Fecal Estrogen Research Articles

Noninvasive monitoring of female reproductive hormone metabolites in the endangered European mink (Mustela lutreola)

This study examined the reproductive physiology of female European mink (Mustela lutreola) to augment the available information on estrus, ovulation, and pregnancy with the long-term goal of supporting ex situ breeding management of this highly endangered species. Fecal reproductive hormone metabolites were measured using EIAs for estrogen and 20-oxo-pregnane metabolites. Seasonal hormone profiles were established. A comparison of hormone fluctuations in pregnant and nonpregnant females showed that both estrogen and 20-oxo-pregnane metabolites were significantly elevated during gestation, which is 42 days in length. Delayed implantation or embryonic diapause does not occur in this species. Litter size was correlated with 20-oxo-pregnane levels but not with estrogen concentrations. During lactation, 20-oxo-pregnane metabolite levels remained higher than in nonpregnant females. The breeding season was characterized by peaks in vaginal cornified cells and fecal estrogen metabolite levels. Up to four peaks in estrogen levels were identified and confirmed that European mink are seasonally polyestrous. The results of 20-oxo-pregnane measurements indicated that hCG can be applied to induce ovulation. With the establishment of this noninvasive method, we present a new tool to support population management of this species.

The use of altrenogest to avoid hyperestrogenism after eCG-hCG ovulation induction in southern tigrina (Leopardus guttulus)

The goal of this study was to optimize an ovulation induction protocol for use with artificial insemination in the southern tigrina (Leopardus guttulus). The specific aims were to report the efficacy of using altrenogest, an oral progestin (Regumate, MSD Animal Health, Merck Animal Health), to suppress ovarian activity and prevent follicular hyperstimulation and hyperestrogenism after the administration of exogenous eCG and hCG. To monitor ovarian responses, fecal estrogen and progestogen metabolites were quantified by enzyme immunoassay in females before and after intramuscular administration of 200-IU eCG and 150-IU hCG in two trials, 4 months apart. During the first trial, there was no use of altrenogest, only the eCG-hCG ovulation induction protocol. In the second trial, the ovulation induction protocol was preceded by the administration of oral altrenogest for 14 days (minimum of 0.192 mg per kg per day). Altrenogest administration resulted in a suppression of follicular activity in three out of six females before eCG-hCG administration on the basis of lower mean estrogen concentrations (P < 0.05). It also resulted in four out of six females presenting lower fecal estrogen metabolite concentrations (P < 0.05) after ovulation induction, and two out of six individuals showed a reduction (P < 0.05) in postovulatory fecal progestogen metabolite concentrations, all when compared to the same female's cycles without the progestin. Fecal estrogen metabolite concentrations were closer to baseline in 50% of these individuals after altrenogest and eCG-hCG treatments when compared to basal concentrations before gonadotropins without the use of altrenogest. This study reported that use of altrenogest in southern tigrina can suppress ovarian activity and avoid hyperestrogenism after administration of eCG and hCG treatment. However, not all females responded uniformly, so more studies are needed to increase the efficacy of ovulation induction for use with artificial insemination in this species.

What is the best test for cryptorchidism?

