possible steroidogenic function, but none has been defined. The endocrine events leading to parturition are unknown. In the Asian elephant, a fall in iP precedes parturition; oestrogen levels decline and cortisol increases at the time of birth. The paper concludes with a brief selection of priorities for future research.

Rasmussen, B., 1998. The Chemical Identification of a Preovulatory Pheromone: A Reproductive Chemosignal from Female to Male Asian Elephants. Journal of the Elephant Managers Association 7, 52-56.

1997. Laboratory Procedures for Veterinary Technicians. Mosby, St. Louis MO.

Heisterman, M., Trohorsch, B., Hodges, J.K., 1997. Assessment of ovarian function in the African elephant (Loxodonta africana) by measurement of 5-reduced progesterone metabolites in serum and urine. Zoo Biology 16, 273-284.
Abstract: We have previously shown that 5-pregnane-3,20-dione (5-DHP) and 5-pregnane-3-ol-20-one (5-OH) are the major luteal and circulating progestins in the African elephant. Therefore, the aim of the present study was to determine 1) circulating levels and patterns of secretion of 5-DHP in relation to progesterone (P4) throughout the ovarian cycle, 2) the presence and relative abundance of 5-reduced progestins in urine and 3) whether their measurement in urine would provide a non-invasive method for monitoring luteal function. Urine samples were collected weekly throughout a total of 13 ovarian cycles from 5 females. In addition, matched blood samples were collected during 6 cycles from 2 of the 5 animals. All hormone measurements were carried out by enzymeimmunoassay following extraction. Urine was hydrolized prior to extraction. Profiles of P4 and 5-DHP in serum followed a similar cyclic pattern and both measurements were significantly correlated (r = 0.78-0.98, mean 0.89, P < 0.001). Concentrations of 5-DHP were, however 10-20-fold higher than those if P4. In addition, 5-DHP measurements showed a more pronounced luteal phase increase than that of immunoreactive P4. HPLC co-chromatography confirmed the presence of large amounts of 5-P-3-OH in urine as a single immunoreactive peak, whereas 5-DHP was present in very low levels and measurable only as one of several immunoreactive substances. Measurements of urinary 5-P-3-OH were significantly correlated to serum 5 -DHP measurements in each of the 6 cycles (r = 0.72-0.93, mean 0.81, P < 0.001), whereas correlation coefficients between urinary and serum 5-DHP values were generally lower (r = 0.34-0.83, mean 0.69) and significant in only 4 of 6 cycles. Accordingly, only urinary excretion of 5-P-3-OH, but not of 5-DHP, exhibited a clear cyclic pattern, with consistently low levels of 0.15-.020 g/mg Cr in the follicular phase and 10-fold elevated levels (1.8-2.2 g/mg Cr) in the luteal phase. Based on the intervals between successive luteal phase increases in urinary 5-P-3-OH, a mean cycle length of 14.1+/- 1.8 weeks, comprising a follicular phase of 5.0 +/- 0.9 weeks and a luteal phase of 9.10+/-01.4 weeks was determined for the 13 cycles studied. The results indicate that measurements of 5-P-3-OH in urine provide a reliable non-invasive method for monitoring luteal function in the African elephant.

Niemuller, C., Shaw, H.J., Hodges, J.K., 1997. Pregnancy determination in the Asian elephant (Elephas maximus): A change in the plasma progesterone to 17 hydroxyprogesterone ratio. Zoo Biology 16, 415-426.
Abstract: The measurement of circulating progesterone (P) is widely used to confirm and monitor pregnancy in the captive Asian elephant. Consistently elevated progesterone (P) concentrations for a minimum of 12 weeks can be used as a positive indication of pregnancy, although two instances of prolonged P secretion during the cycle have been recorded [Rubel, 1987; Olsen et al, 1994]. Previously, we demonstrated that pregnanetriol was the major urinary gestagen metabolite enabling the noninvasive monitoring of the reproductive cycle of the Asian elephant [Niemuller et al., 1993] as well as pregnancy [unpublished data]. the importance of this unusual urinary metabolite triggered an investigation into the secretion of circulating concentrations of 17 hydroxyprogesterone (17 OHP) during pregnancy and reproductive cycles, as this steroid is the only steroid precursor of pregnanetriol. Comparison of the profiles between 17 OHP and P during early pregnancy (n = 5) and nonconceptive cycles (n = 15) demonstrated a decline in 17 OHP , but not P, as early as week 3 postmating (designated as week 1) and lasting up to week 13. Otherwise, secretions of 17 OHP mimicked P concentrations throughout pregnancy and in nonconceptive cycles. Examination of the mean ratio values of 17 OHP to P demonstrated a significant drop in the ratio during weeks 2-7 of early pregnancy from >/= 0.7 to < 0.7 compared with the same time period in a nonreceptive cycles (p < 0.05m N = 5), A 2x2 table analysis of the 17 OHP:P ratio during weeks 2-7 indicated that the possibility of a false positive or false negative result was 3.4 and 6.5%, respectively, based on the sensitivity and specificity of the test. Overall, gestation lengths of the pregnancies completed during this study (N = 4) ranged from 91 to 98 weeks, with a mean of 93+/-2.9 (SEM) weeks. A birth-to-conception interval of 47 weeks was noted in one animal. The results described in this report provide additional data on the reproductive endocrinology of the pregnant Asian elephant and also present the earliest means to date of determining pregnancy by analysis of the 17 OHP:P ratio.

