Fecal analysis of ovarian cycles in female black-handed spider monkeys (Ateles geoffroyi)

An enzyme immunoassay (EIA) was applied to characterize the reproductive endocrinology of adult female black-handed spider monkeys (Ateles geoffroyi). Analysis of paired urine and fecal samples, collected from two females housed at San Diego Zoo, confirmed that the EIAs employed provided quantitative measurements of ovarian sex steroid hormones. Fecal metabolite levels were significantly correlated with those in urine, confirming that feces are a valid source of steroid metabolites in this species. The excretion of these metabolites in feces lagged urinary excretion by 1-2 days. The ovarian cycle profiles of the two captive females and five free-ranging females are comparable, with an average length of approximately 20-23 days. Cyclical bleeding, as previously reported, was observed in one of the two captive females. Pregnancy was detected in four free-ranging females, and early fetal loss for one female was indicated by hormonal data.     

Steroid excretion during the ovarian cycle in captive and wild muriquis,Brachyteles arachnoides

Urine, feces, and copulation frequency were collected from two captive muriqui females, Brachyteles arachnoides, at the Centro de Primatologia do Rio de Janeiro following the resumption of postpartum ovarian cycles. Fecal steroid profiles from seven wild muriqui females at the Estação Biologica de Caratinga, Minas Gerais, Brazil, were compared to the captive females to determine the approximate patterns of steroid excretion relative to the urinary LH peak. Hormonal profiles from one of the captive female muriquis revealed a discrete urinary LH peak. For this female, fecal progesterone increased on the same day as the urinary LH peak, while fecal estradiol increased 6 days later and urinary steroids increased 5 days later. For both captive females, the onset of fecal progesterone increase was preceded by the onset of copulations, which occurred during at least a 5-day period. The complete fecal hormonal profiles of the one captive female for which continuous data were available were similar to those found in wild muriqui monkeys, with the onset of an increase in sustained progesterone levels occurring several days prior to the onset of sustained estradiol increase. These patterns suggest that fecal progesterone may be excreted rapidly in this species. The onset of sustained increase in fecal progesterone levels, together with the consistent delay in the onset of the sustained increase in estradiol, may provide the best indicators of the periovulatory period for muriqui females. Am. J. Primatol. 42:311–321, 1997. © 1997 Wiley-Liss, Inc.     

Diurnal variation on the excretion patterns of fecal steroids in common marmoset (Callithrix jacchus) females

The use of fecal steroid analysis to assess gonadal and adrenal function in primates has rapidly increased in recent years due to the ability to collect feces from nonhuman primates living in wild conditions. These techniques offer an exciting new potential for enhancing our knowledge of the endocrine status of free-living animals. Prior to using these techniques under field conditions, it is important to determine the diurnal variation of fecal excreted steroids for assessing possible time limitations on fecal collections. The following study investigates the diurnal frequency of defecation and patterns of steroid levels excreted in feces from four female common marmosets, Callithrix jacchus, living in a family group. These females represented three reproductive conditions: early pregnancy, ovarian cycling, and noncycling (postpubertal). Cortisol, estradiol, and progesterone were extracted and analyzed by enzyme immunoassay. Diurnal variations in steroid levels were found by ANOVA for cortisol and progesterone but not for estradiol. Significantly higher levels of cortisol were found in the afternoon, while the reverse was found for progesterone. All females showed the same pattern of steroid level change, except for cortisol in the pregnant female. Since all females defecated within the first hour after they awoke in the morning, this time was determined to be the most effective time to collect feces. The consistency of our findings reinforces the usefulness of this approach for studying reproductive and adrenocortical function in marmosets and also indicates that fecal collection should be limited to either morning or afternoon collections. Am. J. Primatol. 46:105–117, 1998. © 1998 Wiley-Liss, Inc.     