Cryptorchidism is a common heritable condition in horses and ponies and while retained testicles rarely release viable sperm, Leydig cells produce testosterone leading to undesirable stallion-like behaviour. Endocrine assays are frequently required to definitively diagnose retained testicular tissue. A literature search of the Medline database was performed using the PubMed search engine. The search term ‘equine cryptorchidism’ was entered and the results filtered using the contents of the abstracts to retain articles relating specifically to diagnostic testing for cryptorchidism. The initial search yielded 139 articles which after filtering were reduced to 6 articles included for analysis. Retrospective analysis of clinical cases comprising level IV evidence was used to evaluate the use of serum oestrone sulphate and serum testosterone (alone and after human chorionic gonadotropin (hCG) administration) for the detection of cryptorchidism (Cox et al. 1986). Leung et al. (2011) also analysed a large number of normal horses but few cryptorchid animals (n = 25, only 18 confirmed) were diagnosed using urine steroid profiling. The remaining prospective experimental studies evaluated the use of serum testosterone following administration of hCG (Silberzahn et al. 1989), serum anti-Müllerian hormone (AMH) (Claes et al. 2013), serum androstenedione (Illera et al. 2003) and faecal oestrogens (Palme et al. 1994) to distinguish between gelded, entire and cryptorchid horses. This data was then variably extrapolated to provide inferred relevance for the use of each test for the detection of suspected cryptorchid animals. Of the 4 prospective experimental studies those by Silberzahn et al. (1989) and Palme et al. (1994) lack sufficient numbers or appropriate methodology to provide quality evidence. Illera et al. (2003) evaluated the use of serum androstenedione but inconsistency in their results significantly weakens the evidence provided by this paper. The most recent experimental study by Claes et al. (2013) provides the strongest evidence for the use of serum AMH. The quality of the evidence provided by the retrospective studies (Cox et al. 1986; Leung et al. 2011) is weak as the cryptorchid status of all the animals in the study, particularly those testing negative, was not confirmed. Testosterone may be measured directly but serum levels vary greatly between individuals and fluctuate with age and season (Cox et al. 1986; Claes et al. 2013) making it unsuitable as a definitive test. Measurement of testosterone pre and 30 min post administration of hCG was reported to be more reliable with an accurate diagnosis achieved in 95% of cases (Cox et al. 1986). However, other studies have questioned the reliability of this elevation in testosterone so soon after hCG administration (Silberzahn et al. 1989; Illera et al. 2003) and accuracy may be improved by testosterone measurement up to 72 h post hCG (Silberzahn et al. 1989). This claim has not been validated and such a time delay is less suitable for use in clinical practice. Measurement of serum oestrone sulphate in horses and ponies (not donkeys) >3-years-old was 96% accurate in weak early retrospective studies (Cox et al. 1986). However, evaluation of serum oestrone sulphate in a more clearly defined population yielded reduced positive predictive values (PPV) and negative predictive values (NPV) of 85% and 91% respectively for this test (Illera et al. 2003). Illera et al. (2003) reported serum androstenedione to have increased accuracy (92%) compared with both oestrone sulphate (86%) and testosterone pre and post hCG stimulation (88%) for the detection of cryptorchids. However, this finding is weakened by anomalies in the results of the study and, on the basis of this, evidence serum androstenedione cannot be recommended. Anti-Müllerian hormone (AMH) is produced in elevated amounts from Sertoli cells in cryptorchid testes (Ball et al. 2008) making it a potentially reliable and specific test for the presence of testicular tissue. Claes et al. (2013) evaluated the serum AMH concentrations in stallions, geldings and cryptorchids. Serum AMH concentrations were significantly higher in cryptorchid stallions (32.7 ± 5.3 ng/ml) compared with intact stallions (14.7 ± 2.4 ng/ml); geldings were below or fractionally above the lower limit of detection for the test. Only animals of >2 years of age were included in the study and AMH levels require investigation in cryptorchid pubertal animals. The calculated biological half-life was 1.5 days (Claes et al. 2013) making AMH a useful test if there is doubt over the success of a recent castration procedure. The evidence for many of the available endocrine assays is based on poor quality retrospective or experimental studies. Currently, the strongest evidence exists for the use of AMH for the detection of cryptorchidism, although the test warrants validation in a large number of confirmed cryptorchid animals and in animals <2 years of age. No conflicts of interests have been declared.

Assessment of ovarian activity in captive goral (Naemorhedus griseus) using noninvasive fecal steroid monitoring

To date, there is no information on gonadal steroidogenic activity of female goral (Naemorhedus griseus), a threatened species of Thailand. Captive goral populations have been established to produce animals for ex situ conservation and reintroduction, but as yet none are self-sustaining. The objectives of the present study were to (1) determine the influence of season on ovarian steriodogenic function; and (2) examine the relationship between gonadal hormone excretion and sexual behaviors throughout the year. Fecal samples were collected 5 to 7 days/wk for 15 months from 8 adult females housed at Omkoi Wildlife Breeding Center in Thailand and analyzed for ovarian steroid metabolites using validated enzyme immunoassays. Observations of sexual behaviors and mating were conducted each morning for 30 min/session. Based on fecal estrogen and progestagen metabolite concentrations, the overall estrous cycle length was about 21 days, with a 2- to 3-day follicular phase and an 18- to 20-day luteal phase. Sexual behaviors, most notably tail-up, increased for 2 to 3 days during the time estrogens were elevated during mating. Fecal progestagens were elevated during luteal phases and increased further during gestation, which lasted approximately 7 months. The lactation period was 5 months, and females were anestrus for 2 to 5 of those months, with the exception of one that cycled continuously throughout. Two females conceived around 2 months postpartum and so were pregnant during lactation. Birth records over the past 21 years indicated young are born throughout the year. This combined with the hormonal data suggests that female gorals are not strongly seasonal, at least in captivity, although there was considerable variation among females in estrogen and progestagen patterns. In conclusion, fecal steroid metabolite monitoring is an effective means of assessing ovarian function in this species and will be a useful tool for breeding management and planned development of assisted reproductive techniques such as artificial insemination and embryo transfer.

