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Mesoamerican jaguars (Panthera onca) have been extirpated from over 77% of their historic range, inhabiting fragmented landscapes at potentially reduced population sizes. Maintaining and restoring genetic diversity and connectivity across human-altered landscapes has become a major conservation priority; nonetheless large-scale genetic monitoring of natural populations is rare. This is the first regional conservation genetic study of jaguars to primarily use fecal samples collected in the wild across five Mesoamerican countries: Belize, Costa Rica, Guatemala, Honduras, and Mexico. We genotyped 445 jaguar fecal samples and examined patterns of genetic diversity and connectivity among 115 individual jaguars using data from 12 microsatellite loci. Overall, moderate levels of genetic variation were detected (NA = 4.50 ± 1.05, AR = 3.43 ± 0.22, HE = 0.59 ± 0.04), with Mexico having the lowest genetic diversity, followed by Honduras, Guatemala, Belize, and Costa Rica. Population-based gene flow measures (FST = 0.09 to 0.15, Dest = 0.09 to 0.21), principal component analysis, and Bayesian clustering applied in a hierarchical framework revealed significant genetic structure in Mesoamerican jaguars, roughly grouping individuals into four genetic clusters with varying levels of admixture. Gene flow was highest among Selva Maya jaguars (northern Guatemala and central Belize), whereas genetic differentiation among all other sampling sites was moderate. Genetic subdivision was most pronounced between Selva Maya and Honduran jaguars, suggesting limited jaguar movement between these close geographic regions and ultimately refuting the hypothesis of contemporary panmixia. To maintain a critical linkage for jaguars dispersing through the Mesoamerican landscape and ensure long-term viability of this near threatened species, we recommend continued management and maintenance of jaguar corridors. The baseline genetic data provided by this study underscores the importance of understanding levels of genetic diversity and connectivity to making informed management and conservation decisions with the goal to maintain functional connectivity across the region.

Bat communities in the Neotropics are some of the most speciose assemblages of mammals on Earth, with regions supporting more than 100 sympatric species with diverse feeding ecologies. Because bats are small, nocturnal, and volant, it is difficult to directly observe their feeding habits, which has resulted in their classification into broadly defined dietary guilds (e.g., insectivores, carnivores, and frugivores). Apart from these broad guilds, we lack detailed dietary information for many species and therefore have only a limited understanding of interaction networks linking bats and their diet items. In this study, we used DNA metabarcoding of plants, arthropods, and vertebrates to investigate the diets of 25 bat species from the tropical dry forests of Lamanai, Belize. Our results report some of the first detection of diet items for the focal bat taxa, adding rich and novel natural history information to the field of bat ecology. This study represents a comprehensive first effort to apply DNA metabarcoding to bat diets at Lamanai and provides a useful methodological framework for future studies testing hypotheses about coexistence and niche differentiation in the context of modern high‐throughput molecular data. Inferring the exact diets of small, nocturnal mammals such as bats is notoriously difficult. Limited knowledge of what bats actually eat leaves important trophic interactions and food web networks unclear. Using molecular barcoding of plants, vertebrates, and invertebrates, we report fine‐scale dietary detection for 25 species of Belizean bats.

With increasing anthropogenic impact and landscape change, terrestrial carnivore populations are becoming more fragmented. Thus, it is crucial to genetically monitor wild carnivores and quantify changes in genetic diversity and gene flow in response to these threats. This study combined the use of scat detector dogs and molecular scatology to conduct the first genetic study on wild populations of multiple Neotropical felids coexisting across a fragmented landscape in Belize, Central America. We analyzed data from 14 polymorphic microsatellite loci in 1053 scat samples collected from wild jaguars (Panthera onca), pumas (Puma concolor), and ocelots (Leopardus pardalis). We assessed levels of genetic diversity, defined potential genetic clusters, and examined gene flow for the three target species on a countrywide scale using a combination of individual- and population-based analyses. Wild felids in Belize showed moderate levels of genetic variation, with jaguars having the lowest diversity estimates (HE = 0.57 ± 0.02; AR = 3.36 ± 0.09), followed by pumas (HE = 0.57 ± 0.08; AR = 4.20 ± 0.16), and ocelots (HE = 0.63 ± 0.03; AR = 4.16 ± 0.08). We observed low to moderate levels of genetic differentiation for all three target species, with jaguars showing the lowest degree of genetic subdivision across the country, followed by ocelots and pumas. Although levels of genetic diversity and gene flow were still fairly high, we detected evidence of fine-scale genetic subdivision, indicating that levels of genetic connectivity for wild felids in Belize are likely to decrease if habitat loss and fragmentation continue at the current rate. Our study demonstrates the value of understanding fine-scale patterns of gene flow in multiple co-occurring felid species of conservation concern, which is vital for wildlife movement corridor planning and prioritizing future conservation and management efforts within human-impacted landscapes.

