Optimizing waterborne eDNA capture from waterholes in savanna systems under remote field conditions.

Environmental DNA (eDNA) is used for biodiversity assessments in a variety of ecosystems across the globe, whereby different eDNA concentration, preservation and extraction methods can outperform others depending on the sampling conditions and environment. Tropical and subtropical ecosystems in Africa are among the less studied systems concerning eDNA-based monitoring. Waterholes in arid parts of southern Africa represent important agglomeration points for terrestrial mammals, and the eDNA shed into such waterbodies provides a powerful source of information for monitoring mammalian biodiversity in the surrounding area. However, the applied methods for eDNA sampling, preservation and filtering in different freshwater systems vary greatly, and rigorous protocol testing in African freshwater systems is still lacking. This study represents the first attempt to examine variations in eDNA concentration, preservation and extraction methods under remote field conditions using waterborne eDNA in a savanna system. Collected samples were heavily affected by microalgal and bacterial growth, impeding eDNA capture and PCR success. We demonstrate clear effects of the methodological choices, which also depend on the state of eDNA. A preliminary metabarcoding run showed little taxonomic overlap in mammal species detection between two metabarcoding primers tested. We recommend water filtering (using filters with pore sizes >1 µm) over centrifugation for eDNA concentration, Longmire's solution for ambient temperature sample preservation and Qiagen's DNeasy PowerSoil Pro Kit for DNA extraction of these inhibitor-prone samples. Furthermore, at least two independent metabarcoding markers should be utilized in order to maximize species detections in metabarcoding studies.

Spatial density estimate of the snow leopard, Panthera uncia, in the Central Tibetan Plateau, China.

Knowledge of large carnivore population abundance is essential for wildlife management and conservation, but these data are often difficult to obtain in inherently low-density species. In particular, the snow leopard, Panthera uncia, an enigmatic cat occupying remote mountains in Central Asia, has received insufficient assessments of its population abundance because of logistical and methodological challenges. Here, we aimed to develop a robust density estimation of snow leopards based on 81 days of camera trapping within a contiguous and previously unsurveyed 1950 km2 area of habitat on the Tibetan Plateau (Mayue Township, Shenzha County, Tibet Autonomous Region, China). By applying spatially explicit capture-recapture models, we produced an estimate of 1.40 (95%CI: 1.06-1.84) individuals per 100 km2 . Results also suggested sex-specific variation in the range of movement around activity centers, with male (N = 10, σ = 4.02) movement considerably greater than female (N = 8, σ = 1.84) movement. The findings can serve as a reliable baseline reference for assessing the population trends of this endangered felid species with future estimates. This study will provide context to contribute toward a better understanding of ecological factors shaping the distribution and abundance of snow leopards and the related conservation measures needed to sustain their long-term survival on the Tibetan Plateau.

Ungulate spatiotemporal responses to contrasting predation risk from wolves and snow leopards.

Spatial responses to risk from multiple predators can precipitate emergent consequences for prey (i.e. multiple-predator effects, MPEs) and mediate indirect interactions between predators. How prey navigate risk from multiple predators may therefore have important ramifications for understanding the propagation of predation-risk effects (PREs) through ecosystems. The interaction of predator and prey traits has emerged as a potentially key driver of antipredator behaviour but remains underexplored in large vertebrate systems, particularly where sympatric prey share multiple predators. We sought to better generalize our understanding of how predators influence their ecosystems by considering how multiple sources of contingency drive prey distribution in a multi-predator-multi-prey system. Specifically, we explored how two sympatric ungulates with different escape tactics-vertically agile, scrambling ibex Capra sibirica and sprinting argali Ovis ammon-responded to predation risk from shared predators with contrasting hunting modes-cursorial wolves Canis lupus and vertical-ambushing, stalking snow leopards Panthera uncia. Contrasting risk posed by the two predators presented prey with clear trade-offs. Ibex selected for greater exposure to chronic long-term risk from snow leopards, and argali for wolves, in a nearly symmetrical manner that was predictable based on the compatibility of their respective traits. Yet, acute short-term risk from the same predator upended these long-term strategies, increasing each ungulates' exposure to risk from the alternate predator in a manner consistent with a scenario in which conflicting antipredator behaviours precipitate risk-enhancing MPEs and mediate predator facilitation. By contrast, reactive responses to wolves led ibex to reduce their exposure to risk from both predators-a risk-reducing MPE. Evidence of a similar reactive risk-reducing effect for argali vis-à-vis snow leopards was lacking. Our results suggest that prey spatial responses and any resulting MPEs and prey-mediated interactions between predators are contingent on the interplay of hunting mode and escape tactics. Further investigation of interactions among various drivers of contingency in PREs will contribute to a more comprehensive understanding and improved forecasting of the ecological effects of predators.