Measuring fecal testosterone in females and fecal estrogens in males: Comparison of RIA and LC/MS/MS methods for wild baboons (Papio cynocephalus)

The development of non-invasive methods, particularly fecal determination, has made possible the assessment of hormone concentrations in wild animal populations. However, measuring fecal metabolites needs careful validation for each species and for each sex. We investigated whether radioimmunoassays (RIAs) previously used to measure fecal testosterone (fT) in male baboons and fecal estrogens (fE) in female baboons were well suited to measure these hormones in the opposite sex. We compared fE and fT concentrations determined by RIA to those measured by liquid chromatography combined with triple quadropole mass spectrometry (LC/MS/MS), a highly specific method. Additionally, we conducted a biological validation to assure that the measurements of fecal concentrations reflected physiological levels of the hormone of interest. Several tests produced expected results that led us to conclude that our RIAs can reliably measure fT and fE in both sexes, and that within-sex comparisons of these measures are valid: (i) fTRIA were significantly correlated to fTLC/MS/MS for both sexes; (ii) fTRIA were higher in adult than in immature males; (iii) fTRIA were higher in pregnant than non-pregnant females; (iv) fERIA were correlated with 17β-estradiol (fE2) and with estrone (fE1) determined by LC/MS/MS in pregnant females; (v) fERIA were significantly correlated with fE2 in non-pregnant females and nearly significantly correlated in males; (vi) fERIA were higher in adult males than in immature males. fERIA were higher in females than in males, as predicted, but unexpectedly, fTRIA were higher in females than in males, suggesting a difference in steroid metabolism in the two sexes; consequently, we conclude that while within-sex comparisons are valid, fTRIA should not be used for intersexual comparisons. Our results should open the field to important additional studies, as to date the roles of testosterone in females and estrogens in males have been little investigated.

Induction of ovarian activity and ovulation in an induced ovulator, the maned wolf (Chrysocyon brachyurus), using GnRH agonist and recombinant LH

Assisted reproductive techniques, such as ovarian manipulation and artificial insemination, are useful for enhancing genetic management of threatened wildlife maintained ex situ. In this study, we used noninvasive fecal hormone monitoring to investigate (1) the influence of pairing with a male on endocrine responses of female maned wolves (Chrysocyon brachyurus) to a GnRH agonist (deslorelin) and (2) the efficiency of recombinant LH (reLH) on ovulation induction in females housed alone. Deslorelin (2.1 mg Ovuplant) was given to females that were either paired with a male (n = 4) or housed alone (n = 7); the implant was removed 7 to 11 days postimplantation. Three of seven singleton females were injected with reLH (0.0375 mg) on the day of implant removal, whereas the remaining females (n = 4) did not receive the additional treatment. Fecal samples were collected 5 to 7 days/wk from all females starting 11 days prior to hormone insertion until at least 70 days post implant removal for a total of 11 hormone treatment cycles. Fecal estrogen and progestagen metabolites were extracted and analyzed by enzyme immunoassay. Evidence of ovulation, demonstrated by a surge of estrogen followed by a significant rise in progestagen, occurred in all paired females. Three of the four singleton females that did not receive reLH treatment exhibited no rise in progestagen after an estrogen surge. All singleton females treated with reLH exhibited a rise in fecal progestagen after injection, indicating ovulation. In conclusion, deslorelin is effective at inducing ovarian activity and ovulation in paired female maned wolves; however, exogenous reLH is needed to induce ovulation in females housed alone. The findings obtained from this study serve as a foundation for future application of artificial insemination to enhance genetic management of this threatened species ex situ.

