Bighorn Sheep (Ovis canadensis) with Higher Whole Blood Selenium Levels Have Improved Survival and Altered Immune Responses.

Bighorn sheep (Ovis canadensis) are herbivorous ungulates that live in forage-poor areas of the American west. The trace minerals that herbivores derive from forage are important for immune function. Therefore, identifying trace minerals that affect immune function in bighorn sheep could provide important insights into disease susceptibility and population health in threatened populations. We sought to determine whether trace mineral composition in blood or plasma correlates to survival and determine whether immunologic parameters correlate with any trace minerals that affect survival. We used data collected from 2016 to 2018 as part of a large study on bighorn sheep in southeastern Oregon and northern Nevada, US. We measured the survival of 135 bighorn sheep during the 8-mo monitoring period, including general metrics of immune function and trace mineral levels. We found that animals with higher selenium had improved survival over the monitoring period, with higher peripheral blood mononuclear cell activity (lymphocytes and monocytes) and lower bacterial killing ability in an in vitro assay. This suggests that bighorn sheep may have altered immune function when selenium levels are low, making them more likely to die during the 8-mo monitoring period. Future work should consider whether habitat management strategies that increase selenium intake might improve disease resistance and survival in bighorn sheep in selenium-poor areas.

Animal movement and associated infectious disease risk in a metapopulation.

Animal movements among habitat patches or populations are important for maintaining long-term genetic and demographic viability, but connectivity may also facilitate disease spread and persistence. Understanding factors that influence animal movements is critical to understanding potential transmission risk and persistence of communicable disease in spatially structured systems. We evaluated effects of sex, age and Mycoplasma ovipneumoniae infection status at capture on intermountain movements and seasonal movement rates observed in desert bighorn sheep (Ovis canadensis nelsoni) using global positioning system collar data from 135 individuals (27 males, 108 females) in 14 populations between 2013 and 2018, following a pneumonia outbreak linked to the pathogen M. ovipneumoniae in the Mojave Desert, California, USA. Based on logistic regression analysis, intermountain movements were influenced by sex, age and most notably, infection status at capture: males, older animals and uninfected individuals were most likely to make such movements. Based on multiple linear regression analysis, females that tested positive for M. ovipneumoniae at capture also had lower mean daily movement rates that were further influenced by season. Our study provides empirical evidence of a pathogenic infection decreasing an individual's future mobility, presumably limiting that pathogen's ability to spread, and ultimately influencing transmission risk within a spatially structured system.

Population connectivity patterns of genetic diversity, immune responses and exposure to infectious pneumonia in a metapopulation of desert bighorn sheep.

Habitat fragmentation is an important driver of biodiversity loss and can be remediated through management actions aimed at maintenance of natural connectivity in metapopulations. Connectivity may protect populations from infectious diseases by preserving immunogenetic diversity and disease resistance. However, connectivity could exacerbate the risk of infectious disease spread across vulnerable populations. We tracked the spread of a novel strain of Mycoplasma ovipneumoniae in a metapopulation of desert bighorn sheep Ovis canadensis nelsoni in the Mojave Desert to investigate how variation in connectivity among populations influenced disease outcomes. M. ovipneumoniae was detected throughout the metapopulation, indicating that the relative isolation of many of these populations did not protect them from pathogen invasion. However, we show that connectivity among bighorn sheep populations was correlated with higher immunogenetic diversity, a protective immune response and lower disease prevalence. Variation in protective immunity predicted infection risk in individual bighorn sheep and was associated with heterozygosity at genetic loci linked to adaptive and innate immune signalling. Together, these findings may indicate that population connectivity maintains immunogenetic diversity in bighorn sheep populations in this system and has direct effects on immune responses in individual bighorn sheep and their susceptibility to infection by a deadly pathogen. Our study suggests that the genetic benefits of population connectivity could outweigh the risk of infectious disease spread and supports conservation management that maintains natural connectivity in metapopulations.

Dating of a large tool assemblage at the Cooper's Ferry site (Idaho, USA) to ~15,785 cal yr B.P. extends the age of stemmed points in the Americas.

The timing and character of the Pleistocene peopling of the Americas are measured by the discovery of unequivocal artifacts from well-dated contexts. We report the discovery of a well-dated artifact assemblage containing 14 stemmed projectile points from the Cooper's Ferry site in western North America, dating to ~16,000 years ago. These stemmed points are several thousand years older than Clovis fluted points (~13,000 cal yr B.P.) and are ~2300 years older than stemmed points found previously at the site. These points date to the end of Marine Isotope Stage 2 when glaciers had closed off an interior land route into the Americas. This assemblage includes an array of stemmed projectile points that resemble pre-Jomon Late Upper Paleolithic tools from the northwestern Pacific Rim dating to ~20,000 to 19,000 years ago, leading us to hypothesize that some of the first technological traditions in the Americas may have originated in the region.