Guidelines for Telemetry Studies on Snow Leopards.

Animal-borne tracking devices have generated a wealth of new knowledge, allowing us to better understand, manage and conserve species. Fitting such tracking devices requires that animals are captured and often chemically immobilized. Such procedures cause stress and involve the risk of injuries and loss of life even in healthy individuals. For telemetry studies to be justifiable, it is vital that capture operations are planned and executed in an efficient and ethical way. Project objectives must be clearly articulated to address well-defined knowledge gaps, and studies designed to maximize the probability of achieving those goals. We provide guidelines for how to plan, design, and implement telemetry studies with a special emphasis on snow leopards that are typically captured using foot snares. We also describe the necessary steps to ensure that captures are conducted safely, and with minimal stress to animals.

Multiscale patterns of isolation by ecology and fine-scale population structure in Texas bobcats.

Patterns of spatial genetic variation can be generated by a variety of ecological processes, including individual preferences based on habitat. These ecological processes act at multiple spatial and temporal scales, generating scale-dependent effects on gene flow. In this study, we focused on bobcats (Lynx rufus), a highly mobile, generalist felid that exhibits ecological and behavioral plasticity, high abundance, and broad connectivity across much of their range. However, bobcats also show genetic differentiation along habitat breaks, a pattern typically observed in cases of isolation-by-ecology (IBE). The IBE observed in bobcats is hypothesized to occur due to habitat-biased dispersal, but it is unknown if this occurs at other habitat breaks across their range or at what spatial scale IBE becomes most apparent. Thus, we used a multiscale approach to examine isolation by ecology (IBE) patterns in bobcats (Lynx rufus) at both fine and broad spatial scales in western Texas. We genotyped 102 individuals at nine microsatellite loci and used partial redundancy analysis (pRDA) to test if a suite of landscape variables influenced genetic variation in bobcats. Bobcats exhibited a latitudinal cline in population structure with a spatial signature of male-biased dispersal, and no clear barriers to gene flow. Our pRDA tests revealed high genetic similarity in similar habitats, and results differed by spatial scale. At the fine spatial scale, herbaceous rangeland was an important influence on gene flow whereas mixed rangeland and agriculture were significant at the broad spatial scale. Taken together, our results suggests that complex interactions between spatial-use behavior and landscape heterogeneity can create non-random gene flow in highly mobile species like bobcats. Furthermore, our results add to the growing body of data highlighting the importance of multiscale study designs when assessing spatial genetic structure.

Genomics, environment and balancing selection in behaviourally bimodal populations: The caribou case.

Selection forces that favour different phenotypes in different environments can change frequencies of genes between populations along environmental clines. Clines are also compatible with balancing forces, such as negative frequency-dependent selection (NFDS), which maintains phenotypic polymorphisms within populations. For example, NFDS is hypothesized to maintain partial migration, a dimorphic behavioural trait prominent in species where only a fraction of the population seasonally migrates. Overall, NFDS is believed to be a common phenomenon in nature, yet a scarcity of studies were published linking naturally occurring allelic variation with bimodal or multimodal phenotypes and balancing selection. We applied a Pool-seq approach and detected selection on alleles associated with environmental variables along a North-South gradient in western North American caribou, a species displaying partially migratory behaviour. On 51 loci, we found a signature of balancing selection, which could be related to NFDS and ultimately the maintenance of the phenotypic polymorphisms known within these populations. Yet, remarkably, we detected directional selection on a locus when our sample was divided into two behaviourally distinctive groups regardless of geographic provenance (a subset of GPS-collared migratory or sedentary individuals), indicating that, within populations, phenotypically homogeneous groups were genetically distinctive. Loci under selection were linked to functional genes involved in oxidative stress response, body development and taste perception. Overall, results indicated genetic differentiation along an environmental gradient of caribou populations, which we found characterized by genes potentially undergoing balancing selection. We suggest that the underlining balancing force, NFDS, plays a strong role within populations harbouring multiple haplotypes and phenotypes, as it is the norm in animals, plants and humans too.

Environmental and anthropogenic drivers of connectivity patterns: A basis for prioritizing conservation efforts for threatened populations.

Ecosystem fragmentation and habitat loss have been the focus of landscape management due to restrictions on contemporary connectivity and dispersal of populations. Here, we used an individual approach to determine the drivers of genetic differentiation in caribou of the Canadian Rockies. We modelled the effects of isolation by distance, landscape resistance and predation risk and evaluated the consequences of individual migratory behaviour (seasonally migratory vs. sedentary) on gene flow in this threatened species. We applied distance-based and reciprocal causal modelling approaches, testing alternative hypotheses on the effects of geographic, topographic, environmental and local population-specific variables on genetic differentiation and relatedness among individuals. Overall, gene flow was restricted to neighbouring local populations, with spatial coordinates, local population size, groups and elevation explaining connectivity among individuals. Landscape resistance, geographic distances and predation risk were correlated with genetic distances, with correlations threefold higher for sedentary than for migratory caribou. As local caribou populations are increasingly isolated, our results indicate the need to address genetic connectivity, especially for populations with individuals displaying different migratory behaviours, whilst maintaining quality habitat both within and across the ranges of threatened populations.