Background Animals evolved in a microbial world, and their gut microbial symbionts have played a role in their ecological diversification. While many recent studies report patterns of phylosymbiosis between hosts and their gut bacteria, fewer studies examine the potentially adaptive functional contributions of these microbes to the dietary habits of their hosts. In this study, we examined predicted metabolic pathways in the gut bacteria of more than 500 individual bats belonging to 60 species and compare the enrichment of these functions across hosts with distinct dietary ecologies. Results We found that predicted microbiome functions were differentially enriched across hosts with different diets. Using a machine-learning approach, we also found that inferred microbiome functions could be used to predict specialized host diets with reasonable accuracy. We detected a relationship between both host phylogeny and diet with respect to microbiome functional repertoires. Because many predicted functions could potentially fill nutritional gaps for bats with specialized diets, we considered pathways discriminating dietary niches as traits of the host and fit them to comparative phylogenetic models of evolution. Our results suggest that some, but not all, predicted microbiome functions may evolve toward adaptive optima and thus be visible to the forces of natural selection operating on hosts over evolutionary time. Conclusions Our results suggest that bats with specialized diets may partially rely on their gut microbes to fulfill or augment critical nutritional pathways, including essential amino acid synthesis, fatty acid biosynthesis, and the generation of cofactors and vitamins essential for proper nutrition. Our work adds to a growing body of literature suggesting that animal microbiomes are structured by a combination of ecological and evolutionary processes and sets the stage for future metagenomic and metabolic characterization of the bat microbiome to explore links between bacterial metabolism and host nutrition.

With fewer than 200 tigers (Panthera tigris tigris) left in Nepal, that are generally confined to five protected areas across the Terai Arc Landscape, genetic studies are needed to provide crucial information on diversity and connectivity for devising an effective country-wide tiger conservation strategy. As part of the Nepal Tiger Genome Project, we studied landscape change, genetic variation, population structure, and gene flow of tigers across the Terai Arc Landscape by conducting Nepal's first comprehensive and systematic scat-based, non-invasive genetic survey. Of the 770 scat samples collected opportunistically from five protected areas and six presumed corridors, 412 were tiger (57%). Out of ten microsatellite loci, we retain eight markers that were used in identifying 78 individual tigers. We used this dataset to examine population structure, genetic variation, contemporary gene flow, and potential population bottlenecks of tigers in Nepal. We detected three genetic clusters consistent with three demographic sub-populations and found moderate levels of genetic variation (He = 0.61, AR = 3.51) and genetic differentiation (FST = 0.14) across the landscape. We detected 3-7 migrants, confirming the potential for dispersal-mediated gene flow across the landscape. We found evidence of a bottleneck signature likely caused by large-scale land-use change documented in the last two centuries in the Terai forest. Securing tiger habitat including functional forest corridors is essential to enhance gene flow across the landscape and ensure long-term tiger survival. This requires cooperation among multiple stakeholders and careful conservation planning to prevent detrimental effects of anthropogenic activities on tigers.

Carnivores are currently colonizing cities where they were previously absent. These urban environments are novel ecosystems characterized by habitat degradation and fragmentation, availability of human food, and different prey assemblages than surrounding areas. Coyotes ( Canis latrans ) established a breeding population in New York City (NYC) over the last few decades, but their ecology within NYC is poorly understood. In this study, we used non-invasive scat sampling and DNA metabarcoding to profile vertebrate, invertebrate, and plant dietary items with the goal to compare the diets of urban coyotes to those inhabiting non-urban areas. We found that both urban and non-urban coyotes consumed a variety of plants and animals as well as human food. Raccoons ( Procyon lotor ) were an important food item for coyotes within and outside NYC. In contrast, white-tailed deer ( Odocoileus virginianus ) were mainly eaten by coyotes inhabiting non-urban areas. Domestic chicken ( Gallus gallus ) was the human food item found in most scats from both urban and non-urban coyotes. Domestic cats ( Felis catus ) were consumed by urban coyotes but were detected in only a small proportion of the scats (<5%), which differs markedly from high rates of cat depredation in some other cities. In addition, we compared our genetic metabarcoding analysis to a morphological analysis of the same scat samples. We found that the detection similarity between the two methods was low and it varied depending on the type of diet item.

Bengal tigers (Panthera tigris tigris) serve a pivotal role as an apex predator in forest ecosystems. To increase our knowledge on factors impacting the viability and health of this endangered species, we studied the gut microbiota in 32 individual Bengal tigers from three geographically separated areas (Chitwan National Park (CNP), Bardia National Park (BNP) and Suklaphanta Wildlife Reserve (SWR)) in Nepal, using noninvasive genetic sampling methods. Gut microbiota influence the immune system, impact various physiological functions, and modulates metabolic reactions, that ultimately impact the host health, behavior and development. Across the tiger populations in Nepal, we found significant differences in the composition of microbial communities based on their geographic locations. Specifically, we detected significant differences between CNP and the other two protected areas (CNP vs BNP: pseudo t = 1.944, P = 0.006; CNP vs SWR: pseudo t = 1.9942, P = 0.0071), but no differences between BNP and SWR. This mirrors what has been found for tiger gene flow in the same populations, suggesting gut microbiota composition and host gene flow may be linked. Furthermore, predictive metagenome functional content analysis (PICRUSt) revealed a higher functional enrichment and diversity for significant gut microbiota in the Chitwan tiger population and the lowest enrichment and diversity in Suklaphanta. The CNP tiger population contained higher proportions of microbiota that are associated with predicted functions relevant for metabolism of amino acid, lipid, xenobiotics biodegradation, terpenoides and polyketides than the SWR population. We conclude the tiger population structure, gut microbiota profile and associated functional metabolic categories are correlated, with geographically most separated CNP and SWR tiger population having the most distinct and different host genotype and microbiota profiles. Our work dramatically expands the understanding of tiger microbiota in wild populations and provides a valuable case study on how to investigate genetic diversity at different hierarchical levels, including hosts as well as their microbial communities.