Kirkpatrick, J.F., Turner, J.W., Jr., Liu, I.K., Fayrer-Hosken, R., 1996. Applications of pig zona pellucida immunocontraception to wildlife fertility
control. J Reprod Fertil Suppl 50, 183-189.
Abstract: A unique application of pig zona pellucida (PZP) immunocontraception is the control of wildlife populations. A native PZP vaccine has been successfully applied to wild horse and donkey populations. A single annual booster inoculation was capable of maintaining contraception. Seven consecutive years of PZP treatment in wild mares resulted in no detectable debilitating side effects, and reversibility of contraception has been documented among mares treated for up to 4 consecutive years. Long-term treatment (5-7 years) is associated with some ovulation failure and depressed urinary oestrogen concentrations. Complex social behaviours in horses were unaffected by treatment. PZP immunocontraception has also been successfully applied to white-tailed deer, with no detectable changes in ovarian histology after 2 years of treatment. Seventy-four species of captive zoo animals have been treated with the PZP vaccine, with documented success in 27 species, including members of the orders
Perissodactyla (Equidae), Artiodactyla (Cervidae, Capridae, Giraffidae, Bovidae), and Carnivora (Ursidae, Mustelidae, Felidae). Immunocytochemistry studies have demonstrated a high degree of crossreactivity between anti-PZP antibodies and African elephant zona pellucida. The need for a one-inoculation form of the vaccine has led to the incorporation of PZP into lactide-glycolide microspheres, which cause a delayed release of the PZP. PZP immunocontraception of wildlife has potential because of (1) > 90% effectiveness, (2) the ability for remote delivery, via darts, (3) reversibility after short-term use, (4) a wide breadth of effectiveness across many species, (5) a lack of debilitating side-effects even after long-term treatment, and (6) minimal effects upon social behaviours.

Murray, S., Bush, M., Tell, L.A., 1996. Medical management of postpartum problems in an Asian elephant (Elephas maximus) cow and calf. Journal of Zoo and Wildlife Medicine 27, 255-258.
Abstract: An 18-yr old female Asian elephant (Elephas maximus) gave birth to a 120-kg female calf following 22 mo of gestation. Immediately after parturition, the cow became agitated and aggressive towards the calf. Before the keepers were able to safely intervene and remove the calf, the cow stepped on the calf's head and right front leg. Within 30 min, the cow calmed down, allowing the calf's safe reintroduction under close keeper supervision and control. The cow had a retained placenta, poor mammary development, and low milk production. The calf's injuries, in combination with the cow's low milk production, impeded the calf's ability to nurse and gain weight. Within 10 days, the calf lost 10% of its weight. Serum protein electrophoresis indicated failure of passive transfer of maternal immunoglobulin. On day 10, the calf received a transfusion of concentrated immunoglobulin extracted and concentrated from the cow's previously banked plasma. On day 13, the calf developed a urinary tract infection, as diagnosed by white blood cells and bacteria in the urine. Following immunoglobulin administration and antibiotic therapy, clinical signs slowly resolved and the calf gained weight. The cow passed the fetal membranes during parturition, but the placenta was retained. Despite prophylactic systemic antibiotics and vaginal flushing, the cow became depressed and developed a leukocytosis and anemia. A mucopurulent vaginal discharge and ventral edema were noted on day 3, and milk production was minimal. Because decreased milk production has been reported as a common sequel to retained placenta, efforts were focused on removing the placenta. Intermittent oxytocin therapy on days 2-14 did not result in expulsion of the placenta and produced only transient abdominal contractions and minimal increases in milk letdown. On day 15, 10 mg estradiol cypionate was administered i.m. followed by 200 IU oxytocin i.v. An additional 10 IU oxytocin was administered i.v. on day 16. The friable placenta was palpable within the vaginal vault on day 17. The remaining placenta was removed by gentle traction applied by a modified weighted pressure cuff. Once the placenta was removed, the cow's clinical problems slowly resolved and the calf continued to gain weight.