Measurement of urinary and fecal steroid metabolites during the ovarian cycle in captive and wild Japanese macaques, Macaca fuscata

We measured the concentration of steroid hormones from urine, feces, and blood samples of two captive Japanese macaques, Macaca fuscata, during nonconceptive ovarian cycles to compare the patterns of the excreted steroids with those of circulating steroids. Urine and feces were analyzed for estrone conjugates (E1C) and pregnanediol-3-glucronide (PdG) using enzyme immunoassays (EIAs), while plasma was analyzed for estradiol-17beta(E2), progesterone (P), and luteinizing hormone (LH) using radioimmunoassays (RIAs). Urinary and fecal E1C and PdG levels were approximately parallel to plasma E2 and P levels, respectively. The E1C profiles of daily urinary and fecal samples revealed a midcycle peak, followed by a sustained PdG increase lasting up to two weeks from the E1C peak. A fecal E1C peak was one day later than the urinary E1C peak. One of the captive females exhibited a discrete plasma LH peak, one indicator that ovulation has occurred, on the day following the urinary E1C peak, i.e., the same day of fecal E1C peak. We measured excreted steroids in nine wild females and determined the timing of ovulation by comparing fecal steroid profiles to those obtained in captive monkeys. Data from wild females indicated that eight of nine females conceived during their first ovulatory cycle of the sampling period, whereas the remaining female failed to conceive during the sampling period even though she ovulated. In the eight females that conceived, E1C increased again following the detected or estimated E1C peak, with levels comparable to the preovulatory peak levels, and sustained elevations of PdG for over 40 days. These data illustrate that the urinary and fecal profiles of ovarian steroid excretion obtained through the application of these noninvasive techniques provide an accurate approach for monitoring conceptive and nonconceptive ovarian cycle in captive and free-living Japanese macaques.     

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±72.6ng/g feces) was five-fold higher than baseline (87.3±14.0ng/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±4.7μg/g feces), increasing nearly five-fold above baseline (4.1±0.8μg/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±5.2μg/g feces. Two additional females were administered exogenous gonadotropins (150IU eCG; 100IU 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.     

Comparative analysis of gonadal and adrenal activity in the black and white rhinoceros in North America by noninvasive endocrine monitoring

Patterns of fecal reproductive steroid metabolites and adrenal corticoids were characterized for 12‐ to 24‐month periods in black (n = 10 male, 16 female) and white (n = 6 male, 13 female) rhinoceroses at 14 institutions. All black rhinoceros females exhibited at least some ovarian cyclicity on the basis of fecal progestogen analysis (range, 2–12 cycles/yr). However, cycles often were erratic, with many being shorter (<20 days; 18% of cycles) or longer (>32 days; 21%) than the average of 26.8 ± 0.5 days (n = 104 cycles). Five females exhibited periods of acyclicity of 2–10‐month duration that were unrelated to season. One complete and seven partial pregnancies were evaluated in the black rhinoceros. Fecal progestogens increased over luteal phase concentrations after 3 months of gestation. Females resumed cyclicity within 3 months postpartum, before calves were weaned (n = 5). Approximately half of white rhinoceros females (6 of 13) showed no evidence of ovarian cyclicity. Of the cycles observed, 5 were “short” (32.8 ± 1.2 days) and 24 were “long” (70.1 ± 1.6 days). Only two females cycled continuously throughout the study. One had both long (n = 9) and short (n = 2) cycles, whereas the other exhibited long cycles only (n = 5). Fecal estrogen excretion was variable, and profiles were not useful for characterizing follicular activity or diagnosing pregnancy in either species. Males of both species showed no evidence of seasonality on the basis of fecal androgen profiles. Androgen metabolite concentrations were higher (P < 0.05) in the black (27.6 ± 6.9 ng/g) than in the white (16.8 ± 3.1 ng/g) rhinoceros. An adrenocorticotropin hormone challenge in four black rhinoceros males demonstrated that the clearance rate of corticoid metabolites into feces was ～24 hours. Fecal corticoid concentrations did not differ between males and females, but overall means were higher in the black (41.8 ± 3.1 ng/g) than in the white (31.2 ± 1.7 ng/g) rhinoceros. In summary, fecal steroid analysis identified a number of differences in hormonal secretory dynamics between the black and white rhinoceros that may be related to differences in reproductive rates in captivity. Most black rhinoceros females exhibited some cyclic ovarian activity. In contrast, few white rhinoceroses demonstrated evidence of regular estrous cyclicity, and those females that were active had comparatively long cycles. Results also suggest that fecal corticoid concentrations reflect adrenal activity and may be species specific. Continued studies are needed to determine whether fecal corticoid measurements will be useful for understanding the cause of inconsistent gonadal activity in these two species. Because all but three (15.8%) of the white rhinoceroses evaluated in this study were less than 20 years of age compared to 73.1% (19 of 26) of the black rhinoceroses, the impact of age on reproductive and adrenal activity also needs to be evaluated further. Zoo Biol 20:463–486, 2001. © 2002 Wiley‐Liss, Inc.     

