Thirty years of conservation breeding: Assessing the genetic diversity of captive Livingstone's fruit bats.

Fruit bats (genus Pteropus) are typically island-endemic species important in seed dispersal and reforestation that are vulnerable to increased extinction risk. An effective method of reducing extinction risk in vulnerable species that cannot be conserved in their native habitat is establishing an ex-situ captive breeding programme. Due to anthropogenic threats and low population numbers, in the early 1990s, a captive breeding programme was established at Jersey Zoo, British Isles, for Critically Endangered Livingstone's fruit bats (Pteropus livingstonii). Here we use six polymorphic microsatellite loci to assess genetic diversity in the captive breeding population of Livingstone's fruit bats (P. livingstonii), 30 years after the programme's establishment, investigating change over generations and comparing our findings with published data from the wild population. We found no significant difference between the genetic diversity in the captive and wild populations of Livingstone's fruit bats (P. livingstonii), in both expected heterozygosity and allelic richness. The captive population has retained a comparable level of genetic diversity to that documented in the wild, and there has been no significant decline in genetic diversity over the last 30 years. We advise that a full pedigree of the paternal lineage is created to improve the management of the captive breeding programme and further reduce the possibility of inbreeding. However, it appears that the captive breeding programme is currently effective at maintaining genetic diversity at levels comparable to those seen in the wild population, which suggests reintroductions could be viable if genetic diversity remains stable in captivity.

Weight as an indicator of enclosure suitability in Livingstone's fruit bats (Pteropus livingstonii).

Obesity is common in zoo animals, and both dietary management and the provision of adequate opportunities for exercise are needed to tackle it. We used 30 years of records from Jersey Zoo to compare the weight and forearm length of wild and captive-born Livingstone's fruit bats (Pteropus livingstonii), and to assess the impact on weight of enclosure space. The mean capture weight of wild-caught male Livingstone's bats was 657 g, significantly higher than that of females (544 g). In both wild and captive-born bats, males had significantly longer forearms than females, but there was no effect of birth location. Males weighed more in the mating season than at other times of year. Both sexes gained more weight during development if born in enclosures that restricted flight rather than a large aviary; this was particularly noticeable in females. After reaching maturity at 3 years, weights of bats born in restricted enclosures continued to increase, reached a peak of over 1000 g at 8-10 years, and then declined in both sexes. The weight of bats born in the aviary remained more stable after the age of three. Like wild bats, adult females born in the aviary weighed less than males. However, females born in restricted enclosures weighed more than males born in the same enclosures. Enclosure designs that maximize opportunities for flight can limit excessive weight gain in captive fruit bats and may therefore improve fitness and health, essential in planning for future reintroduction programs.

Social roles influence cortisol levels in captive Livingstone's fruit bats (Pteropus livingstonii).

A critical component of conserving and housing species ex situ is an explicit scientific understanding of the physiological underpinnings of their welfare. Cortisol has been repeatedly linked to stress, and therefore used as an indicator of welfare for many species. In order to measure cortisol in the Livingstone's fruit bat (Pteropus livingstonii; a critically endangered keystone species) without disturbing the captive population, we have developed and validated a non-invasive, novel hormone extraction procedure and faecal glucocorticoid assay. A total of 92 faecal samples, 73 from the P. livingstonii breeding colony at Jersey Zoo, Channel Islands and 19 samples from P. livingstonii housed at Bristol Zoological Gardens, UK, have been collected and analyzed. Mixed-effect modelling of the influence of physiological state variables on cortisol concentration revealed that lactating females had higher cortisol levels than non-lactating females, indicating that our assay is measuring biologically relevant hormone concentrations. Males and older bats also had higher cortisol than non-lactating females and younger individuals. Further analysis applied social network methodology to compare the cortisol levels of bats with different social roles. We found that individuals that linked social groups possessed higher than average cortisol levels and conversely, individuals with high-quality, positive relationships had lower cortisol levels. These results demonstrate, for the first time in a bat species, social mediation of stress hormones. Lastly, the frequency of vocalisation was found to positively correlate with cortisol concentration in males, suggesting that this behaviour may be used by animal management as a visual indicator of a bat's hormonal status. Hence, this research has provided unique insights and empirical scientific knowledge regarding the relationship between the physiology and social behaviour of P. livingstonii, therefore allowing for recommendations to be made to optimise bat welfare at the individual level.

Individual, social, and environmental factors affecting salivary and fecal cortisol levels in captive pied tamarins (Saguinus bicolor).