Rasmussen, L.E.L., Lee, T.D., Roelofs, W.L., Zhang, A., Daves, G.D., 1996. Insect pheromone in elephants. Nature 379, 1.

Brown, J.L., Wemmer, C.M., Lehnhardt, J., 1995. Urinary Cortisol Analysis for Monitoring Adrenal Activity in Elephants. Zoo Biology 14 , 533-542.
Abstract: Cortisol was measured in dichloromethane-extracted elephant urine using an ¹²⁵I solid-phase radioimmunoassay (RIA). The cortisol RIA was validated by demonstrating 1) parallelism between dilutions of pooled urinary extracts and the standard curve, 2) significant recovery of exogenous cortisol added to elephant urine, and 3) a relationship between changes in the peripheral and urinary cortisol after an adrenocorticotropin hormone (ACTH) challenge. One African (Loxodonta africana) and one Asian (Elephas maximus) elephant were given three injections of ACTH (1.25 mg) at 2 h intervals. Serum cortisol increased four- to eightfold within 30 min after the first injection and peaked (nine- to twelvefold increase) after the second injection. Serum concentrations began to decline 2-3 h after the last injection but were still approximately fourfold higher than baseline at the end of the collection period (hour 8). In the urine, cortisol concentrations were increased in the first sample postinjection (1.5 - 4 h) and peaked twenty- to fortyfold by ~6 h. Urinary cortisol remained elevated at 8 h, but returned to baseline by the following morning. Analysis of high performance liquid chromatography fractions of extracted urine revealed that immunoactivity was associated with free cortisol (~90% of total immunoactivity) and a more polar, unidentified metabolite. A method for preserving urine was developed to allow storing unfrozen samples. One pool of urine from each of one African and two Asian elephants was divided into aliquots, placed in tubes containing absolute ethanol (10%), sodium azide (0.1%) or distilled water (control), and frozen after 0, 1 , 2, 3, 4, 6, 8, 10, 12, and 24 weeks of storage at ~25^ºC. In unpreserved samples, cortisol concentrations were reduced 46% by 2 weeks and 95% by 24 weeks. In contrast, ethanol- and sodium azide-preserved samples retained 100 and 95% of cortisol immunoactivity through 8 weeks and 93 and 85% of activity through 12 weeks, respectively. We infer from these data that changes in urinary cortisol excretion in the elephant reflect fluctuations in adrenal activity and may be a useful indicator of stress. Additionally, urine samples can be collected and stored unfrozen for at least 2 months before any appreciable loss in cortisol immunoactivity occurs, a finding potentially useful to field application of this technique.

Brown, J.L., Lehnhardt, J., 1995. Serum and urinary hormones during pregnancy and the peri- and postpartum period in an Asian elephant (Elephas maximus). Zoo Biology 14, 555-564.

Maluf, N.S.R., 1995. Kidney of elephants. Anatomical Record 242, 491-514.
Abstract: BACKGROUND: Elephants are an important and isolated order. Their kidneys need substantial investigation and hitherto have not been portrayed even by a pyelogram. METHODS: Pyelograms and injection of vessels with colored acrylic emulsions were done initially. Dissection was under fiberoptics using a dissecting microscope with frequent measurements. Special areas were cut for microscopy (light and electron) and photography. Glomerular counts were done by macerating weighted pieces of cortex and later finding the cortical fraction of the renal parenchyma. RESULTS: The elephant kidney is devoid of dorsoventral symmetry. It is composed of 8 +/- 2 lobes separated by fine interlobar septa. There is no reduction of lobes with maturity. The pelvis bifurcates at the sinus into primary branches or infundibula which dispatch a secondary branch or infundibulum into every lobe. Interlobar arteries and veins, nerves, fat, and connective tissue generally accompany every secondary infundibulum into its lobe. A major branch of the renal artery may perforate the renal capsule and course to the cortico-medullary (C-M) border independently of the secondary infundibulum to that lobe. The number of glomeruli per kidney is approximately 15 x 10(6). In adults the glomerular mass is 4.9 +/- 0.5% of the renal parenchyma and 6.7 +/- 0.3% of the cortex. Areae cribrosae occur generally at low papillae. They are the outlets of numerous terminal collecting ducts which may be accompanied by a tubus maximus (T.M.) A T.M. of diameter 1.6 mm and length 10 mm may act as the only substitute for an area cribrosa. Wide anastomoses between the two main renal veins occur within the renal sinus. Intralobar arteries and veins often course right through the outer medulla to and from, respectively, the C-M border. CONCLUSIONS: Anatomically, an elephant's kidneys appear to be able to concentrate urine only moderately. Their kidneys tend to resemble those of the manatee but not of the dugong.

