Factors influencing terrestriality in primates of the Americas and Madagascar.

Among mammals, the order Primates is exceptional in having a high taxonomic richness in which the taxa are arboreal, semiterrestrial, or terrestrial. Although habitual terrestriality is pervasive among the apes and African and Asian monkeys (catarrhines), it is largely absent among monkeys of the Americas (platyrrhines), as well as galagos, lemurs, and lorises (strepsirrhines), which are mostly arboreal. Numerous ecological drivers and species-specific factors are suggested to set the conditions for an evolutionary shift from arboreality to terrestriality, and current environmental conditions may provide analogous scenarios to those transitional periods. Therefore, we investigated predominantly arboreal, diurnal primate genera from the Americas and Madagascar that lack fully terrestrial taxa, to determine whether ecological drivers (habitat canopy cover, predation risk, maximum temperature, precipitation, primate species richness, human population density, and distance to roads) or species-specific traits (body mass, group size, and degree of frugivory) associate with increased terrestriality. We collated 150,961 observation hours across 2,227 months from 47 species at 20 sites in Madagascar and 48 sites in the Americas. Multiple factors were associated with ground use in these otherwise arboreal species, including increased temperature, a decrease in canopy cover, a dietary shift away from frugivory, and larger group size. These factors mostly explain intraspecific differences in terrestriality. As humanity modifies habitats and causes climate change, our results suggest that species already inhabiting hot, sparsely canopied sites, and exhibiting more generalized diets, are more likely to shift toward greater ground use.

An integrated passive acoustic monitoring and deep learning pipeline for black-and-white ruffed lemurs (Varecia variegata) in Ranomafana National Park, Madagascar.

The urgent need for effective wildlife monitoring solutions in the face of global biodiversity loss has resulted in the emergence of conservation technologies such as passive acoustic monitoring (PAM). While PAM has been extensively used for marine mammals, birds, and bats, its application to primates is limited. Black-and-white ruffed lemurs (Varecia variegata) are a promising species to test PAM with due to their distinctive and loud roar-shrieks. Furthermore, these lemurs are challenging to monitor via traditional methods due to their fragmented and often unpredictable distribution in Madagascar's dense eastern rainforests. Our goal in this study was to develop a machine learning pipeline for automated call detection from PAM data, compare the effectiveness of PAM versus in-person observations, and investigate diel patterns in lemur vocal behavior. We did this study at Mangevo, Ranomafana National Park by concurrently conducting focal follows and deploying autonomous recorders in May-July 2019. We used transfer learning to build a convolutional neural network (optimized for recall) that automated the detection of lemur calls (57-h runtime; recall = 0.94, F1 = 0.70). We found that PAM outperformed in-person observations, saving time, money, and labor while also providing re-analyzable data. Using PAM yielded novel insights into V. variegata diel vocal patterns; we present the first published evidence of nocturnal calling. We developed a graphic user interface and open-sourced data and code, to serve as a resource for primatologists interested in implementing PAM and machine learning. By leveraging the potential of this pipeline, we can address the urgent need for effective primate population surveys to inform conservation strategies.

Functional relationships between estradiol and paternal care in male red-bellied lemurs, Eulemur rubriventer.

Fathers contribute substantially to infant care, yet the mechanisms facilitating paternal bonding and interactions with infants are not as well understood as they are in mothers. Several hormonal changes occur as males transition into parenthood, first in response to a partner's pregnancy, and next in response to interacting with the newborn. These changes may prepare fathers for parenting and help facilitate and maintain paternal care. Experimental studies with monkeys and rodents suggest that paternal care requires elevated estradiol levels, which increase when a male's partner is pregnant and are higher in fathers than non-fathers, but its role in the expression of paternal behaviors throughout infant development is unknown. To assess estradiol's role in paternal care, we analyzed the relationship between paternal estradiol metabolites and 1) offspring age, and 2) paternal care behavior (holding, carrying, huddling, playing, grooming), in wild, red-bellied lemurs (Eulemur rubriventer). We collected 146 fecal samples and 1597 h of behavioral data on 10 adult males who had newborn infants during the study. Estradiol metabolites increased four-fold in expectant males, and in new fathers they fluctuated and gradually decreased with time. Infant age, not paternal behavior, best predicted hormone levels in new fathers. These results suggest that hormonal changes occur in expectant males with facultative paternal care, but they do not support the hypothesis that estradiol is directly associated with the day-to-day expression of paternal care. Future research should explore estradiol's role in facilitating behaviors, including infant-directed attention and responsiveness, or preparing fathers for infant care generally.