Emigration effects on estimates of age- and sex-specific survival of two sciurids.

Age- and sex-specific survival estimates are crucial to understanding important life history characteristics, and variation in these estimates can be a key driver of population dynamics. When estimating survival using Cormack-Jolly-Seber (CJS) models, emigration is typically unknown but confounded with apparent survival. Consequently, especially for populations or age classes with high dispersal rates, apparent survival estimates are often biased low and temporal patterns in survival might be masked when site fidelity varies temporally. We used 9 years of annual mark-recapture data to estimate age-, sex-, and time-specific apparent survival of Humboldt's flying squirrels (Glaucomys oregonensis) and Townsend's chipmunks (Neotamias townsendii). For Humboldt's flying squirrels, these estimates support a small body of research investigating potential variation in survival among age and sex classes, but age- and sex-specific survival has not been evaluated for Townsend's chipmunks. We also quantified the effects of age- and sex-specific emigration on confounded estimates of apparent survival. Our estimates of juvenile flying squirrel survival were high relative to other small mammal species and estimates for both species were variable among years. We found survival differed moderately among age and sex classes for Humboldt's flying squirrels, but little among age and sex classes for Townsend's chipmunks, and that the degree to which emigration confounded apparent survival estimates varied substantially among years. Our results demonstrate that emigration can influence commonly used estimates of apparent survival. Unadjusted estimates confounded the interpretation of differences in survival between age and sex classes and masked potential temporal patterns in survival because the magnitude of adjustment varied among years. We conclude that apparent survival estimators are robust during some time periods; however, when emigration rates vary in time, the effects of emigration should be carefully considered and accounted for.

Host, Microbiome, and Complex Space: Applying Population and Landscape Genetic Approaches to Gut Microbiome Research in Wild Populations.

In recent years, emerging sequencing technologies and computational tools have driven a tidal wave of research on host-associated microbiomes, particularly the gut microbiome. These studies demonstrate numerous connections between the gut microbiome and vital host functions, primarily in humans, model organisms, and domestic animals. As the adaptive importance of the gut microbiome becomes clearer, interest in studying the gut microbiomes of wild populations has increased, in part due to the potential for discovering conservation applications. The study of wildlife gut microbiomes holds many new challenges and opportunities due to the complex genetic, spatial, and environmental structure of wild host populations, and the potential for these factors to interact with the microbiome. The emerging picture of adaptive coevolution in host-microbiome relationships highlights the importance of understanding microbiome variation in the context of host population genetics and landscape heterogeneity across a wide range of host populations. We propose a conceptual framework for understanding wildlife gut microbiomes in relation to landscape variables and host population genetics, including the potential of approaches derived from landscape genetics. We use this framework to review current research, synthesize important trends, highlight implications for conservation, and recommend future directions for research. Specifically, we focus on how spatial structure and environmental variation interact with host population genetics and microbiome variation in natural populations, and what we can learn from how these patterns of covariation differ depending on host ecological and evolutionary traits.

Sex chromosome transformation and the origin of a male-specific X chromosome in the creeping vole.

The mammalian sex chromosome system (XX female/XY male) is ancient and highly conserved. The sex chromosome karyotype of the creeping vole (Microtus oregoni) represents a long-standing anomaly, with an X chromosome that is unpaired in females (X0) and exclusively maternally transmitted. We produced a highly contiguous male genome assembly, together with short-read genomes and transcriptomes for both sexes. We show that M. oregoni has lost an independently segregating Y chromosome and that the male-specific sex chromosome is a second X chromosome that is largely homologous to the maternally transmitted X. Both maternally inherited and male-specific sex chromosomes carry fragments of the ancestral Y chromosome. Consequences of this recently transformed sex chromosome system include Y-like degeneration and gene amplification on the male-specific X, expression of ancestral Y-linked genes in females, and X inactivation of the male-specific chromosome in male somatic cells. The genome of M. oregoni elucidates the processes that shape the gene content and dosage of mammalian sex chromosomes and exemplifies a rare case of plasticity in an ancient sex chromosome system.

Previously Unrecognized Exposure of Desert Bighorn Sheep (Ovis canadensis nelsoni) to Mycoplasma ovipneumoniae in the California Mojave Desert.