1994. Veterinary Laboratory Medicine. Iowa State University Press, Ames, IA.

Lodwick, L.J., Dubach, J.M., Phillips, L.G., Brown, C.S., Jandreski, M.A., 1994. Pharmacokinetics of amikacin in African elephants (Loxodonta africana). Journal of Zoo and Wildlife Medicine 25, 367-375.
Abstract: Two adult females were given single i.v. injections of 8 mg/kg. Trials using 3 mg/kg and 6 mg/kg i.m. were conducted with three adult females. Serum concentrations of amikacin were measured serially over a 24-49 h period. After i.v. administration of 8 mg/kg, the elimination half-lives (t0.5) were 4.0 and 3.7 h, the volumes of distribution at steady state were 0.21 and 0.18 litres/kg, and total body clearances were 41.8 and 40.8 ml/h/kg. At i.m. doses of 3 and 6 mg/kg, the peak concentrations observed ranged from 4.8 to 8.4 µg/ml and 14.2 to 21.8 µg/ml, respectively. The time at observed peak concentration was between 1 and 3 h, and t0.5 ranged from 3.8 to 5.9 h for the lower dose and from 3.7 to 6.3 h for the higher dose. Following the single dose trials, one elephant was treated with amikacin at a dose of 7 mg/kg i.m. at 24 h intervals for 21 days, and serum amikacin concentrations were determined 2 to 4 times on each of 11 days. Mean (SD) peak serum concentration for this elephant was 19.0±2.8 µg/ml, and mean serum concentration at 24 h (through) was 1.7±0.4 µg/ml. There was indication in this one elephant of a mild, reversible renal tubular insult based on a slight transient elevation in serum creatinine and the presence of casts in the urine. These changes resolved soon after the end of treatment. These preliminary results suggest that amikacin administered at 6-8 mg/kg i.m. once every 24 h would be appropriate for elephants with bacterial infections suspected to be susceptible to amikacin.

Plouzeau, E., daCunha, S., Shaw, H.J., 1994. The ovarian cycle in Asian and African elephants (Elephas maximus and Loxodonta africana). Techniques for the monitoring of female fertility in captivity. Revue de Medecine Veterinaire 145, 905-911.
Abstract: A discussion. During the oestrous cycle, circulating blood progesterone in elephants alternates between high and low concentrations over a cycle of 16±2 weeks. Some of the data have suggested a 3-week cycle and other data a 15- to 16-week cycle. Captive females show no signs of oestrus, although males show a Flehmen-like response to female urine, which is inversely related to plasma progesterone concentration. Pregnancy is associated with a sustained increase in circulating progesterone concentration and with an increase in total oestrogen, prolactin and oestrogen concentrations during the second half of pregnancy. The most reliable method of monitoring the oestrous cycle and pregnancy is by weekly analysis of plasma progesterone in both species, or of its urinary metabolite (pregnanetriol) in Asian elephants.

Wallace, C., Byron, T.H., Foerner, J.J., Weston, H., Kilpatrick, J., Jastremski, M.S. Clinical case report: the medical management and treatment of a 36 year old premiparturient Asian elephant cow with a dystocia and following a Caesarian section. 1994.
Ref Type: Unpublished Work
Abstract: The medical history and management of a 36 year old premiparturient Asian elephant (Elephas maximus indicus) with a dystocia requiring a caesarian section are discussed. The examination and complete medical evaluation to determine the health of the cow and viability and position of the calf are described. The medical management of the post-operative complications and the changes in complete blood counts, differential, serum biochemistry values and urinalysis prior to the elephant's death are described. Complications included peritonitis with systemic sepsis, renal failure, and hepatic failure. Pertinent necropsy findings on the cow included severe diffuse subacute peritonitis, uterine transmural necrosis, diffuse renal tubular nephrosis, and hepatic centrolobular degeneration.