Preferred habitat and effective population size drive landscape genetic patterns in an endangered species.

Landscape genetics provides a framework for pinpointing environmental features that determine the important exchange of migrants among populations. These studies usually test the significance of environmental variables on gene flow, yet ignore one fundamental driver of genetic variation in small populations, effective population size, N(e). W(e) combined both approaches in evaluating genetic connectivity of a threatened ungulate, woodland caribou. We used least-cost paths to calculate matrices of resistance distance for landscape variables (preferred habitat, anthropogenic features and predation risk) and population-pairwise harmonic means of N(e), and correlated them with genetic distances, FST and D(c). Results showed that spatial configuration of preferred habitat and Ne were the two best predictors of genetic relationships. Additionally, controlling for the effect of Ne increased the strength of correlations of environmental variables with genetic distance, highlighting the significant underlying effect of Ne in modulating genetic drift and perceived spatial connectivity. We therefore have provided empirical support to emphasize preventing increased habitat loss and promoting population growth to ensure metapopulation viability.

Reconstruction of caribou evolutionary history in Western North America and its implications for conservation.

The role of Beringia as a refugium and route for trans-continental exchange of fauna during glacial cycles of the past 2million years are well documented; less apparent is its contribution as a significant reservoir of genetic diversity. Using mitochondrial DNA sequences and 14 microsatellite loci, we investigate the phylogeographic history of caribou (Rangifer tarandus) in western North America. Patterns of genetic diversity reveal two distinct groups of caribou. Caribou classified as a Northern group, of Beringian origin, exhibited greater number and variability in mtDNA haplotypes compared to a Southern group originating from refugia south of glacial ice. Results indicate that subspecies R. t. granti of Alaska and R. t. groenlandicus of northern Canada do not constitute distinguishable units at mtDNA or microsatellites, belying their current status as separate subspecies. Additionally, the Northern Mountain ecotype of woodland caribou (presently R. t. caribou) has closer kinship to caribou classified as granti or groenlandicus. Comparisons of mtDNA and microsatellite data suggest that behavioural and ecological specialization is a more recently derived life history characteristic. Notably, microsatellite differentiation among Southern herds is significantly greater, most likely as a result of human-induced landscape fragmentation and genetic drift due to smaller population sizes. These results not only provide important insight into the evolutionary history of northern species such as caribou, but also are important indicators for managers evaluating conservation measures for this threatened species.

Effects of chronic wasting disease on reproduction and fawn harvest vulnerability in Wisconsin white-tailed deer.

Chronic wasting disease (CWD) is a fatal, transmissible spongiform encephalopathy that affects free-ranging and captive North American cervids. Although the impacts of CWD on cervid survival have been documented, little is known about the disease impacts on reproduction and recruitment. We used genetic methods and harvest data (2002-04) to reconstruct parentage for a cohort of white-tailed deer (Odocoileus virginianus) fawns born in spring 2002 and evaluate the effects of CWD infection on reproduction and fawn harvest vulnerability. There was no difference between CWD-positive and CWD-negative male deer in the probability of being a parent. However, CWD-positive females were more likely to be parents than CWD-negative females. Because our results are based on harvested animals, we evaluated the hypothesis that higher parentage rates occurred because fawns with CWD-positive mothers were more vulnerable to harvest. Male fawns with CWD-positive mothers were harvested earlier (>1 mo relative to their mother's date of harvest) and farther away from their mothers than male fawns with CWD-negative mothers. Male fawns with CWD-positive mothers were also harvested much earlier and farther away than female fawns from CWD-positive mothers. Most female fawns (86%) with CWD-positive mothers were harvested from the same section as their mothers, while almost half of male and female fawns with CWD-negative mothers were farther away. We conclude that preclinical stages of CWD infection do not prohibit white-tailed deer from successfully reproducing. However, apparently higher harvest vulnerability of male fawns with CWD-positive mothers suggests that CWD infection may make females less capable of providing adequate parental care to ensure the survival and recruitment of their fawns.

Going coastal: shared evolutionary history between coastal British Columbia and Southeast Alaska wolves (Canis lupus).