A 2013 outbreak of respiratory disease in bighorn sheep from California's Mojave Desert metapopulation caused high mortality in at least one population. Subsequent PCR and strain-typing indicate widespread infection of a single strain of Mycoplasma ovipneumoniae throughout this region. Serosurvey of archived samples showed that some populations have had antibodies to M. ovipneumoniae since at least 1986, although pre-2013 strain-type data are unavailable.

Impact of Mycoplasma ovipneumoniae on juvenile bighorn sheep (Ovis canadensis) survival in the northern Basin and Range ecosystem.

Determining the demographic impacts of wildlife disease is complex because extrinsic and intrinsic drivers of survival, reproduction, body condition, and other factors that may interact with disease vary widely. Mycoplasma ovipneumoniae infection has been linked to persistent mortality in juvenile bighorn sheep (Ovis canadensis), although mortality appears to vary widely across subspecies, populations, and outbreaks. Hypotheses for that variation range from interactions with nutrition, population density, genetic variation in the pathogen, genetic variation in the host, and other factors. We investigated factors related to survival of juvenile bighorn sheep in reestablished populations in the northern Basin and Range ecosystem, managed as the formerly-recognized California subspecies (hereafter, "California lineage"). We investigated whether survival probability of 4-month juveniles would vary by (1) presence of M. ovipneumoniae-infected or exposed individuals in populations, (2) population genetic diversity, and (3) an index of forage suitability. We monitored 121 juveniles across a 3-year period in 13 populations in southeastern Oregon and northern Nevada. We observed each juvenile and GPS-collared mother semi-monthly and established 4-month capture histories for the juvenile to estimate survival. All collared adult females were PCR-tested at least once for M. ovipneumoniae infection. The presence of M. ovipneumoniae-infected juveniles was determined by observing juvenile behavior and PCR-testing dead juveniles. We used a known-fate model with different time effects to determine if the probability of survival to 4 months varied temporally or was influenced by disease or other factors. We detected dead juveniles infected with M. ovipneumoniae in only two populations. Derived juvenile survival probability at four months in populations where infected juveniles were not detected was more than 20 times higher. Detection of infected adults or adults with antibody levels suggesting prior exposure was less predictive of juvenile survival. Survival varied temporally but was not strongly influenced by population genetic diversity or nutrition, although genetic diversity within most study area populations was very low. We conclude that the presence of M. ovipneumoniae can cause extremely low juvenile survival probability in translocated bighorn populations of the California lineage, but found little influence that genetic diversity or nutrition affect juvenile survival. Yet, after the PCR+ adult female in one population died, subsequent observations found 11 of 14 ( 79%) collared adult females had surviving juveniles at 4-months, suggesting that targeted removals of infected adults should be evaluated as a management strategy.

Bighorn sheep gut microbiomes associate with genetic and spatial structure across a metapopulation.

Studies in laboratory animals demonstrate important relationships between environment, host traits, and microbiome composition. However, host-microbiome relationships in natural systems are understudied. Here, we investigate metapopulation-scale microbiome variation in a wild mammalian host, the desert bighorn sheep (Ovis canadensis nelsoni). We sought to identify over-represented microbial clades and understand how landscape variables and host traits influence microbiome composition across the host metapopulation. To address these questions, we performed 16S sequencing on fecal DNA samples from thirty-nine bighorn sheep across seven loosely connected populations in the Mojave Desert and assessed relationships between microbiome composition, environmental variation, geographic distribution, and microsatellite-derived host population structure and heterozygosity. We first used a phylogenetically-informed algorithm to identify bacterial clades conserved across the metapopulation. Members of genus Ruminococcaceae, genus Lachnospiraceae, and family Christensenellaceae R7 group were among the clades over-represented across the metapopulation, consistent with their known roles as rumen symbionts in domestic livestock. Additionally, compositional variation among hosts correlated with individual-level geographic and genetic structure, and with population-level differences in genetic heterozygosity. This study identifies microbiome community variation across a mammalian metapopulation, potentially associated with genetic and geographic population structure. Our results imply that microbiome composition may diverge in accordance with landscape-scale environmental and host population characteristics.

Pan-tissue transcriptome analysis of long noncoding RNAs in the American beaver Castor canadensis.