Characterization of reproductive cycles and adrenal activity in the black‐footed ferret (Mustela nigripes) by fecal hormone analysis

The reproductive cycle of the black‐footed ferret (Mustela nigripes) was characterized by enzyme immunoassay (EIA) analysis of ovarian fecal steroids (estradiol, progestins) in 29 females over two consecutive breeding seasons. Estrous status was determined by measuring the vulva size and examining the percentage of superficial cells in vaginal lavages. Mean fecal estradiol concentrations were correlated with vulval area (r = 0.370, P < 0.0001) and the percentage of superficial cells (r = 0.380, P < 0.0001). Ovulation resulted in a rise in fecal progestin concentrations 5 days after breeding that differed (P < 0.05) between pregnant (n = 14) and pseudopregnant (n = 12) females during the late luteal phase (days 12–40), with concentrations remaining higher in pregnant animals. Gestation length was 41.3 ± 0.7 days with 3.6 ± 0.4 kits produced per female. Litter size correlated significantly (P < 0.05) with fecal estradiol, but not progestins during the 12 to 40 days after breeding. Females failing to breed (n = 3) remained in estrus for 31 ± 6.2 days before ovulation induction with human chorionic gonadotropin. Adrenal activity in male (n = 4) and female (n = 6) black‐footed ferrets was monitored by quantifying fecal corticoid metabolites after a series of manipulations (physical restraint, intramuscular saline, intramuscular gel adrenocorticotropic hormone (ACTH), intramuscular liquid ACTH). A significant (P < 0.0001) increase in fecal corticoids above the pre‐treatment baseline occurred 20 to 44 hours after restraint (five of 10 animals), saline (six of nine), gel ACTH (seven of 10), and liquid ACTH (nine of 10) treatments. Immunoreactivity of high‐performance liquid chromatography–separated fecal elutes was compared using antibodies against cortisol and corticosterone. The cortisol EIA demonstrated immunoreactivity that co‐eluted with ³H‐cortisol, whereas a corticosterone radioimmunoassay detected a metabolite peak that co‐eluted with ³H‐corticosterone in addition to a slightly less polar and one considerably more polar peak. Despite recognizing different metabolites, both assays produced similar temporal profiles of corticoid excretion after manipulation. This study provides new information on the black‐footed ferret regarding differences in fecal steroid excretion patterns between pregnancy and pseudopregnancy and the potential application of fecal corticoid metabolite monitoring for evaluating responses to stressors associated with practices used in breeding management. Zoo Biol 20:517–536, 2001. © 2002 Wiley‐Liss, Inc.     

Perineal swellings: a social and endocrine advantage for Barbary macaque females (Macaca sylvanus)

This study focuses the relationship between social rank, intersexual behavior, age, fecal cortisol equivalent excretion rates and perineal swelling size in contraceptive-treated Macaca sylvanus females. Behavioral data and fecal samples were collected during a twenty-week period from 24 females. Females were categorized with regard to perineal swelling (enlarged or reduced), and to their dominance rank (high or low); we found that swelling size was not associated with it. However, females with an enlarged perineum received more male grooming interactions, had closer spatial relations to males, and received significantly more interventions from males than did the others. Endocrinologically, females with enlarged swelling showed decreased fecal cortisol equivalent excretion rates. Multiple regression models showed a positive relationship between the extent of the swelling size and grooming, inspection and spatial relation. There was a negative relationship between the extent of swelling size and fecal cortisol equivalents. These results imply that enlarged perineal swellings among implanted Barbary macaque females have a greater impact on intersexual contact and adrenocortical activity than does social rank or age.     