Nutrient balancing in a fruit-specialist primate, the black-and-white ruffed lemur (Varecia variegata).

Animals' foraging behavior and dietary choices are, in part, driven by their ultimate function: to meet nutritional demands. However, depending on their degree of dietary specialization and the availability and distribution of food resources in their environment, species may utilize different nutritional strategies. With shifting plant phenology, increasing unpredictability of fruiting, and declining food quality in response to anthropogenic climate change, existing nutritional constraints may become exacerbated. Such changes are especially concerning for Madagascar's endemic fruit specialists given the nutrient-limitation of the island's landscapes. In this study, we examined the nutritional strategy of one such fruit-specialist primate, the black-and-white ruffed lemur (Varecia variegata), over a 12-month period (January to December 2018) in Ranomafana National Park, Madagascar. We hypothesized that Varecia would balance nonprotein energy (NPE) to protein (AP) at a high ratio similar to other frugivorous primates, and that they would prioritize protein intake given their high degree of frugivory. We found that Varecia balance NPE:AP at a ratio of 11:1, higher than in any other primate studied to date; however, diets shifted such that nutrient balancing varied seasonally (12.6:1 abundant-9.6:1 lean). Varecia meet NRC suggested recommendations of 5-8% of calories from protein, despite having a diet mostly comprising fruits. However, seasonal shifts in NPE intakes result in significant energy shortfalls during fruit-lean seasons. Flowers provide an important source of NPE during these periods, with flower consumption best predicting lipid intake, suggesting this species' ability to shift resource use. Nevertheless, achieving adequate and balanced nutrient intakes may become precarious in the face of increasing unpredictability in plant phenology and other environmental stochasticities resulting from climate change.

Urinary C-peptide and total triiodothyronine as energetic biomarkers for studies of lemurs.

Measuring energy balance and energy metabolism can provide crucial information for understanding the ecological and behavioral drivers of an animal's energetic and physiological condition. Both urinary C-peptide (uCP) of insulin and urinary total triiodothyronine (uTT3) have been validated as noninvasive biomarkers of energy balance and metabolic activity in haplorrhine primates. This study attempts to validate uCP and uTT3 measures in strepsirrhines, a phylogenetically distinct primate clade, using the ruffed lemur (genus Varecia) as a model. We experimentally manipulated the diet of captive black-and-white (Varecia variegata) and red (Varecia rubra) ruffed lemurs at Duke Lemur Center across a 4-week period. We collected urine samples from subjects (n = 5) each day during 1 week of control diet, 2 weeks of calorie-restricted diet and 1 week of refeeding, designed to temporarily reduce energy balance and metabolism. We also tested the outcome of filter paper as a storage method by comparing to controls (frozen at -20°C) to assess its suitability for studies of wild populations. We successfully measured uCP and uTT3 levels in frozen urine samples using commercial enzyme immunoassay kits and found that both biomarkers were excreted at lower concentrations (C-peptide: 1.35 ng/mL, 54% reduction; TT3: 1.5 ng/mL, 37.5% reduction) during calorie-restricted periods compared to normal diet periods. Filter paper recovery for uCP was 19%, though values were significantly positively correlated with frozen control samples. uTT3 could not be recovered at measurable concentrations using filter paper. These methods enable noninvasive measurement of energetic conditions in wild strepsirrhines and subsequent assessment of relationships between energy balance and numerous socioecological drivers in primate populations.

Primate landscape genetics: A review and practical guide.

Landscape genetics is an emerging field that integrates population genetics, landscape ecology, and spatial statistics to investigate how geographical and environmental features and evolutionary processes such as gene flow, genetic drift, and selection structure genetic variation at both the population and individual levels, with implications for ecology, evolution, and conservation biology. Despite being particularly well suited for primatologists, this method is currently underutilized. Here, we synthesize the current state of research on landscape genetics in primates. We begin by outlining how landscape genetics has been used to disentangle the drivers of diversity, followed by a review of how landscape genetic methods have been applied to primates. This is followed by a section highlighting special considerations when applying the methods to primates, and a practical guide to facilitate further landscape genetics studies using both existing and de novo datasets. We conclude by exploring future avenues of inquiry that could be facilitated by recent developments as well as underdeveloped applications of landscape genetics to primates.