Niemuller, C.A., Shaw, H.J., Hodges, J.K., 1993. Non-invasive monitoring of ovarian function in Asian elephants (Elephas maximus) by measurement of urinary 5beta-pregnanetriol. Journal of Reproduction and Fertility 99, 617-625.
Abstract: An enzymeimmunoassay for 5beta-pregnanetriol is described. Immunoreactive pregnanetriol concentrations were significantly correlated with the concentrations of progesterone (0.98, n = 269, P<0.01) and 17alpha-hydroxyprogesterone (0.95, n = 205, P<0.01), the precursor of pregnanetriol. The duration of cycles as determined by measurements of plasma progesterone, plasma 17alpha-hydroxyprogesterone and urinary pregnanetriol averaged 15.54±1.5 weeks (23 cycles), 15.21±1.7 weeks (15 cycles) and 15.45±0.94 weeks (20 cycles) respectively. The results demonstrated that it is possible to monitor ovarian function in Asian elephants by measuring urinary pregnanetriol concentration.

Okewole, P.A., Oyetunde, I.L., Irokanulo, E.A., Chima, J.C., Nwankpa, N., Laleye, Y., Bot, C., 1993. Anthrax and cowdriosis in an African elephant (Loxodonta africana). Veterinary Record 133, 168.
Abstract: In February 1992, a 15-year-old African elephant died; it was the second elephant that had died within 2 weeks at a wildlife park. Clinical signs in both elephants included frequent micturition, restlessness and weakness of the hindquarters with frequent falls. PM examination revealed ecchymosis of the epicardium, atrioventricular surfaces of the heart and serosal surfaces of the intestines and bladder with sloughing of intestinal mucosae. The liver was enlarged, ecchymotic and congested. A serosanguinous exudate with fibrin was present in the thoracic and abdominal cavities. The meninges were congested. Bacillus anthracis was cultured from tissue samples and from tissue samples from guineapigs inoculated with broth cultures of the tissue samples from the elephant. Cowdria ruminantium was identified in stained impression smears from the elephant brain. This appears to be the first report of the simultaneous occurrence of anthrax and cowdriosis in an African elephant.

Turczynski, C.J. The endocrinology of musth in the male Asiatic elephant (Elephas maximus): Serum estradiol, serum LH and serum, fecal and urinary testosterone. 1993. College Station, TX. USA, Texas A&M University.
Ref Type: Thesis/Dissertation

Czekala, N.M., Roocroft, A., Bates, M., Allen, J., Lasley, B.L., 1992. Estrogen metabolism in the Asian elephant (Elephas maximus). Zoo Biology 11, 75-80.
Abstract: Estradiol-17B metabolism was studied in two female Asian elephants (Elephas maximus). In an initial study, 500 microCi of tritiated estradiol-17B was injected iv into a single animal, and 0, 30, and 60 min serum samples were collected as well as all excreted urine and feces for 24hr. In a second study, 1.5 mg unlabeled estradiol-17B was injected iv into a second animal and 0, 5, 15, 30 and 60 min serum samples and a 30 min urine sample were collected postinjection. Analyses of samples from both studies demonstrated a rapid conversion of free estradiol to conjugated forms in the serum. The first (5 min) serum sample following the injection of unlabelled estradiol contained unconjugated estradiol:conjugated estradiol: conjugated estrone at a ratio of 60:29:10, respectively, and at 30 min a ratio of 33:43:24. The urinary estrogen metabolites were in the conjugated form with an estradiol:estrone ratio of 60:40. No radiolabelled estrogen was found in the fecal samples during the 24 hr following administration of the radiolabelled estradiol. These data indicate a rapid clearance of circulating free estradiol in the elephant, with a major metabolite in the serum and urine being estradiol conjugate.

Hagenbeck, D. Attempts to monitor oestrus and pregnancy in Indian elephants by urine testing. Diagnostische Zyklusansprache und Trachtigkeitsbestimmung beim asiatischen Elefanten (Elephas maximus). 1-66. 1992. Hannover, Germany, Tierarztliche Hochschule Hannover.
Ref Type: Thesis/Dissertation

Chakraborty, A., Islam, S., Gogoi, A.R., Chaudhury, B., 1991. A note on clinical examination of elephants in Manas Tiger Project and Kaziranga National Park in Assam. Zoos' Print Journal November.
Abstract:
The diseases of elephants have been recorded by Steel (1885) and Evans (1910) and till then many works have done on elephants. Compared to the extent of work done in other states of our country, very little investigation seems to have been done in Assam. The present communication relates the report of investigation carried out in the domesticated elephants of Manas Tiger Project and Kaziranga National Park in Assam.