Excretion and measurement of estradiol and progesterone metabolites in the feces and urine of female squirrel monkeys (Saimiri sciureus)

The first objective of the present study was to determine the metabolic form and rate of excretion of ovarian hormone metabolites in the urine and feces of female squirrel monkeys injected with radiolabeled progesterone (Po) and estradiol. The major portion of the urinary metabolites of both hormones was excreted within 16-24 hr post-injection. Estrogen and Po isotopes in feces exhibited an excretion peak at 16 hr post-injection. The majority of recovered radiolabel of both hormones was excreted in feces. Chromatographic separation of fecal extractions indicated that the major estrogen metabolites in feces are in the free as opposed to the conjugated form. The radioactivity and immunoreactivity for estrone and estradiol (E(1) and E(2), respectively) in eluates of fecal samples subjected to celite co-chromatography indicated that both free E(1) and E(2) exist as excretion products in the feces of female squirrel monkeys. The major radioactive peaks for Po metabolites showed peaks in the elution profile at or very near the Po standard, and corresponded with the celite co-chromatography elution profile of Po standard when subjected to enzyme immunoassay (EIA). The second objective was to validate the application of EIA systems to measure fecal metabolites. Reproductive events of one female squirrel monkey across one annual reproductive cycle are described using the endocrine profile generated from fecal steroid assays. Examination of this profile confirmed that longitudinal fecal sampling and steroid hormone metabolite measurement in feces was not only feasible and practical, but accurately detected known reproductive events as well.     

Steroid metabolism and validation of noninvasive endocrine monitoring in the African wild dog (Lycaon pictus)

The purpose of this study was to validate noninvasive endocrine monitoring techniques for African wild dogs (Lycaon pictus) and to establish physiological validity of these methods by evaluating longitudinal reproductive-endocrine profiles in captive individuals. To determine the primary excretory by-products of ovarian steroid metabolism, [¹⁴C]-progesterone and [³H]-estradiol were co-administered to a female and all excreta were collected for 80 hr postinjection. Radiolabel excretion peaked ≤ 18 hr postinfusion, and progesterone and estradiol metabolites were excreted in almost equivalent proportions in urine (39.7 and 41.1%, respectively) and feces (60.3 and 58.9%, respectively). Most of the urinary metabolites were conjugated (estradiol, 94.3 ± 0.3%; progesterone, 90.4 ± 0.5%), so that immunoassays for pregnanediol-3α-glucuronide (PdG) and estrogen conjugates (EC) were effective for assessing steroid metabolites. Two immunoreactive estrogens (estradiol and estrone) and at least one immunoreactive progesterone metabolite (3α-hydroxy-5α, pregnan-20-one) were detected in feces. Urine and fecal samples were collected (1–3 times per week) for 1.5 yr from one adult female and two adult males to assess longitudinal steroid metabolite excretion. Overall correlation of urinary PdG to matched, same-day fecal progesterone metabolites immunoreactivity was 0.38 (n = 71, P < 0.05). Similarly, urinary EC was correlated (P < 0.05) with same-day fecal estrogen immunoreactivity (r = 0.49, n = 71). During pregnancy and nonpregnant cycles, copulation occurred at the time of peak (or declining) estrogen metabolites and increasing progesterone metabolites concentrations. Estrus duration was 6–9 days and gestation lasted 69 days with parturition occurring coincident with a drop in progesterone metabolites. Males exhibited seasonal trends in fecal testosterone excretion with maximal concentrations from July to September coincident with peak mating activity. Although these limited longitudinal hormone profiles should be interpreted cautiously, noninvasive gonadal steroid monitoring suggests that: (1) both female and male wild dogs may exhibit reproductive seasonality in North America, (2) females are monoestrous, and (3) peak testicular activity occurs between August and October coincident with mating behavior. From a conservation perspective, noninvasive endocrine monitoring techniques should be useful for augmenting captive breeding programs, as well as for developing an improved understanding of the physiological mechanisms underlying reproductive suppression in response to social and ecological pressures. Zoo Biol 16:533–548, 1997. © 1997 Wiley-Liss, Inc.     