Seasonal variability in the diet and feeding ecology of black-and-white ruffed lemurs (Varecia variegata) in Ranomafana National Park, southeastern Madagascar.

OBJECTIVES: We characterized the diet and foraging ecology of the black-and-white ruffed lemur (Varecia variegata), a specialized frugivore, and investigated behavioral strategies exhibited in response to seasonal changes in resource availability. MATERIALS AND METHODS: Behavioral data were collected from the same two adjacent communities across 29 months during two observation periods (2007-2008; 2017-2018) in Mangevo, a primary rainforest habitat in southeastern Madagascar. To analyze feeding in the context of energy maximization versus time minimization strategies, we used nonparametric tests to compare plant part constituents, dietary diversity, activity budgets, and canopy strata use between fruit-abundant versus fruit-lean seasons. RESULTS: Individuals dedicated ~30% of their time to feeding year-round, mostly in the middle canopy (11-20 m). Animals fed primarily on fruits (74% of diet), but frugivory decreased and folivory increased markedly during fruit-lean seasons. Abundant season dietary diversity (98 taxa, H' = 0.71-1.37) was greater than lean season diversity (70 taxa, H' = 0.56-1.06), which coincided with less traveling, more resting, and higher canopy use-though interannual variation was observed. CONCLUSIONS: Herein, we describe behavioral and dietary patterns that are concordant with a time minimizing behavioral strategy. Black-and-white ruffed lemur diets comprised lower taxonomic diversity, fewer fruits, and more leaves during fruit-lean months. Further, shifts toward less travel, more resting, and greater use of higher canopy levels during this time were most likely for thermoregulatory benefits.

Fermented food consumption in wild nonhuman primates and its ecological drivers.

OBJECTIVES: Although fermented food use is ubiquitous in humans, the ecological and evolutionary factors contributing to its emergence are unclear. Here we investigated the ecological contexts surrounding the consumption of fruits in the late stages of fermentation by wild primates to provide insight into its adaptive function. We hypothesized that climate, socioecological traits, and habitat patch size would influence the occurrence of this behavior due to effects on the environmental prevalence of late-stage fermented foods, the ability of primates to detect them, and potential nutritional benefits. MATERIALS AND METHODS: We compiled data from field studies lasting at least 9 months to describe the contexts in which primates were observed consuming fruits in the late stages of fermentation. Using generalized linear mixed-effects models, we assessed the effects of 18 predictor variables on the occurrence of fermented food use in primates. RESULTS: Late-stage fermented foods were consumed by a wide taxonomic breadth of primates. However, they generally made up 0.01%-3% of the annual diet and were limited to a subset of fruit species, many of which are reported to have mechanical and chemical defenses against herbivores when not fermented. Additionally, late-stage fermented food consumption was best predicted by climate and habitat patch size. It was more likely to occur in larger habitat patches with lower annual mean rainfall and higher annual mean maximum temperatures. DISCUSSION: We posit that primates capitalize on the natural fermentation of some fruits as part of a nutritional strategy to maximize periods of fruit exploitation and/or access a wider range of plant species. We speculate that these factors contributed to the evolutionary emergence of the human propensity for fermented foods.

Maternal feeding benefits of allomaternal care in black-and-white colobus (Colobus guereza).

Caring for infants involves lactation, protection, provisioning, and carrying-all energetically taxing states for primate mothers. Holding and carrying clinging infants often constrains mothers from moving and traveling, potentially reducing their food and energy intake; however, when separated from its mother an infant is at risk of predation. This separation therefore requires that mothers be vigilant, further deterring them from feeding. Allomaternal care (AMC) is hypothesized to allow mothers to safely detach from their infants to feed, permitting them to increase energy intake, which is particularly needed for lactation. We examined the nutritional benefits of AMC in black-and-white colobus monkeys (Colobus guereza) by estimating energy intake by lactating mothers during AMC versus non-AMC. We studied seven mother-infant dyads in three groups of C. guereza during six months in Kibale National Park, Uganda. Immature group members handled infants more often than adults, and females handled infants more often than males. An infant's distance to its mother and its nearest neighbor's age and sex best predicted the occurrence of AMC. Lactating mothers fed more often, fed and rested for longer durations, and consumed more metabolizable energy during AMC compared to when they were caring for their infants. These results demonstrate that AMC in C. guereza provides mothers with feeding opportunities that increase their energy intake.