Pied tamarins (Saguinus bicolor) are endangered New World primates, and in captivity appear to be very susceptible to stress. We measured cortisol in 214 saliva samples from 36 tamarins and in 227 fecal samples from 27 tamarins, and investigated the effects of age, sex, pregnancy, rearing history, social status, weight, group composition, and enclosure type using generalized linear mixed models. There was no effect of age on either fecal or salivary cortisol levels. Female pied tamarins in late pregnancy had higher fecal cortisol levels than those in early pregnancy, or nonpregnant females, but there was no effect of pregnancy on salivary cortisol. Females had higher salivary cortisol levels than males, but there was no effect of rearing history. However, for fecal cortisol, there was an interaction between sex and rearing history. Hand-reared tamarins overall had higher fecal cortisol levels, but while male parent-reared tamarins had higher levels than females who were parent-reared, the reverse was true for hand-reared individuals. There was a trend towards lower fecal cortisol levels in subordinate individuals, but no effect of status on salivary cortisol. Fecal but not salivary cortisol levels declined with increasing weight. We found little effect of group composition on cortisol levels in either saliva or feces, suggesting that as long as tamarins are housed socially, the nature of the group is of less importance. However, animals in off-show enclosures had higher salivary and fecal cortisol levels than individuals housed on-show. We suggest that large on-show enclosures with permanent access to off-exhibit areas may compensate for the effects of visitor disturbance, and a larger number of tamarins of the same species housed close together may explain the higher cortisol levels found in tamarins living in off-show accommodation, but further research is needed.

A comparative study of litter size and sex composition in a large dataset of callitrichine monkeys.

In many birds and mammals, the size and sex composition of litters can have important downstream effects for individual offspring. Primates are model organisms for questions of cooperation and conflict, but the factors shaping interactions among same-age siblings have been less-studied in primates because most species bear single young. However, callitrichines (marmosets, tamarins, and lion tamarins) frequently bear litters of two or more, thereby providing the opportunity to ask whether variation in the size and sex composition of litters affects development, survival, and reproduction. To investigate these questions, we compiled a large dataset of nine species of callitrichines (n = 27,080 individuals; Callithrix geoffroyi, Callithrix jacchus, Cebuella pygmaea, Saguinus imperator, Saguinus oedipus, Leontopithecus chrysomelas, Leontopithecus chrysopygus, Leontopithecus rosalia, and Callimico goeldii) from zoo and laboratory populations spanning 80 years (1938-2018). Through this comparative approach, we found several lines of evidence that litter size and sex composition may impact fitness. Singletons have higher survivorship than litter-born peers and they significantly outperform litter-born individuals on two measures of reproductive performance. Further, for some species, individuals born in a mixed-sex litter outperform isosexually-born individuals (i.e., those born in all-male or all-female litters), suggesting that same-sex competition may limit reproductive performance. We also document several interesting demographic trends. All but one species (C. pygmaea) has a male-biased birth sex ratio with higher survivorship from birth to sexual maturity among females (although this was significant in only two species). Isosexual litters occurred at the expected frequency (with one exception: C. pygmaea), unlike other animals, where isosexual litters are typically overrepresented. Taken together, our results indicate a modest negative effect of same-age sibling competition on reproductive output in captive callitrichines. This study also serves to illustrate the value of zoo and laboratory records for biological inquiry.

Flight patterns in zoo-housed fruit bats (Pteropus spp.).

Maintaining the capacity for sustained flight in captivity is a key goal for the management of threatened fruit bats. We developed quantifiable descriptions of flight complexity and used them to assess the suitability of an enclosure for two species of fruit bat of differing size, the large Livingstone's fruit bat, Pteropus livingstonii, and the smaller Rodrigues fruit bat, Pteropus rodricensis, in a two-part study. In Phase 1, Rodrigues fruit bats flew more often than Livingstone's fruit bats and although the majority of flights in both species were linear, Rodrigues fruit bats were more likely to display complex flight paths involving turns, while flights by Livingstone's fruit bats were more likely to end in a crash-landing than Rodrigues fruit bat flights. The enclosure may therefore not have been large enough for Livingstone's fruit bats to display a full range of flight behavior over longer distances. In Phase 2, juvenile Livingstone's fruit bats ( < 3 years old) flew more than twice as often as younger adults (3-10 years old) did. Older adult Livingstone's fruit bats over the age of 10 years were not observed to fly. We could not separate out the effects of age, weight and environment during development as these factors were strongly correlated in our study; future work in this area will be very important in understanding the factors that affect flight in captive bats, and how it can be encouraged by appropriate enclosure design.

Genetic assessment for the endangered black lion tamarin Leontopithecus chrysopygus (Mikan, 1823), Callitrichidae, Primates.