Estrus Cycle Status Defined by Vaginal Cytology Does Not Correspond to Fluctuations of Circulating Estrogens in Female Mice

Gender-oriented studies in shock, trauma, and/or sepsis require accurate monitoring of hormonal fluctuations as estrogens may influence various end points. Yet, monitoring is challenging in small laboratory animals: e.g., despite its subjectivity, vaginal smears are the major method for determination of estrus cycle phases in mice. Using female mice of different age, we aimed to (a) characterize general age-related changes in systemic estrogens and (b) examine the utility of determination of the estrus cycle by vaginal smears and/or impedance simultaneously comparing them with oscillation of systemic estrogens. In this study, 3-, 15-, and 20-month-old mice underwent vaginal smear and impedance examination each morning for 22 days. Ten hours after each morning checkup, feces were collected, and a second vaginal smear performed. Blood was collected on days 15 and 22. In 3-month-old females, estrus (by smears) was three times more frequent than in older mice, but mean concentrations of plasma and fecal estrogens never decreased with age. Collectively (not individually) plotted fecal estrogens values increased in the proestrus/estrus interphase (by smears) in 3-month-old mice only. Impedance typically peaked (4.5 Ω in 3-month-old mice) in the estrus phase, and only the prediction of estrus (highest area under the curve = 0.87 in 3-month-old) but not of other phases was possible. Regardless of age, individual cycle phase (by smears) never correlated with corresponding fecal estrogens, and estrus could not be predicted. In conclusion, while the fecal estrogens oscillation and frequency of estrus phase were affected by age, the systemic hormone release persisted. In mice, vaginal cytology did not reflect changes of systemic (fecal) estrogens, whereas impedance accurately identified estrus. The flaws and advantages of the examined monitoring methods should be considered in the design of future shock studies.

Presencia de doble estro anual en una hembra de lobo mexicano (Canis lupus baileyi) en cautiverio

Resumen As in most of the wolf subspecies, sexual behavior in Mexican wolf is shown in late winter once a year. Reproductive cycle in female wolves includes seasonality and monoestrum, with a long anestrus period. Estrogens and progesterone are responsible for estrus appearance and presence of both seems to be necessary for the full stimulation of sexual behavior in female wolves. In previous studies we made quantification of fecal estrogens and progesterone to several couples of Mexican wolves in captivity, in all the cases peaks on both hormones are correlated with reproductive behavior and shown in winter. Lately, another couple hosted in Aragon Zoo, Mexico City, shown, on in an unexpected way, sexual behavior during August in addition to the normal one in February; then we performed a sexual hormones analysis since this last periods in fecal samples. Results show a significant rise on female sexual steroid hormones concentrations in both, winter and summer. Female was checked for clinical and physical disorders, finding no evidence of pathologies. Pedigree was reviewed looking for possible hybridization, but only a pure wolf lineage was found, environmental factors were also studied providing no evidence that could explain such hormones concentrations. This is first report in summer of estrus behavior in Mexican wolf.

Factors influencing breeding success, ovarian cyclicity, and cub survival in zoo-managed tigers (Panthera tigris)

Understanding factors that influence reproduction and offspring survival in zoo populations is critical for management of threatened and endangered species. Examination of long-term data (1989–2011) compiled from the Association of Zoos and Aquarium's zoo-managed tiger breeding program provides the basis for a more thorough understanding of reproduction and scientifically based decisions for effective population management in this endangered felid. Biological and management-related factors that could influence tiger breeding success and cub survival were evaluated using logistic mixed models. Breeding success improved with female age until approximately age five, then declined thereafter. Experienced female breeders had greater breeding success than inexperienced females. Litter size was most predictive of cub survival, with average-sized litters (3–4 cubs) experiencing the highest proportional survival. Management-related factors, such as whether the breeding institution had a recent tiger litter and whether both animals were already located at the same institution, also influenced breeding success and cub survival. These results highlight the importance of institutional husbandry experience and the need to retain knowledge through staff turnovers to achieve optimal reproductive success. Using fecal estrogen data, frequency of ovarian cyclicity and mean cycle length did not differ by female age or parity; thus, lack of cyclicity and/or increased cycle duration are not likely explanations for declining breeding success with age. These results provide valuable reproductive information that should improve scientific management of zoo-based tiger populations.