Sex-Segregated Range Use by Black-and-White Ruffed Lemurs (Varecia variegata) in Ranomafana National Park, Madagascar.

Ranging behavior is one important strategy by which nonhuman primates obtain access to resources critical to their biological maintenance and reproductive success. As most primates live in permanent social groups, their members must balance the benefits of group living with the costs of intragroup competition for resources. However, some taxa live in more spatiotemporally flexible social groups, whose members modify patterns of association and range use as a method to mitigate these costs. Here, we describe the range use of one such taxon, the black-and-white ruffed lemur (Varecia variegata), at an undisturbed primary rain forest site in Ranomafana National Park, Madagascar, and characterize sex differences in annual home range area, overlap, and daily distances traveled. Moreover, we characterize seasonal variability in range use and ask whether ranging behaviors can be explained by either climatic or reproductive seasonality. We found that females used significantly larger home ranges than males, though sexes shared equal and moderate levels of home range overlap. Overall, range use did not vary across seasons, although within sexes, male range use varied significantly with climate. Moreover, daily path length was best predicted by day length, female reproductive state, and sex, but was unrelated to climate variables. While the patterns of range use and spatial association presented here share some similarities with "bisexually bonded" community models described for chimpanzees, we argue that ruffed lemurs best conform to a "nuclear neighborhood" community model wherein nuclear (core) groups share the highest levels of home range overlap, and where these groups cluster spatially into adjacent "neighborhoods" within the larger, communally defended territory.

HEALTH ASSESSMENT OF BLACK-AND-WHITE RUFFED LEMURS (VARECIA VARIEGATA) IN RANOMAFANA NATIONAL PARK, MADAGASCAR.

The Anthropocene is a major threat to biodiversity worldwide. Human pressures including climate change and emerging infectious diseases are presenting new challenges to wildlife, requiring vigilance and monitoring of wild populations to ensure their persistence. In order to monitor fluctuations in health, baseline data from long-term studies are required. Clinical laboratory data on 80 black-and-white ruffed lemurs (Varecia variegata), derived from the capture of 98 individuals spanning six field seasons are presented. Serum biochemical profiles showed variation between years that remained within published reference intervals for the species, with the exception of total bilirubin in 2008, and creatine kinase and chloride in 2019. Serum trace minerals and fat-soluble vitamin values also fluctuated between years and are within ranges seen in other lemur species. These results, combined with previously published data on ectoparasite load and population genetic diversity, suggest that the Mangevo ruffed lemur population is healthy and can provide important and valuable baseline data for comparisons moving forward.

Implications of lemuriform extinctions for the Malagasy flora.

Madagascar's lemurs display a diverse array of feeding strategies with complex relationships to seed dispersal mechanisms in Malagasy plants. Although these relationships have been explored previously on a case-by-case basis, we present here the first comprehensive analysis of lemuriform feeding, to our knowledge, and its hypothesized effects on seed dispersal and the long-term survival of Malagasy plant lineages. We used a molecular phylogenetic framework to examine the mode and tempo of diet evolution, and to quantify the associated morphological space occupied by Madagascar's lemurs, both extinct and extant. Using statistical models and morphometric analyses, we demonstrate that the extinction of large-bodied lemurs resulted in a significant reduction in functional morphological space associated with seed dispersal ability. These reductions carry potentially far-reaching consequences for Malagasy ecosystems, and we highlight large-seeded Malagasy plants that appear to be without extant animal dispersers. We also identify living lemurs that are endangered yet occupy unique and essential dispersal niches defined by our morphometric analyses.

SMARTA: Automated testing apparatus for visual discrimination tasks.