Social and reproductive conditions modulate urinary cortisol excretion in black tufted-ear marmosets (Callithrix kuhli)

The links between psychosocial stress, social status, reproductive function, and urinary cortisol were assessed in social groups of black tufted-ear marmosets (Callithrix kuhli). Urinary cortisol excretion was monitored in cases of intrafamily conflict (“sibling fights”) and in daughters in four distinct social contexts: in the family group, while housed singly or in same-sex pairs, and while paired with a male pairmate. Cortisol excretion was elevated in participants in intra-family conflict on the day of and the day following the conflict, relative to concentrations a week prior to or following the conflict. Daughters in natal family groups had concentrations of cortisol that did not differ from reproductively active adult females. This finding held for daughters who were either anovulatory or undergoing ovulatory cycles while in the natal family group. Natal family members and male pairmates exerted buffering effects on levels of activity in the hypothalamic pituitary adrenal axis (HPA) in female C. kuhli. Placing females in solitary housing led to significantly increased cortisol excretion. In the 2 months subsequent to pairing with a male partner, excreted cortisol concentrations in females declined significantly. Daughters removed from their natal family group and housed with a sister did not exhibit increased cortisol levels. These data reveal that activity in the (HPA) axis in marmosets is sensitive to psychosocial stressors, and that urinary cortisol can provide a useful quantitative measure of HPA reactivity. As in other callitrichids, delayed breeding in daughters and reproductive anomalies in C. kuhli appear to be mediated by mechanisms other than elevated HPA activity. Am. J. Primatol. 42:253–267, 1997. © 1997 Wiley-Liss, Inc.     

Non-invasive detection and monitoring of estrus,pregnancy and the postpartum period in pygmy loris (Nycticebus pygmaeus) using fecal estrogen metabolites

Estrone-conjugates (E₁C) were measured in the feces of six female pygmy lorises (Nycticebus pygmaeus) during estrus (n = 12), pregnancy (n = 4) and the postpartum period (n = 3). Noninvasive feces collection permitted frequent sampling throughout estrus and pregnancy, without disturbance of animals. The estrous period was defined as an increase in fecal E₁C levels above an average of 70 ng/g feces with peaks above 100 ng/g feces obtained in consecutive fecal samples collected over a 6- to 11-day period between the end of July and the first third of October. Comparison of the periovulatory profile of E₁C and the stage of labial opening of the vagina revealed a high agreement (P < 0.001). In all pregnant females, an E₁C rise was found approximately 47 days postestrus, the source of which may be the growing fetal placental unit. Estimated gestation lengths ranged between 187 and 198 days (n = 4). Am. J. Primatol. 41:103–115, 1997. © 1997 Wiley-Liss, Inc.     

Behavioral and endocrine characteristics of the reproductive cycle in wild muriqui monkeys,Brachyteles arachnoides

The analysis of fecal ovarian steroids provides a powerful noninvasive method to obtain insights into ovulatory cycles, gestation length, and the timing of sexual interactions relative to the periovulatory period in wild primates. Techniques developed to collect and assay feces from free-ranging muriqui monkeys (Brachyteles arachnoides) for estradiol and progesterone yield the first explicit reproductive data on this species, and provide the first opportunity to evaluate the timing of observed copulations with muriqui ovarian cycles. Hormonal profiles from seven females indicate average cycle lengths of 21.0 ± 5.4 days (n=20). Females conceived after 3–6 ovulatory cycles. Gestation length averaged 216.4 ± 1.5 days for the five females for which conception cycles were sampled. Discrete copulation periods spanned an average of 2.1 ± 1.2 days (n=29), with intervals between these concentrated periods of copulations averaging 15.6 ± 6.7 days (n=20). There were no significant differences among females in cycle lengths, copulation period lengths, or copulation interval lengths. Ejaculation was visible following 71.8 ± 26.7% of copulations during the females' preovulatory periods. All females copulated outside the periovulatory period. The proportion of copulation days outside the periovulatory period was slightly greater (p=0.08) for primiparous females (64.8 ± 28.3%) than for multiparous females (28.7 ± 19.7%). Am. J. Primatol. 42:299–310, 1997. © 1997 Wiley-Liss, Inc.     