Fecal estrogen, progestagen and glucocorticoid metabolites during the estrous cycle and pregnancy in the giant anteater (Myrmecophaga tridactyla): evidence for delayed implantation

BackgroundDeclining numbers of wild giant anteaters highlight the importance of sustainable captive populations. Unfortunately, captive reproductive management is limited by the lack of external physical indicators of female reproductive status and the aggressive behavior of males. We examined the endocrinology of the estrous cycle and pregnancy, and whether delayed implantation is a gestational strategy for giant anteaters as described for other xenarthrans.MethodsFeces were collected from seven captive females 3–5 times weekly and mating was recorded. Concentrations of estrogen (estrone–glucuronide, E1, and estradiol–17β, E2), progestagen (20–oxo–progestagens, P4), and glucocorticoid (GC) metabolites were examined in fecal extracts by enzyme immunoassay.ResultsEstrous cycles for nulliparous females (6 cycles, n = 2) compared to the multiparous female (6 cycles, n = 1) were shorter (47.3 +/− 4.3 days versus 62.5 +/− 2.6 days) with relatively lower luteal phase concentrations of P4 (49.4 +/− 2.9 ng/g versus 136.8 +/− 1.8 ng/g). The four remaining females had unclear ovarian activity: two females exhibited apparent luteal activity but unclear fluctuations in estrogens, while the other two females had parallel fecal P4 and estrogens concentrations. Pregnancy ranged 171–183 days with females returning to estrus post–partum as early as 60 days (n = 3, 1.8-4 years of age at mating). Delayed implantation was indicated by a biphasic elevation in fecal P4 metabolites: the initial 4–fold increase occurred for 81–105 days and was followed by a 26–fold secondary rise in P4 metabolites lasting 66–94 days prior to parturition. Fecal GC was correlated with fecal estrogens and greatest during estrus, late pregnancy, and six days prior to parturition (estrous cycle GC, 14.4-62.8 ng/g; pregnancy GC, 13.6-232.7 ng/g).ConclusionsEstrous cycles of giant anteaters occurred year–round, but were shorter and more intermittent in younger nulliparous animals compared to a multiparous female. A pronounced elevation in fecal P4, estrogen, and GC occurred during late gestation after an initial post-mating delay providing evidence for delayed implantation. Adrenocorticoid activity indicated impending parturition. Differences in estrous cycle characteristics with age and the protracted but variable gestation length must be considered to improve reproductive success and neonatal survival in giant anteaters.

Detection of estrus in Indian blackbuck: Behavioural, hormonal and urinary volatiles evaluation

The determination of the reproductive status is one of the most important factors for effective wild life conservation and management, and effective use of assisted reproductive techniques like artificial insemination, in vitro fertilization/embryo transfer depends on the knowledge of the basic reproductive physiology. In this context the reproductive status of female blackbucks (Antelope cervicapra L.) was assessed by behaviour and determination, sex steroid hormones in faeces and urinary volatile compounds. The male and female blackbucks exhibited as many as 31 different reproductive/courtship behaviour patterns. Particularly, the males showed a more extensive repertoire: i.e. 23 behavioural patterns by territorial males, 11 by bachelor males and 4 by females. The behaviours such as, mounting, Flehmen, clockwise and anticlockwise movements were significantly higher in male blackbuck when exposed to estrus. By contrast, such courtship behaviours were completely absent in male when exposed to diestrus. It clearly indicates that, the estrus female produces specific chemical cues (pheromone) through urine, which would involve in attracting the conspecifics. In addition, the average faecal oestrogen concentration was significantly higher (p<0.05) during the estrus faecal than the proestrus and diestrus periods. In contrast, the faecal progesterone concentration was significantly higher (p<0.05) during the diestrus faecal sample than that of proestrus and estrus faecal sample. Twenty-eight volatiles are identified, across the three reproductive phases (i.e. proestrus, estrus and diestrus) of sexually mature and prepubertal females. Amongst, the compounds 2-methyl-3-butyn-2-ol, 3,7-dimethylnonane, 3-phenyl-2-propen-1-ol and 2-hydroxybenzoic acid occurred only during estrus which may be considered as marker for detection of estrus which would ultimately help for artificial insemination in captive condition. The findings of the present study suggest that the non-invasive approaches like reproductive behaviours, faecal steroids and estrus-specific urinary volatiles could serve as good indicators for detection of estrus for blackbuck.

Changes in Fecal Estrogen Levels and Sexual Behavior in Captive Sichuan Snub‐Nosed Monkeys (Rhinopithecus roxellana) Following a Male Replacement

Rhinopithecus roxellana are generally seasonal breeders, although copulation can occur throughout the year. Previous studies suggest that estradiol modulates female sexual behavior during the mating season. However, the effects of social context on estrogen levels and behavior have not been fully explored. We studied the relationship between sexual behaviors and fecal estrogens in a group of captive R. roxellana during a period of social instability. We collected behavioral data for six months and collected fecal samples at 2-3-day intervals for four months spanning the mating and nonmating seasons, and analyzed fecal estrogen levels via RIA. Females showed clear cyclic solicitation and copulation peaks in the mating season, which corresponded with sharp peaks in fecal estrogens. During the nonmating season, solicitation rates, copulation rates, and fecal estrogens were generally low. However, one nonpregnant female displayed a sharp peak in solicitations, copulations, and estrogens during the nonmating season 10-14 days after a male replacement. Our results provide preliminary evidence that social and behavioral changes affect estrogen levels in R. roxellana.