This article introduces the open-source Subject-Mediated Automatic Remote Testing Apparatus (SMARTA) for visual discrimination tasks, which aims to streamline and ease data collection, eliminate or reduce observer error, increase interobserver agreement, and automate data entry without the need for an internet connection. SMARTA is inexpensive and easy to build, and it can be modified to accommodate a variety of experimental designs. Here we describe the utility and functionality of SMARTA in a captive setting. We present the results from a case study of color vision in ruffed lemurs (Varecia spp.) at the Duke Lemur Center in Durham, North Carolina, in which we demonstrate SMARTA's utility for two-choice color discrimination tasks, as well as its ability to streamline and standardize data collection. We also include detailed instructions for constructing and implementing the fully integrated SMARTA touchscreen system.

Social behaviour and gut microbiota in red-bellied lemurs (Eulemur rubriventer): In search of the role of immunity in the evolution of sociality.

Vertebrate gut microbiota form a key component of immunity and a dynamic link between an individual and the ecosystem. Microbiota might play a role in social systems as well, because microbes are transmitted during social contact and can affect host behaviour. Combining methods from behavioural and molecular research, we describe the relationship between social dynamics and gut microbiota of a group-living cooperative species of primate, the red-bellied lemur (Eulemur rubriventer). Specifically, we ask whether patterns of social contact (group membership, group size, position in social network, individual sociality) are associated with patterns of gut microbial composition (diversity and similarity) between individuals and across time. Red-bellied lemurs were found to have gut microbiota with slight temporal fluctuations and strong social group-specific composition. Contrary to expectations, individual sociality was negatively associated with gut microbial diversity. However, position within the social network predicted gut microbial composition. These results emphasize the role of the social environment in determining the microbiota of adult animals. Since social transmission of gut microbiota has the potential to enhance immunity, microbiota might have played an escalating role in the evolution of sociality.

Novel opsin gene variation in large-bodied, diurnal lemurs.

Some primate populations include both trichromatic and dichromatic (red-green colour blind) individuals due to allelic variation at the X-linked opsin locus. This polymorphic trichromacy is well described in day-active New World monkeys. Less is known about colour vision in Malagasy lemurs, but, unlike New World monkeys, only some day-active lemurs are polymorphic, while others are dichromatic. The evolutionary pressures underlying these differences in lemurs are unknown, but aspects of species ecology, including variation in activity pattern, are hypothesized to play a role. Limited data on X-linked opsin variation in lemurs make such hypotheses difficult to evaluate. We provide the first detailed examination of X-linked opsin variation across a lemur clade (Indriidae). We sequenced the X-linked opsin in the most strictly diurnal and largest extant lemur, Indri indri, and nine species of smaller, generally diurnal indriids (Propithecus). Although nocturnal Avahi (sister taxon to Propithecus) lacks a polymorphism, at least eight species of diurnal indriids have two or more X-linked opsin alleles. Four rainforest-living taxa-I. indri and the three largest Propithecus species-have alleles not previously documented in lemurs. Moreover, we identified at least three opsin alleles in Indri with peak spectral sensitivities similar to some New World monkeys.

Comparing primate crania: The importance of fossils.

OBJECTIVES: Extant primate crania represent a small subset of primate crania that have existed. The main objective here is to examine how the inclusion of fossil crania changes our understanding of primate cranial diversity relative to analyses of extant primates. We hypothesize that fossil taxa will change the major axes of cranial shape, occupy new areas of morphospace, change the relative diversity of major primate clades, and fill in notable gaps separating major primate taxa/clades. MATERIALS AND METHODS: Eighteen 3D landmarks were collected on 157 extant and fossil crania representing 90 genera. Data were subjected to a Generalized Procrustes Analysis then principal components analysis. Relative diversity between clades was assessed using an F-statistic. RESULTS: Fossil taxa do not significantly alter major axes of cranial shape, but they do occupy unique areas of morphospace, change the relative diversity between clades, and fill in notable gaps in primate cranial evolution. Strepsirrhines remain significantly less diverse than anthropoids. Fossil hominins fill the gap in cranial morphospace between extant great apes and modern humans. DISCUSSION: The morphospace outlined by living primates largely includes that occupied by fossil taxa, suggesting that the cranial diversity of living primates generally encompasses the total diversity that has evolved in this Order. The evolution of the anthropoid cranium was a significant event allowing anthropoids to achieve significantly greater cranial diversity compared to strepsirrhines. Fossil taxa fill in notable gaps within and between clades, highlighting their transitional nature and eliminating the appearance of large morphological distances between extant taxa, particularly in the case of extant hominids.