Relationship between ovarian cycle phase and sexual behavior in female Japanese macaques (Macaca fuscata)

We conducted behavioral observations simultaneously with fecal sample collection on eight nonlactating females 2-3 times per week, October 1997-March 1998, to examine the relationship between ovarian hormones and the sexual behavior of female Japanese macaques (Macaca fuscata) during the mating season. We analyzed samples by enzyme immunoassay for fecal hormone levels. Hormone profiles of estrone-glucuronide (E1) and pregnanediol-glucuronide (PdG) were used to separate ovarian cycles into three phases (follicular, periovulatory, and luteal). Hormonal profiles indicate average cycle lengths of 27.6 +/- 4.2 days (+/- SD; n = 26). Average lengths of the luteal and follicular phases were 12.3 +/- 3.8 days (+/- SD) and 8.3 +/- 3.4 days (+/- SD), respectively. We observed female Japanese macaques engaging in sexual activity throughout the ovarian cycle, with the highest rates occurring during the follicular and periovulatory phases as compared to the luteal phase. The attractivity of female Japanese macaques increased significantly during the follicular and periovulatory phases of the ovarian cycle, when E1 levels are peaking and PdG levels drop to baseline. In addition, females displayed a significant increase in proceptive behavior during the follicular and periovulatory phases. Grooming bouts, as well as proximity between female and male macaques, also increased significantly during the follicular and periovulatory phases. We conclude that fluctuating levels of ovarian hormones in different phases of the cycle are significantly associated with variable rates of copulatory and pericopulatory behaviors in these Japanese macaque females.     

Ovarian Cycle and Effect of Social Changes on Adrenal and Ovarian Function in Pygathrix nemaeus

To assist captive breeding of the endangered red-shanked douc langur (Pygathrix nemaeus), basic knowledge on female reproductive physiology is important. We aimed 1) to characterize the pattern of fecal estrogens and progestogens during the ovarian cycle and 2) to use the information to provide reliable data on ovarian cycle characteristics. Moreover, we examined the potential impact of a change in group composition on ovarian activity and adrenal function, the latter being measured by fecal glucocorticoid excretion. We collected fecal samples during 2 mo from 5 adult females of 3 groups under undisturbed conditions and from the same 5 adult females and 2 juvenile females and 2 adult males during 3 mo during which there were changes in housing and group composition. During the undisturbed period, measurement of fecal 5-P-3OH, a major progesterone metabolite, and total estrogens revealed in all adult females a cyclic pattern, from which we documented a cycle length of 26.4 ± 1.8 days, comprising a follicular phase of 13.1 ± 2.9 days and a luteal phase of 14.8 ± 1.2 days. In contrast, during the period of social changes, 3 of the 5 females showed clear signs of cycle irregularity as indicated by extended follicular phases, abbreviated luteal phases and/or anovulation. Moreover, the period of social changes was associated with an overall significant elevation in fecal glucocorticoid excretion. Our data provide the first detailed information on the endocrine characterization of the ovarian cycle in Pygathrix nemaeus and suggest that social changes have the potential to impair ovarian function, likely as a result of increased activation of the HPA-axis due to stressful situations. However, because of relatively small sample size, particularly concerning the latter finding, more data are needed to confirm these results.     