1990. The story of Babe, the Asian elephant. Veterinary Viewpoints 2.

Diaz-Samayoa-de-Aguirre, L. Sex hormones in blood plasma, urine and faeces from female and male Indian elephants. Vergleichende Hormonuntersuchungen in Blutplasma, Harn und Kot beim weiblichen und mannlichen asiatischen Elefanten (Elephas maximus). 1-145. 1990. Hannover, Germany, Tierarztliche Hochschule.
Ref Type: Thesis/Dissertation

Mainka, S.A., Lothrop, C.D., Jr., 1990. Reproductive and hormonal changes during the estrous cycle and pregnancy in Asian elephants. Zoo Biology 9, 411-419.
Abstract: Serum progesterone and urinary total estrogen concentrations were determined weekly to bi-weekly in 2 female Asian elephants for 96 weeks. The mean estrous interval was approximately 16 weeks in the nonpregnant animal. A total of 5 cycles were observed in the 96 weeks study period. The serum progesterone concentration ranged from 150 pg/ml to greater than 350 pg/ml during the luteal phase of the estrous cycle. The serum progesterone was elevated for 8-12 weeks of the 16 week estrous cycle. The urinary total estrogen concentration ranged from less than 10 to greater than 300 pg/microgram creatinine. The second animal was pregnant at the beginning of the study period. The serum progesterone concentration was elevated (> 100 pg/ml) in the pregnant animal until parturition. The urinary total estrogens increased from approximately 50 pg/microgram creatinine to greater than 400 pg/microgram creatinine during the first year of pregnancy and remained elevated until parturition. Estrous cycling had not resumed by 3 months post partum.

Brannian, J.D., Griffin, F., Terranova, P.F., 1989. Urinary androstenedione and luteinizing hormone concentrations during musth in a mature African elephant. Zoo Biology 8, 165-170.
Abstract: Musth has not been well documented in captive African elephants. A 37-year-old African bull elephant in the Kansas City Zoological Park was observed during periods of behavioral musth and non-musth. Androstenedione and luteinizing hormone (LH) concentrations in urine were measured by radioimmunoassay. Urinary androstenedione and LH levels were significantly higher in musth urine than in non-musth samples. A positive correlation (P> 0.001) existed between urinary LH and androstenedione concentrations. These results indicate that musth can occur in a zoo-maintained African elephant and that urinary androgen levels are elevated during musth, possibly as a result of LH stimulation of testicular steroidogenesis.

Czekala, N.M., Roocroft, A., Bates, M. Estrogen metabolism in the Asian elephant (Elephas maximus). Biology of Reproduction 40[suppl.1], 119. 1989.
Ref Type: Abstract
Abstract: Understanding follicular development in elephants has been confounded by an inability to detect serum estradiol (E₂) in cyclic patterns. Serum levels remain < 20 pg/ml throughout the cycle. To understand E₂ dynamics in the elephant, two metabolic studies were initiated. First 500 microCi tritiated E₂ was injected (iv). All urine and feces voided during the following 24 hrs. and two blood samples (30 min and 2 hrs) were collected. No radiolabel appeared in feces during the 24 hrs. Urinary radiolabel analyzed by HPLC appeared 100% conjugated, 52% E₂-3-conjugate (E₂C) and 48% estrone conjugate (E₁C). Peak levels appeared in urine 30 min post-injection, decreased rapidly during the next 2 hrs, then gradually to 16.5 hrs when levels stabilized. Serum radiolabel at 30 min appeared as conjugated and free (66:34). In the second study, unlabelled E₂ (1.5 mg) was injected (iv). Blood was taken at 0, 5, 15, 30, and 60 min and urine at 30 min. In serum, peak estrogen levels appeared at 5 min (E₂:E₂C:E₁C, 38:39:22 or 61:38 conjugated:free). Half-life of E₂ is ca.10 min and 60 min for E₂C. E₁C peaked at 15 min and declined by 20% in 45 min. Urine yielded similar results as the label study (E₂C:E₁C, 60:40). Daily urine E₂C and weekly serum E₂C and progesterone (P) were measured by RIA in five mature female elephants. Preliminary results indicated elevated levels of E₂C during the luteal phase similar to serum P profile. The follicular phase contains a biphasic profile of E₂C. These results suggest that E₂ is rapidly converted to E₂C and excreted in urine.