Resource seasonality and reproduction predict fission-fusion dynamics in black-and-white ruffed lemurs (Varecia variegata).

Ruffed lemurs (genus Varecia) are often described as having a flexible social organization, such that both cohesive (low fission-fusion dynamics) and fluid (high fission-fusion dynamics) grouping patterns have been observed. In ruffed lemur communities with high fission-fusion dynamics, group members vary in their temporal and spatial dispersion throughout a communally defended territory. These patterns have been likened to those observed in several haplorrhine species that exhibit the most fluid types of fission-fusion social organization (e.g., Pan and Ateles). To substantiate and further refine these claims, we describe the fission-fusion dynamics of a black-and-white ruffed lemur (Varecia variegata) community at Mangevo, an undisturbed primary rainforest site in Ranomafana National Park, Madagascar. We collected instantaneous group scan samples from August 2007-December 2008 (4,044 observation hours) to study and characterize patterns of subgroup size, composition, cohesion, and social association. In 16 consecutive months, we never found all members of the community together. In fact, individuals spent nearly half of their time alone. Subgroups were small, cohesive, and typically of mixed-sex composition. Mixed-sex subgroups were significantly larger, less cohesive, and more common than either male-only or female-only subgroups. Subgroup dynamics were related to shifts in climate, phenology of preferred fruit species, and female reproductive state. On average, association indices were low. Males and females were equally gregarious; however, adult male-male associations were significantly weaker than any other association type. Results presented herein document striking differences in fission-fusion dynamics between black-and-white ruffed lemurs and haplorrhines, while also demonstrating many broad-scale similarities to haplorrhine taxa that possess the most fluid fission-fusion societies.

Terrain Ruggedness and Canopy Height Predict Short-Range Dispersal in the Critically Endangered Black-and-White Ruffed Lemur.

Dispersal is a fundamental aspect of primates' lives and influences both population and community structuring, as well as species evolution. Primates disperse within an environmental context, where both local and intervening environmental factors affect all phases of dispersal. To date, research has primarily focused on how the intervening landscape influences primate dispersal, with few assessing the effects of local habitat characteristics. Here, we use a landscape genetics approach to examine between- and within-site environmental drivers of short-range black-and-white ruffed lemur (Varecia variegata) dispersal in the Ranomafana region of southeastern Madagascar. We identified the most influential drivers of short-range ruffed lemur dispersal as being between-site terrain ruggedness and canopy height, more so than any within-site habitat characteristic evaluated. Our results suggest that ruffed lemurs disperse through the least rugged terrain that enables them to remain within their preferred tall-canopied forest habitat. Furthermore, we noted a scale-dependent environmental effect when comparing our results to earlier landscape characteristics identified as driving long-range ruffed lemur dispersal. We found that forest structure drives short-range dispersal events, whereas forest presence facilitates long-range dispersal and multigenerational gene flow. Together, our findings highlight the importance of retaining high-quality forests and forest continuity to facilitate dispersal and maintain functional connectivity in ruffed lemurs.

Frugivory in four sympatric lemurs: implications for the future of Madagascar's forests.

Although some conservationists accept that not all species can be saved, we illustrate the difficulty in deciding which species are dispensable. In this article, we examine the possibility that the integrity of a forest relies on its entire faunal assemblage. In Madagascar, one faunal group, the lemurs, accounts for the greatest biomass and species richness among frugivores. For example, 7 of the 13 sympatric lemur species in Madagascar's eastern rainforests consume primarily fruit. Because of this, we suggest that some tree species may rely heavily on particular lemur taxa for both seed dispersal and germination. In Ranomafana National Park, the diets for four of the day-active lemur frugivores have been documented during annual cycles over a 5-year period. We predicted that, although the fruit of some plant taxa would be exploited by multiple lemur species, the fruit of others would be eaten by one lemur species alone. Analyses reveal that while lemurs overlap in a number of fruit taxa exploited, 46% (16/35) of families and 56% (29/52) of genera are eaten exclusively by one lemur species. We, therefore, predict local changes in forest composition and structure if certain of these lemur species are eliminated from a forest owing to hunting, disease, or habitat disturbance. We also suggest that this result may be of global significance because carbon sequestration by the tropical forests in Madagascar may be reduced as a result of this predicted change in forest composition.