This is the first study analyzing genetic diversity in captive individuals of the endangered black lion tamarin, Leontopithecus chrysopygus, and also comparing genetic diversity parameters between wild populations and captive groups using the same set of molecular markers. We evaluated genetic diversity and differentiation for the Brazilian and European captive groups and a wild population through 15 polymorphic microsatellite markers. The genetic diversity levels were similar among Brazilian captive, European captive and wild animals from the National Forest of Capão Bonito. Expected heterozygosity showed values ranging from 0.403 to 0.462, and significant differences were not observed among the populations. Different allele frequencies were observed among the groups, which showed the presence of distinct private alleles. The PCoA analysis evidenced three main clusters suggesting that the captive Brazilian and European groups are markedly differentiated both from one another and from the wild population of Capão Bonito. Likewise, the most likely number of genetic clusters (K) revealed by Structure was three. Such a structure is probably the result of the strength of drift and non-random reproduction in these small and isolated groups. Despite this differentiation, all groups still have similar genetic diversity levels, comparable to other callitrichids. The data obtained herein are important to increasing knowledge of the genetics of tamarins and supporting breeding programs to prevent loss of genetic diversity and inbreeding depression.

Why do captive pied tamarins give birth during the day?

Diurnal primates typically give birth at night, when it is presumed that they are safer at a very vulnerable time, and this is reflected in an overwhelmingly nocturnal pattern of delivery in most species of Callitrichidae. However, over half (51.1%) of 88 births to pied tamarins (Saguinus bicolor) at Durrell Wildlife Park occurred during the day (0800-1700), almost always in the afternoon. Nearly three quarters of breeding females (17/23) had at least one diurnal birth, including females from all generations in captivity from wild-caught to fifth captive-born generation, and from all six matrilines represented at Durrell. The proportion of diurnal births has remained relatively stable over time despite management changes. We used generalized linear mixed modeling to investigate several factors that we hypothesized could affect time of birth: maternal experience, season, female rearing history, and whether or not the group was on public display. We fitted all possible models to the data, but none explained more than 7.5% of the variation. Daytime delivery had few statistically significant detrimental effects, although infant survival was somewhat lower and parental rejection increased in diurnal births. Pied tamarins do not seem to fit any of the hypotheses previously put forward to explain exceptions to the typical primate circadian pattern of delivery. Zoo Biol. 35:487-494, 2016. © 2016 Wiley Periodicals, Inc.

Predictors of dominance rank and agonistic interactions in captive Livingstone's fruit bats.

Male dominance hierarchies have been studied in many animals but rarely in bats (Chiroptera). The dominance rank of social animals may dictate access to resources and mates; therefore, it has important implications for an individual's fitness and is crucial for successful captive management. Between January and December 2018, at both Bristol Zoo Gardens (Bristol, UK) and Jersey Zoo (Jersey, British Isles), we observed 19 male Livingstone's fruit bats Pteropus livingstonii using focal follows for 345 h overall, noting the outcome of all agonistic interactions. We recorded instigators of interactions, along with winners and losers, and analyzed these data using the R-package "EloRating" to create Elo-rating temporal plots of dominance ranks. We used generalized linear mixed models and multiple linear regression to analyze interaction data and test hypotheses regarding predictors of dominance rank, frequency of agonistic interaction, and choice of interaction partner. Age was positively correlated with dominance rank up to around year 9, when an asymptote was attained. Highly ranked bats instigated the most agonistic interactions, and largely directed these interactions at bats with much lower rankings than themselves. Hierarchies were extremely stable throughout the data collection period at both sites. We conclude that Livingstone's fruit bats have a stable linear dominance hierarchy, with high-ranking, typically older males instigating the most interactions with lowest ranking males to secure dominance rank. This study adds to the limited discourse on Pteropus social behaviors, indicating that some bat species may have social systems similar in complexity to some nonhuman primates.

After short interbirth intervals, captive callitrichine monkeys have higher infant mortality.

Life history theory predicts a trade-off between the quantity and quality of offspring. Short interbirth intervals-the time between successive births-may increase the quantity of offspring but harm offspring quality. In contrast, long interbirth intervals may bolster offspring quality while reducing overall reproductive output. Further research is needed to determine whether this relationship holds among primates, which have intensive parental investment. Using Cox proportional hazards models, we examined the effects of interbirth intervals (short, normal, or long) on infant survivorship using a large demographic dataset (n = 15,852) of captive callitrichine monkeys (marmosets, tamarins, and lion tamarins). In seven of the nine species studied, infants born after short interbirth intervals had significantly higher risks of mortality than infants born after longer interbirth intervals. These results suggest that reproduction in callitrichine primates may be limited by physiologic constraints, such that short birth spacing drives higher infant mortality.