Field-sampling methods for molecular scatology studies must be optimized, especially when working on elusive species in challenging tropical environments where rates of DNA degradation are elevated because of hot and humid weather conditions. To maximize polymerase chain reaction (PCR) amplification success and genotyping accuracy rates and to minimize genotyping error rates for fecal DNA samples of jaguars (Panthera onca) and co-occurring Neotropical felids collected in Belize, Central America, we evaluated the performance of two fecal DNA storage techniques (dimethyl sulfoxide saline solution [DETs buffer] and 95% ethanol [EtOH]) suitable for long-term preservation at remote tropical sites. Additionally, we tested fecal samples collected from 4 different locations on the scat (top, side, bottom, inside) at 2 different tropical forest types (tropical broadleaf and tropical pine forests). DETs buffer was the superior fecal DNA preservation method, with 44% higher PCR amplification success (P= 0.009) and 17% higher genotyping accuracy (P= 0.021) than 95% EtOH-stored samples. Polymerase chain reaction amplification success of fecal DNA collected at the more open, pine-forest (Pinussp.) site differed significantly across locations on the scat, with highest mean success rates obtained from the top (85% ± 6.5%SD), followed by the side (79% ± 9.4%SD), bottom (76% ± 11.9%SD), and inside (69% ± 10.3% SD) of scat samples. Scat samples collected at the more closed-canopy broadleaf site did not show any significant differences in amplification success rates across scat locations. We recommend that researchers optimize field-sampling methods, including collection and storage protocols, by conducting a pilot study prior to their molecular scatology research efforts.

Large‐scale anthropogenic developments in the metropolitan areas of Nepal and the rural to urban influx of people have exacerbated human–wildlife conflicts across human‐altered landscapes of Nepal. The Kathmandu Valley has experienced large‐scale urbanization and has subsequently witnessed substantial incidents of human–wildlife conflicts given the increasing levels of human encroachment into remnant wildlife habitats. Here, we applied DNA metabarcoding in combination with geospatial analysis to study the feeding ecology of two urban carnivores, the leopard (Panthera pardus) and the leopard cat (Prionailurus bengalensis), in the forests surrounding the Kathmandu Valley and to check whether the leopards' predation on domestic animals contributes to human‐leopard conflict in this region and to obtain a baseline data on the dietary habits of the poorly studied leopard cat. We found that leopards were highly dependent on domestic animals in areas dominated by human‐use activities (agricultural and built‐up areas), whereas leopard cats mostly predated on wild rodents. Through our work, we highlight the importance of domestic prey in the diets of urban carnivores like leopards and demonstrate the influence human‐induced habitat disturbance has on the ecology of local wildlife. This study generates critical information which will help to inform conflict mitigation strategies and conservation planning for the two carnivore species, in addition to identifying areas within the region that are susceptible to human–wildlife conflicts. Large‐scale urbanization and the rural‐to‐urban influx in Nepal have increased human–wildlife conflicts, particularly in the Kathmandu Valley. Using DNA metabarcoding and geospatial analysis, we studied the feeding ecology of leopards and leopard cats, finding that leopards rely heavily on domestic animals in human‐dominated areas, whereas leopard cats primarily prey on wild rodents. This study highlights the impact of human‐induced habitat disturbance on urban carnivores and provides critical information for conflict mitigation and conservation planning.

Little is known about the relatedness structure of carnivores living in urban areas, where green spaces may vary in size and resource availability. We examined the minimum population size, relatedness structure, and genetic diversity of a recently established population of eastern coyotes (Canis latrans) inhabiting New York City (NYC). The population has been established for approximately 25 years, and sample collection for genetic analysis has been ongoing since 2010. We genotyped 234 scat, eight tissue, and three blood samples at nine microsatellite loci. We identified 45 individual coyotes with a male-biased sex ratio of 2.2:1. We also found moderate to high levels of genetic diversity, with average observed heterozygosity of 0.779 and mean number of alleles per locus of 7.8. Most of the green spaces surveyed supported a single group of closely related coyotes in each. Relatedness comparisons between parks also indicated that coyotes compared across different parks were also closely related. We identified two unrelated mated pairs and found no support for polygamy. The high incidence of relatedness suggests that the coyote population is descended from a small number of founding individuals. Additionally, we genetically recaptured several coyotes, including one individual sampled in the Bronx and in Queens, with a median of 103 days between resampling. This result indicates that the coyotes are persisting in some of the isolated greenspaces of New York City and able to move successfully between them.