Dahl, K.D., Czekala, N.M., Hsueh, A.J.W. Measurement of urinary bioactive follicle-stimulating hormone (FSH) levels during reproductive cycles in diverse mammalian species. Biology of Reproduction 40[suppl.1], 119. 1989.
Ref Type: Abstract
Abstract: Recently an in vitro granulosa cell aromatase bioassay (GAB) was used to measure FSH in serum and urine samples (JCEM, March, 1987). We now adapted the GAB assay to measure urinary bio-FSH levels in conjunction with the determination of urinary immunoreactive pregnanediol-3-glucoronide (PdG) and/or estrone conjugates (EC). Daily urine samples were collected from 2 monkey species, lion-tailed macaque (Macaca silenus) and golden monkey (Rhinopithecus roxellana), and 3 ungulates, giraffe (Giraffa camelopardalis), okapi (Okapi johnstoni) and Asian elephant (Elephas maximus). The monkeys displayed a follicular phase rise in EC followed by a midcycle bio-FSH surge which declined as PdG concentrations increased during the luteal phase. Although both monkey species had similar cycle lengths (30-35 days), the golden monkey's follicular phase was much longer and displayed 2 bio-FSH peaks compared to 1 peak for the macaque. Although EC was not detectable in the ungulates, a midcycle FSH surge was followed by a luteal phase increase in PdG. The closely related giraffe and okapi had similar cycle lengths (16-17 days), follicular phase lengths (8-9 days), and only 1 follicular phase FSH peak. Conversely, the elephants had cycles lasting 16-19 weeks, and multiple FSH peaks were observed during the 5-6 week follicular phase. Thus, 1) in contrast to the single follicular phase increase and preovulatory surge of bio-FSH seen in the macaque, giraffe and okapi, the finding of multiple follicular phase peaks in the golden monkey and elephants suggests the involvement of complex regulatory mechanisms; 2) the GAB assay provides a noninvasive and practical method for monitoring reproductive cycles in endangered species for future breeding programs.

Morris, P.J., Held, J.P., Jensen, J.M. Clinical pathologic features of chronic renal failure in an African elephant (Loxodonta africana). Proc.1st.Intl.Conf.Zool.Avian Med. 468-472. 1987.
Ref Type: Conference Proceeding

Taylor-Robinson, D., Furr, P.M., Tully, J.G., Barile, M.F., Moller, B.R., 1987. Animal models of Mycoplasma genitalium urogenital infections. Israel Journal of Medical Sciences 23, 561-564.
Abstract: Male and female animals were inoculated urogenitally with Mycoplasma genitalium, recovered originally from men with nongonococcal urethritis. Mice, hamsters and male rhesus monkeys (Macaca mulatta) were resistant. Male cynomolgus monkeys (Macaca fascicularis) were not as sensitive as male chimpanzees (Pan troglodytes): 9 of 11 developed an obvious genital tract infection, some shedding organisms for more than 18 weeks. M. genitalium was recovered from the blood of two of them when large numbers of organisms were in the urethra. Most of the chimpanzees colonized with the organisms had increased numbers of polymorphonuclear leukocytes in the genital tract and developed a fourfold or greater antibody response. Female squirrel monkeys (Saimiri sciureus) and female tamarins (Saguinus mystar) exhibited low-level genital tract infections following intravaginal inoculation, whereas marmosets (Callithrix jacchus) and chimpanzees developed prolonged infections after similar inoculation: thus, female chimpanzees shed organisms for 12 to 15 weeks. Marmosets and grivet monkeys (Ceropithecus aethiops) developed salpingitis with antibody responses after intraoviduct inoculation, and baboons (Papio anubis) developed parametritis after intracervical inoculation. The results offer substantial evidence for the pathogenicity of M. genitalium for the urogenital tract of subhuman primates, and suggest that the microorganism may have a role in human genital tract infections.

Jensen, J. Paralumbar kidney biopsy in a juvenile African elephant. Proc.Am.Assoc.Zoo Vet. 17. 1986.
Ref Type: Conference Proceeding
Abstract: Same case as Ref # 305.

Kahl, V.A.L. The study of the reproductive cycle of the African elephant (Loxodonta africana) through correlation of behavior and cytology with serum and urinary hormonal patterns. 1986. Nebraska, Omaha, USA, University of Nebraska.
Ref Type: Thesis/Dissertation

Munson, L., Heuschele, W., O'Banion, M.K., Sundberg, J.P., Oosterhuis, J.E., 1986. Polyp in the urogenital canal of an African elephant. Journal of the American Veterinary Medical Association 189, 1190-1191.

Poole, J.H., Kasman, L.H., Ramsay, E.C., Lasley, B.L., 1984. Musth and urinary testosterone concentrations in the African elephant (Loxodonta africana). Journal of Reproduction and Fertility 70, 255-260.
Abstract: Urine samples were obtained from free-ranging African elephants that were considered to be in and out of musth. Testosterone concentrations, measured by radioimmunoassay were significantly greater in males that were in or around the time of behavioral musth. This study supports a correlation between the observed behavioral characteristics of musth and urinary testosterone levels.