BACKGROUND: Many coastal species occupying the temperate rainforests of the Pacific Northwest in North America comprise endemic populations genetically and ecologically distinct from interior continental conspecifics. Morphological variation previously identified among wolf populations resulted in recognition of multiple subspecies of wolves in the Pacific Northwest. Recently, separate genetic studies have identified diverged populations of wolves in coastal British Columbia and coastal Southeast Alaska, providing support for hypotheses of distinct coastal subspecies. These two regions are geographically and ecologically contiguous, however, there is no comprehensive analysis across all wolf populations in this coastal rainforest. METHODOLOGY/PRINCIPAL FINDINGS: By combining mitochondrial DNA datasets from throughout the Pacific Northwest, we examined the genetic relationship between coastal British Columbia and Southeast Alaska wolf populations and compared them with adjacent continental populations. Phylogenetic analysis indicates complete overlap in the genetic diversity of coastal British Columbia and Southeast Alaska wolves, but these populations are distinct from interior continental wolves. Analyses of molecular variation support the separation of all coastal wolves in a group divergent from continental populations, as predicted based on hypothesized subspecies designations. Two novel haplotypes also were uncovered in a newly assayed continental population of interior Alaska wolves. CONCLUSIONS/SIGNIFICANCE: We found evidence that coastal wolves endemic to these temperate rainforests are diverged from neighbouring, interior continental wolves; a finding that necessitates new international strategies associated with the management of this species.

Survival in the Rockies of an endangered hybrid swarm from diverged caribou (Rangifer tarandus) lineages.

In North America, caribou (Rangifer tarandus) experienced diversification in separate refugia before the last glacial maximum. Geographical isolation produced the barren-ground caribou (Rangifer tarandus groenlandicus) with its distinctive migratory habits, and the woodland caribou (Rangifer tarandus caribou), which has sedentary behaviour and is now in danger of extinction. Herein we report on the phylogenetics, population structure, and migratory habits of caribou in the Canadian Rockies, utilizing molecular and spatial data for 223 individuals. Mitochondrial DNA analyses show the occurrence of two highly diverged lineages; the Beringian-Eurasian and North American lineages, while microsatellite data reveal that present-day Rockies' caribou populations have resulted from interbreeding between these diverged lineages. An ice-free corridor at the end of the last glaciation likely allowed, for the first time, for barren-ground caribou to migrate from the North and overlap with woodland caribou expanding from the South. The lack of correlation between nuclear and mitochondrial data may indicate that different environmental forces, which might also include human-caused habitat loss and fragmentation, are currently reshaping the population structure of this postglacial hybrid swarm. Furthermore, spatial ecological data show evidence of pronounced migratory behaviour within the study area, and suggest that the probability of being migratory may be higher in individual caribou carrying a Beringian-Eurasian haplotype which is mainly associated with the barren-ground subspecies. Overall, our analyses reveal an intriguing example of postglacial mixing of diverged lineages. In a landscape that is changing due to climatic and human-mediated factors, an understanding of these dynamics, both past and present, is essential for management and conservation of these populations.

Landscape genetics and the spatial distribution of chronic wasting disease.

Predicting the spread of wildlife disease is critical for identifying populations at risk, targeting surveillance and designing proactive management programmes. We used a landscape genetics approach to identify landscape features that influenced gene flow and the distribution of chronic wasting disease (CWD) in Wisconsin white-tailed deer. CWD prevalence was negatively correlated with genetic differentiation of study area deer from deer in the area of disease origin (core-area). Genetic differentiation was greatest, and CWD prevalence lowest, in areas separated from the core-area by the Wisconsin River, indicating that this river reduced deer gene flow and probably disease spread. Features of the landscape that influence host dispersal and spatial patterns of disease can be identified based on host spatial genetic structure. Landscape genetics may be used to predict high-risk populations based on their genetic connection to infected populations and to target disease surveillance, control and preventative activities.

A signal for independent coastal and continental histories among North American wolves.

Relatively little genetic variation has been uncovered in surveys across North American wolf populations. Pacific Northwest coastal wolves, in particular, have never been analysed. With an emphasis on coastal Alaska wolf populations, variation at 11 microsatellite loci was assessed. Coastal wolf populations were distinctive from continental wolves and high levels of diversity were found within this isolated and relatively small geographical region. Significant genetic structure within southeast Alaska relative to other populations in the Pacific Northwest, and lack of significant correlation between genetic and geographical distances suggest that differentiation of southeast Alaska wolves may be caused by barriers to gene flow, rather than isolation by distance. Morphological research also suggests that coastal wolves differ from continental populations. A series of studies of other mammals in the region also has uncovered distinctive evolutionary histories and high levels of endemism along the Pacific coast. Divergence of these coastal wolves is consistent with the unique phylogeographical history of the biota of this region and re-emphasizes the need for continued exploration of this biota to lay a framework for thoughtful management of southeast Alaska.