Reproductive timing and aseasonality in the sun bear (Helarctos malayanus)

Most species of bears exhibit a reproductive strategy that includes a highly defined breeding season, delayed implantation, pseudopregnancy, and no postpartum estrus. Sun bears (Helarctos malayanus), a tropical lowland species, represent a potential departure from the typical ursid reproductive pattern. We studied reproductive timing in captive sun bears by examining fecal hormones and behavior of 13 female and 5 male sun bears, the entire North American population of H. m. euryspilus at the time of the study. Fecal estrogens and progestins in females did not vary by season or month, nor did fecal androgens in males. Estrus occurred in 11 of 12 months; breeding and births occurred year-round. However, birth rates were lower in the spring and higher in the fall and winter. Our study documented 1 complete pregnancy, in which delayed implantation did not occur; the delay between estrus and the progesterone rise (i.e., implantation) in the pregnant bear was similar to that seen in nonpregnant bears. All bears, whether nonpregnant or pregnant, exhibited a prolonged period of high progesterone after estrus, indicating obligate pseudopregnancy. Studbook records show that estrus can occur within 6 months after parturition; these cases were typically associated with loss of the previous cub. Our findings suggest that sun bears are unique among ursids, being polyestrous, nonseasonal breeders that do not appear to exhibit delayed implantation and that are capable of initiating estrus in the event of early loss of a cub.

Sex differences in the mother–neonate relationship in wild baboons: social, experiential and hormonal correlates

In mammals, maternal care is essential for offspring survival, yet individual differences in this care can dramatically affect offspring growth and development. Few studies have, however, investigated the sources, magnitude and consequences of naturally occurring interindividual variation in maternal care during the neonatal period. In this study, we examine several hormonal and nonhormonal predictors of naturally occurring variation in the mothereneonate relationship during the first 8 weeks of infancy in 34 wild baboon (Papio cynocephalus) mothereinfant dyads in Amboseli, Kenya. We use data on physical contact and suckling patterns to assess the quality of the mothereneonate relationship and to evaluate the extent to which variation in this relationship is predictable from perinatal ovarian steroids (i.e. faecal oestrogen and progesterone metabolites), previous infant care experience, maternal dominance rank and offspring sex. We found that newborn infants of more experienced mothers initiated higher rates of changes in mothereinfant contact than newborns of less experienced mothers. However, at each level of maternal experience, newborn males initiated higher rates of changes in mothereinfant contact than newborn females. Moreover, we found evidence suggesting that variation in suckling activity among daughters (but not sons) was predictable from maternal dominance rank and faecal oestrogen (fE) concentrations before birth. To our knowledge, our study provides the first evidence of (1) the influence of cumulative maternal experience on the mothereinfant relationship and (2) the emergence of sex differences in the mothereinfant relationship during the neonatal period in wild primates. Our results suggest that the well-documented sex differences in life history, behaviour and ecology in primates (and other social mammals) may originate very early in life. 2012 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved. For vertebrates, mammalian newborns are unusually dependent on their mothers for nourishment, transport, protection and support (Clutton-Brock 1991). Although offspring eventually transition from complete reliance on their mothers to nutritional, locomotor and social independence, the timing of these events can vary widely across individuals of the same species (Clutton-Brock 1991). Despite the critical importance of the neonatal period in the life of each individual, few studies have investigated the sources, magnitude and consequences of interindividual variation in the mothereoffspring relationship during the neonatal period in wild mammals (Altmann 1980; Bales et al. 2002). In this study, we