Monitoring ovarian function in scimitar-horned oryx (Oryx dammah) by measurement of fecal 20α-progestagen metabolites

The feasibility of monitoring ovarian function in scimitar-horned oryx (Oryx dammah) by measurement of fecal 20α-progestagens was investigated. Fecal samples were collected daily or on alternate days over a 4–11 month period from five oryx during natural (n = 4) or synthetic PGF_2α (cloprostenol)-controlled (n = 1) cycles. Of the four oryx undergoing natural cycles, three had regular access to a vasectomised male, and mating dates were recorded. Ultrasonography was used to monitor changes in reproductive tract morphology in the female administered with cloprostenol. Neutral steroids were extracted from feces with methanol:petroleum ether (2:1 v/v) after first removing phenolic steroids with potassium hydroxide (1 M). The concentration of 20α-progestagens in the methanol phase was measured by enzymeimmunoassay. Excretion of 20α-progestagens in all females followed a cyclic pattern corresponding to the follicular and luteal phases of the ovarian cycle. Concentrations of fecal 20α-progestagens were on average twentyfold greater during the luteal phase compared with the follicular phase. Mean (±SD) ovarian cycle length, based on fecal progestagen profiles, was 24.4 ± 2.2 days with mean (±SD) luteal and follicular phase lengths of 18.7 ± 2.8 and 5.7 ± 1.6 days, respectively. Mating by a vasectomized male occurred when 20α-progestagen concentrations were still elevated or declining. Similarly, fecal progestagens did not return to follicular phase concentrations for 4–5 days after administration of cloprostenol, and a 4 day delay was observed between ovulation, as visualized by ultrasound scanning, and a rise in fecal 20α-progestagens. These data suggest a time lag of approximately 4 days between reproductive events and changes in fecal 20α-progestagen concentrations. We conclude that measurement of immunoreactive 20α-progestagen concentrations in feces has limited application for predicting ovulation or accurately timing inseminations because of delay in steroid excretion, but will enable noninvasive monitoring of ovarian cycles in scimitar-horned oryx for fertility assessment and for determining the outcome of artificial insemination programs. © 1995 Wiley-Liss, Inc.     

Seasonal dynamics of Skrjabinoptera phrynosoma (Nematoda) infection in horned lizards from the Alvord Basin: temporal components of a unique life cycle

The nematode Skrjabinoptera phrynosoma is a stomach parasite of horned lizards in the genus Phrynosoma. This nematode demonstrates a distinctive life cycle wherein entire gravid females harboring infective eggs exit with lizard feces. Pogonomyrmex spp. harvester ants collect these females and feed them to their larvae, which are the only stages of the intermediate host that can become infected. We hypothesized that the seasonal dynamics of nematode abundance within lizard hosts would be correlated with the seasonal availability of suitable intermediate hosts. To describe seasonal variation of nematode population variables and elucidate the timing of critical events in the parasite life cycle, nematodes were collected from both hosts across three collection periods in the ant-and-lizard activity season of 2008 in the Alvord Basin of southeastern Oregon. Among 3 collection periods, and across the activity season, nematodes were harvested from individual Phrynosoma platyrhinos , and the distribution of developmental categories and body lengths of nematodes was analyzed to determine the seasonal change in nematode population composition. Pogonomyrmex spp. ants were collected in pit-fall traps and dissected to determine infection prevalence. The abundance of non-gravid female and juvenile nematodes collected from lizards' stomachs decreased significantly between the early and late collection period, which was likely a consequence of the sequential conversion of these developmental categories to gravid females. The presence of gravid female nematodes peaked in cloacal and fecal collections during mid-season. The body lengths of male nematodes increased as the activity season progressed, perhaps due to growth, but their abundance remained the same. Smaller juvenile nematodes were present in late-season collections from lizards, possibly indicating new acquisitions from infected ants. We propose that once a set population of male nematodes establishes in lizards' stomachs, newly acquired juvenile nematodes develop into non-gravid females that mate, become gravid females, and exit the lizard mid-season. We additionally suggest that the exit of females may be timed with the peak foraging activity of ant intermediate hosts and access to larval ants in the nests. Infection prevalence in the intermediate host was low, with only 1 of 6,000 dissected harvester ants containing a single larval nematode. The temporal dynamics of S. phrynosoma populations within P. platyrhinos at this northern locale is most likely driven by the seasonal availability of harvester ant intermediate hosts.     