Changes in the faecal microbiome of pied tamarins (Saguinus bicolor) associated with chronic, recurrent diarrhoea and weight loss.

BACKGROUND: Chronic recurrent diarrhoea and weight loss is a common problem in captive callitrichids. These symptoms are common clinical features of marmoset wasting syndrome (MWS), a chronic enteric inflammation of unknown aetiology associated with mortality in captive marmosets. The unknown aetiology of the condition presents problems for conservation projects where affected colonies present higher mortality and lower birth rates. Since a role for the microbiome has been established in chronic enteric inflammation of other species it is possible that the intestinal microbiome undergoes similar changes during MWS. RESULTS: The faecal microbiome of pied tamarins (Saguinus bicolor) at Jersey Zoo was determined using 16S rRNA gene amplicon sequencing to compare the composition of the faecal microbiome of tamarins affected by chronic recurrent diarrhoea and weight loss with unaffected individuals. Affected individuals had a higher relative abundance of amplicon sequence variants assigned to Lactobacillus and Helicobacter jaachi while unaffected individuals had a higher relative abundance of some Lachnospiraceae and Ruminococcaceae. CONCLUSIONS: Although Helicobacter has been shown to reside in healthy wild and captive marmosets and tamarins and appears to form part of the normal microbiota, the results of this study raise the prospect that certain species of Helicobacter may be associated with chronic, recurrent diarrhoea in captive callitrichids. The presence of Lactobacillus may also play a role in the development of MWS. Since depletion of Lachnospiraceae and Ruminococcaceae have been linked to chronic gastrointestinal inflammation in humans, this feature of the microbiome of affected tamarins provides another avenue of further research in the pathogenesis of MWS.

Social Experience of Captive Livingstone's Fruit Bats (Pteropus livingstonii).

Social network analysis has been highlighted as a powerful tool to enhance the evidence-based management of captive-housed species through its ability to quantify the social experience of individuals. We apply this technique to explore the social structure and social roles of 50 Livingstone's fruit bats (Pteropus livingstonii) housed at Jersey Zoo, Channel Islands, through the observation of associative, affiliative, and aggressive interactions over two data collection periods. We implement binomial mixture modelling and characteristic-based assortment quantification to describe the complexity and organisation of social networks, as well as a multiple regression quadratic assignment procedural (MRQAP) test to analyse the relationship between network types. We examine the effects of individual characteristics (i.e., sex, age, and dominance rank) on social role by fitting models to explain the magnitude of node metrics. Additionally, we utilize a quadratic assignment procedural (QAP) test to assess the temporal stability of social roles over two seasons. Our results indicate that P. livingstonii display a non-random network structure. Observed social networks are positively assorted by age, as well as dominance rank. The frequency of association between individuals correlates with a higher frequency of behavioural interactions, both affiliative and aggressive. Individual social roles remain consistent over ten months. We recommend that, to improve welfare and captive breeding success, relationships between individuals of similar ages and dominance levels should be allowed to persist in this group where possible, and separating individuals that interact frequently in an affiliative context should be avoided.

Studbook and molecular analyses for the endangered black-lion-tamarin; an integrative approach for assessing genetic diversity and driving management in captivity.

Breeding strategies based on molecular markers have been adopted by ex-situ conservation programs to assess alternative parameters for the genetic diversity estimates. In this work we evaluated molecular and studbook data for captive populations of black-lion-tamarin (BLT), an endangered primate endemic to Brazil's Atlantic Forest. Pedigree analyses were performed using BLT studbook information collected from 1973 to 2018. We analyzed the whole captive population since its foundation; the current captive population (CCP); and all extant BLTs in the Brazilian captive population (BCP), separately. Microsatellite analyses were implemented on the BCP individuals from the eighth generation (BCP-F8) only to avoid generation overlap. The expected heterozygosity for BCP-F8, using molecular, data was 0.45, and the initial expected heterozygosity was 0.69. Kinship parameters showed high genetic relationships in both pedigree and molecular analyses. The genealogy-based endogamy evidenced a high inbreeding coefficient, while the molecular analyses suggested a non-inbreeding signature. The Mate Suitability Index showed detrimental values for the majority of potential pairs in the CCP. Nevertheless, some individuals evidenced high individual heterozygosity and allele representation, demonstrating good potential to be used as breeders. Thus, we propose the use of molecular data as a complementary parameter to evaluate mating-pairs and to aid management decision-making.