Hormonal Correlates of Paternal Care Differences in the Hylobatidae

Only one of the 15 species of monogamous hylobatids, the siamang (Symphalangus syndactylus), demonstrates direct paternal care in the form of infant-carrying, providing a unique model for examining hormonal correlates of paternal care differences between siamangs and gibbons. We used behavioral data and fecal hormone analysis to investigate (1) differences in monthly percent father-infant proximity in relation to monthly fecal androgen metabolite concentrations from infant birth to the late postpartum period between siamangs and gibbons, (2) the pattern of change in fecal androgen and fecal estrogen metabolite concentrations during the 8-week peripartum period between siamangs and gibbons, and (3) the change in mean fecal glucocorticoid metabolite concentrations at 1-month postpartum from individual baseline between siamangs and gibbons. Father-infant proximity increased as androgen concentrations decreased over the postpartum period in siamangs but not in gibbons. Androgen concentrations increased around birth in siamangs during the 8-week peripartum period, but exhibited a decreasing trend around birth in gibbons. Estrogen concentrations increased from pre- to postpartum in siamangs during the 8-week peripartum period, but exhibited a decreasing trend from pre- to postpartum in gibbons. The difference in mean glucocorticoid metabolite concentrations from baseline was greater in siamangs than gibbons. Our data suggest a relationship between specific steroid hormone patterns and differences in paternal care among the hylobatids, warranting further investigation of such proximate mechanisms.

Reproductive gonadal steroidogenic activity in the fishing cat ( Prionailurus viverrinus) assessed by fecal steroid analyses

Non-invasive fecal steroid analyses were used to characterize gonadal activity in the fishing cat ( Prionailurus viverrinus). Estrogen, progestagen and androgen metabolites were quantified in fecal samples collected for 12 months from four males and 10 females housed at seven North American zoological institutions. Male reproductive hormone concentrations did not vary ( P > 0.05) among season, and estrogen cycles were observed year-round in females and averaged (±SEM) 19.9 ± 1.0 days. Mean peak estrogen concentration during estrus (460.0 ± 72.6 ng/g feces) was five-fold higher than baseline (87.3 ± 14.0 ng/g feces). Five of seven females (71.4%) housed alone or with another female demonstrated spontaneous luteal activity (apparent ovulation without copulation), with mean progestagen concentration (20.3 ± 4.7 μg/g feces), increasing nearly five-fold above baseline (4.1 ± 0.8 μg/g feces). The non-pregnant luteal phase averaged 32.9 ± 2.5 days ( n = 13). One female delivered kittens 70 days after natural mating with fecal progestagen concentrations averaging 51.2 ± 5.2 μg/g feces. Two additional females were administered exogenous gonadotropins (150 IU eCG; 100 IU hCG), which caused hyper-elevated concentrations of fecal estrogen and progestagen (plus ovulation). Results indicate that: (1) male and female fishing cats managed in North American zoos are reproductively active year round; (2) 71.4% of females experienced spontaneous ovulation; and (3) females are responsive to exogenous gonadotropins for ovulation induction, but a regimen that produces a normative ovarian steroidogenic response needs to be identified.

Rising fecal glucocorticoid concentrations track reproductive activity in the female giant panda ( Ailuropoda melanoleuca)

To better understand the adaptive significance of adrenal glucocorticoid (GC) variation in the giant panda, we assessed patterns of fecal GC excretion over time as well as during estrus, parturient and non-parturient luteal phases, lactation and acyclicity in 17 adult females. Fecal estrogen and GC patterns were positively correlated ( P < 0.05) in four of five periestrual females ( r = 0.57–0.92). Among all reproductive states, fecal GC was highest ( P < 0.05) during periestrus (non-parturient, 495.9 ± 100.7 ng/g [mean ± SE]; parturient, 654.1 ± 106.5 ng/g; P > 0.05). Concentrations of GC metabolites were lower ( P < 0.05) during the later stage of the luteal phase in non-parturient (334.8 ± 24.8 ng/g) compared to parturient (470.4 ± 54.0 ng/g) females. Although fecal GC concentrations in cyclic, non-parturient females did not differ ( P > 0.05) across all seasons, there were seasonal variations ( P < 0.05) in females that were acyclic and non-lactational. However, the overall lack of difference ( P > 0.05) in GC values between reproductively cyclic and acyclic females did not support the hypothesis that ovarian acyclicity is due to increased adrenal activity (related or unrelated to physiological stress). Furthermore, GCs may play an important role in the normal endocrine milieu associated with sexual receptivity and late pregnancy. These data demonstrate that both reproductive status and seasonal factors are important modulators of adrenal function in this endangered species.