Ovarian cycle of southern brown howler monkey (<Emphasis Type="Italic">Alouatta guariba clamitans</Emphasis>) through fecal progestin measurement

The ovarian cycle in howler monkeys (genus Alouatta) has beean investigated through several biological parameters (ranging between 16.3 and 29.5 days); however, no data exist concerning the ovarian activity of the southern brown howler monkey (Alouatta guariba clamitans). This study aimed to describe the ovarian cycle of A. g. clamitans by profiling fecal progestin concentrations. Over 20 weeks, fecal samples of eight captive adult females of A. g. clamitans were collected. The collections were made at dawn, 5 days a week, and the samples were frozen immediately following collection. Next, they were dried, pulverized and hormonal metabolites were extracted to determine progestin concentrations by enzyme immunoassay. Of the 758 samples tested, the mean concentration of fecal progestins was 2866.40 ± 470.03 ng/g of dry feces, while the mean concentration at baseline was 814.47 ± 164.36 ng/g of dry feces. Among the eight females, one showed no ovarian cyclicity and three presented periods of probable absence of cyclicity and low progestin concentrations. A mean duration of 16 ± 0.52 days was observed for the 35 cycles studied. The interluteal phase lasted 4 ± 0.37 days on average, with a mean concentration of fecal progestins of 467.98 ± 29.12 ng/g of dry feces, while the luteal phase lasted 11 ± 0.50 days, with a mean concentration of 4283.27 ± 193.31 ng/g of dry feces. Besides describing the characteristics of the ovarian cycle, possible causes for the low concentrations of fecal progestins and periods of absence of cyclicity are also discussed.     

Urinary and fecal immunoglobulin A, cortisol and 11-17 dioxoandrostanes, and serum cortisol in metabolic cage housed female cynomolgus monkeys (Macaca fascicularis)

BACKGROUND AND METHODS: Quantitative enzyme-immunoassays of urinary and fecal immunoglobulin A (IgA), cortisol and 11-17-dioxoandrostanes (11,17-DOA), and serum cortisol in eight metabolic-cage-housed female cynomolgus monkeys were performed. The monkeys were divided into two groups, B and NB. Group B animals were blood sampled every 6 hours, whereas Group NB animals were not handled/blood sampled. RESULTS: No differences were recorded between the amounts of feces and urine excreted by the two groups. Group B animals excreted more urinary cortisol than did Group NB animals indicating that restraint-blood sampling resulted in a stress response. Excreted amounts of IgA and 11,17-DOA (urine and feces) did not differ between the groups. CONCLUSIONS: Urinary cortisol was a reliable marker of the stress associated with repeated blood sampling. Declining amounts of excreted urinary cortisol indicated that cynomolgus monkeys acclimated quickly to repeated blood sampling in metabolism cages. Within and between animal variation in amounts of feces voided demonstrated the importance of expressing fecal markers as 'amounts excreted per time unit per kg body weight' rather than just measuring the concentrations in fecal samples.     

Noninvasive monitoring of fecal cortisol metabolites in the eastern chipmunk (Tamias striatus): validation and comparison of two enzyme immunoassays

Monitoring fecal glucocorticoid metabolites in wild animals, using enzyme immunoassays, enables the study of endocrinological patterns relevant to ecology and evolution. While some researchers use antibodies against the parent hormone (which is typically absent from fecal samples), others advocate the use of antibodies designed to detect glucocorticoid metabolites. We validated two assays to monitor fecal cortisol metabolites in the eastern chipmunk (Tamias striatus). We compared an antibody produced against cortisol and one produced against 5α-pregnane-3β, 11β, 21-triol-20-one using a radiometabolism study and an injection with adrenocorticotropic hormone (ACTH). Most cortisol metabolites were excreted in the urine (～83%). Peak excretion in the feces occurred 8 h after injection. Both assays detected an increase in fecal cortisol metabolite levels after injection of ACTH. Males, but not females, exhibited a circadian variation in metabolite levels. The sexes did not exhibit any difference over the time course and route of excretion or the relative increase in fecal cortisol metabolite levels after ACTH injection. The cortisol assay displayed higher reactivity to ACTH injection relative to baseline than did the metabolite assay. While both antibodies gave comparable results, the cortisol antibody was more sensitive to changes in plasma cortisol levels in eastern chipmunks.