Rasmussen, L.E., Schmidt, M.J., Henneous, R., Groves, D., Daves, G.D.Jr., 1982. Asian bull elephants: flehmen-like responses to extractable components in female elephant estrous urine. Science 217, 159-162.
Abstract: Flehmen-like responses (urine tests) are one of the characteristic behavioral reactions of male Asian elephants (Elephas maximus) to cow elephants in estrus. Components of the urine of estrous cow elephants were extracted with organic solvents and partially purified by chromatography and shown to evoke Flehmen-like responses when they were presented to adult bulls.

Ramsay, E.C., Lasley, B.L., Stabenfeldt, G.H., 1981. Monitoring the estrous cycle of the Asian elephant (Elephas maximus), using urinary estrogens. American Journal of Veterinary Research 42, 256-260.
Abstract: The estrous cycle of the Asian elephant (Elephas maximus) was monitored by analysis of urinary estrogens. Daily morning urine samples were analyzed for estrone (E₁), estradiol (E₂), and total immunoreactive estrogen (E_T). The E_T values were shown to correlate poorly with E₁ and E₂ and failed to reveal any patterns of reproductive cycling. Daily E₁ and E₂ values, indexed by creatinine concentrations, demonstrated cyclic profiles in those samples of sufficient concentrations. The technique offered a simple, noninvasive method for determining ovarian function in the elephant.

Nirmalan, G. Physiological peculiarties of elephants. State Level Workshop on Elephants. 21-24. 1979. India, College of Veterinary and Animal Sicences, Kerala Agricultural University.
Ref Type: Conference Proceeding

Fujimoto, E., Koto, N., Imori, T., Nakama, S., 1971. Gonadotropin in the urine of a pregnant Indian elephant -- a case report. Zoologica 55, 73.
Abstract: In 1963, at Takarazuka Zoo, Japan, a young female Indian elephant became pregnant, and in May, 1965, she gave birth to a very large stillborn calf (weighing 133.3 kg, male). The time of conception was problematical, but it was assumed as April or May of 1963, hence the gestation period may have been 24 or 25 months, a little longer than average. Pregnancy diagnosis was attempted during the early and middle gestation period. For exploration, an urinary gonadotropin was checked by the Friedman and Aschheim-Zondek tests on the whole urine samples collected twice in August 1963. Results showed apparently positive results in both tests. However, the samples collected in May and September, 1964, showed negative in three tests, including a male frog (Rana) reaction which was subjected to the concentrated urine samples. So, probably a gondotropic substance many have been excreted in urine of this elephant at some time of the early pregnancy, and this may be more like FSH than LH in its activity.

McCullagh, K.G., 1969. The growth and nutrition of the African elephant I. Seasonal variations in the rate of growth and the urinary excretion of hydroxyproline. East African Wildlife Journal 7, 85-90.

Orbanyi, J., Stohl, G., 1962. The urine of the elephant. UNKNOWN 321-328.

Evans, G.H., 1961. Elephants and Their Diseases: A Treatise on Elephants. Government Printing, Rangoon, Burma.

Simon, K.J., 1959. Further studies on the urine of elephants. Indian Veterinary Journal 36, 209-212.

Simon, K.J., 1958. A preliminary study of the urine of elephants. Indian Veterinary Journal 35, 345.

Schulte, T.L., 1937. The genito-urinary system of the Elephas indicus male. American Journal of Anatomy 61 , 131-157.

Eales, N., 1929. The anatomy of a foetal African elephant, Elephas africanas (Loxodonta africana). Part III. The contents of the thorax and abdomen, and the skeleton. Transactions of the Royal Society of Edinburgh 56, part I, 203-246.

Evans, G.H., 1910. Elephants and Their Diseases: A Treatise on Elephants. Government Printing, Rangoon, Burma.

Paterson, A.M., 1898. The genito-urinary organs of the female Indian elephant. Journal of Anatomy and Physiology 12, 582-604.

Miall, L.C., Greenwood, F., 1879. The anatomy of the Indian elephant. Part III alimentary canal and its appendages. Journal of Anatomy and Physiology 13, 17-50.

Watson, M., 1872. Contributions to the anatomy of the Indian elephant (Elephas indicus), Part II. Urinary and generative organs. Journal of Anatomy and Physiology 7, 60-74.

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