Pattern of faecal 20-oxopregnane and oestrogen concentrations during pregnancy in wild plains zebra mares

Regulative endocrine mechanisms influence the reproductive behaviour and success of mammals, but they have been studied predominantly in domestic and captive animals. The study aims at describing the pattern of faecal 20-oxopregnane and oestrogen concentrations during pregnancy in wild plains zebra Equus quagga chapmani. Data were collected during wet and dry seasons 2007–2009. Enzyme Immunoassays were used to determine 20-oxopregnane and oestrogen concentrations in faecal samples (n=74) collected from individual mares (n=32) whose dates of foaling were known through long-term monitoring. Hormonal profiles were described with a General Additive Model (GAM: Hormone ∼ Days to Foaling). Faecal 20-oxopregnanes have a complex cycle during pregnancy (GAM, n=70, R2=0.616, p<0.001). From −250days to foaling, faecal 20-oxopregnane concentrations were above the baseline levels found in non-pregnant mares, peaking in the last 50days. Faecal oestrogen levels showed a clear peak in mid-pregnancy (GAM, n=62, R2=0.539, p<0.001). The sex of the foetus and season had no detectable effect on hormone concentrations during pregnancy. High levels (>200ng/g DW) of faecal 20-oxopregnanes associated with high (>160ng/g DW) faecal oestrogen levels indicate mid-pregnancy in c.90% of cases (16/17). High faecal 20-oxopregnanes (>200ng/g DW) and low faecal oestrogen levels (<160ng/g DW) indicate late pregnancy, again in c.90% of cases. Two faecal samples would allow the stage of pregnancy to be determined with confidence.

The reproductive endocrinology and behavior of Vancouver Island marmot (Marmota vancouverensis)

The Vancouver Island marmot (Marmota vancouverensis; VIM) is one of North America's most endangered species with fewer than 150 individuals remaining in the wild. A captive breeding program was established across four facilities in Canada as an insurance population and source of animals for reintroduction to the wild. The purpose of this study was to gather information about the basic reproductive biology and behavior of this species, which is essential to improve captive breeding programs. Regular fecal samples were obtained from adult female (n = 14) and male (n = 10) marmots, 2 years of age and older, over 1-3 breeding seasons (2-3 months duration posthibernation) for steroid hormone analysis. Enzyme immunoassays were validated for quantifying fecal testosterone metabolite concentrations for males, and fecal estrogen and progesterone metabolite concentrations for females. Results indicated that fecal progesterone metabolite concentrations can be used to monitor ovulation and pregnancy. Behavioral monitoring through infrared video surveillance was conducted in four breeding pairs over a 2-year period (n = 7 behavioral profiles). Breeding behaviors correlated strongly with changes in reproductive endocrine profiles. A high frequency of play behavior or "wrestling" was observed in conjunction with breeding activity before an elevation in progesterone metabolite concentrations. Impending parturition was associated with increased aggression and exclusion of the male from the maternal nestbox as well as an increase in nesting activity. Observational data combined with hormonal analysis suggest that female VIMs are induced ovulators and that multiple breeding attempts may be required for ovulation and conception. Gestation appears to be approximately 34 days from peak breeding activity (32 days from estimated ovulation). Fecal testosterone concentrations suggest that testicular activity is seasonal with the reproductive activity occurring immediately posthibernation. Monitoring breeding behavior is a useful means of indicating estrus, conception and pregnancy, which can also be supported by the hormonal analysis of daily fecal samples of individual animals.

Daily fecal sex steroid hormonal changes and mating success in captive female cheetahs ( Acinonyx jubatus) in Japan

Daily fecal estrogen and progestin concentrations were measured by enzyme immunoassay in five female cheetahs ( Acinonyx jubatus) for 4–6 months. The animals were housed under different conditions: (1) a female always housed in a group including one or more males; (2) two females isolated individually for short or long periods; (3) the other two females housed together. These females were separately housed with males for mating around the time of the estrogen peaks. The hormone profiles were similar in all five females regardless of the housing conditions. However, only the female that had been isolated from other cheetahs for over a year mated and reproduce cubs successfully, whereas the remaining four did not (one was isolated for only 6 weeks, another was always housed with males and the other two were housed together). In all females, the estrogen peaks were obtained at regular intervals of approximately 8–15 days. Unlike estrogen, the progestin concentrations were always low in all females except during pregnancy and they did not increase following the estrogen surges. These results showed that female cheetahs are typically reflex ovulators and female receptiveness may not be reflected to her hormonal states. It was also suspected that individual housing and long-term separation are advantageous for breeding this wild cat in captivity, mimicking the ecological/behavioral patterns in the wild, though housing condition might have no effect on the estrous cycle.