BACKGROUND: Long noncoding RNAs (lncRNAs) have roles in gene regulation, epigenetics, and molecular scaffolding and it is hypothesized that they underlie some mammalian evolutionary adaptations. However, for many mammalian species, the absence of a genome assembly precludes the comprehensive identification of lncRNAs. The genome of the American beaver (Castor canadensis) has recently been sequenced, setting the stage for the systematic identification of beaver lncRNAs and the characterization of their expression in various tissues. The objective of this study was to discover and profile polyadenylated lncRNAs in the beaver using high-throughput short-read sequencing of RNA from sixteen beaver tissues and to annotate the resulting lncRNAs based on their potential for orthology with known lncRNAs in other species. RESULTS: Using de novo transcriptome assembly, we found 9528 potential lncRNA contigs and 187 high-confidence lncRNA contigs. Of the high-confidence lncRNA contigs, 147 have no known orthologs (and thus are putative novel lncRNAs) and 40 have mammalian orthologs. The novel lncRNAs mapped to the Oregon State University (OSU) reference beaver genome with greater than 90% sequence identity. While the novel lncRNAs were on average shorter than their annotated counterparts, they were similar to the annotated lncRNAs in terms of the relationships between contig length and minimum free energy (MFE) and between coverage and contig length. We identified beaver orthologs of known lncRNAs such as XIST, MEG3, TINCR, and NIPBL-DT. We profiled the expression of the 187 high-confidence lncRNAs across 16 beaver tissues (whole blood, brain, lung, liver, heart, stomach, intestine, skeletal muscle, kidney, spleen, ovary, placenta, castor gland, tail, toe-webbing, and tongue) and identified both tissue-specific and ubiquitous lncRNAs. CONCLUSIONS: To our knowledge this is the first report of systematic identification of lncRNAs and their expression atlas in beaver. LncRNAs-both novel and those with known orthologs-are expressed in each of the beaver tissues that we analyzed. For some beaver lncRNAs with known orthologs, the tissue-specific expression patterns were phylogenetically conserved. The lncRNA sequence data files and raw sequence files are available via the web supplement and the NCBI Sequence Read Archive, respectively.

Effects of trapping effort and trap placement on estimating abundance of Humboldt's flying squirrels.

Live trapping is a common tool used to assess demography of small mammals. However, live-trapping is often expensive and stressful to captured individuals. Thus, assessing the relative tradeoffs among study goals, project expenses, and animal well-being is necessary. Here, we evaluated how apparent bias and precision of estimates for apparent annual survival, abundance, capture probability, and recapture probability of Humboldt's flying squirrels (Glaucomys oregonensis) varied with the number of secondary trapping occasions. We used data from forested sites trapped on 12 consecutive occasions annually in the HJ Andrews Experimental Forest (9 sites, 6 years) and the Siuslaw National Forest (seven sites, three years) in Oregon. We used Huggins robust design models to estimate parameters of interest for the first 4, 8, and 12 trapping occasions. We also estimated the effect of attaching Tomahawk traps to tree boles on site- and year-specific flying squirrel capture frequencies. Our estimates with 12 occasions were similar to those from previous studies. Abundances and capture probabilities were variable among years on both sites; however, variation was much lower on the Siuslaw sites. Reducing the length of primary trapping occasions from 12 to 8 nights had very little impact on parameter estimates, but further reducing the length of primary trapping occasions to four nights caused substantial apparent bias in parameter estimates and decreased precision. We found that attaching Tomahawk traps to tree boles increased the site- and year-specific capture frequency of flying squirrels. Our results suggest that live-trapping studies targeting Humboldt's flying squirrels in the Pacific Northwest of the United States could reduce per-site costs and stress to captured individuals without biasing estimates by reducing the length of primary trapping occasions to 8 nights. We encourage similar analyses for other commonly-trapped species in these and other ecosystems.

Late Upper Paleolithic occupation at Cooper's Ferry, Idaho, USA, ~16,000 years ago.

Radiocarbon dating of the earliest occupational phases at the Cooper's Ferry site in western Idaho indicates that people repeatedly occupied the Columbia River basin, starting between 16,560 and 15,280 calibrated years before the present (cal yr B.P.). Artifacts from these early occupations indicate the use of unfluted stemmed projectile point technologies before the appearance of the Clovis Paleoindian tradition and support early cultural connections with northeastern Asian Upper Paleolithic archaeological traditions. The Cooper's Ferry site was initially occupied during a time that predates the opening of an ice-free corridor (≤14,800 cal yr B.P.), which supports the hypothesis that initial human migration into the Americas occurred via a Pacific coastal route.

Why did the buffalo cross the park? Resource shortages, but not infections, drive dispersal in female African buffalo (Syncerus caffer).

Dispersal facilitates population health and maintains resilience in species via gene flow. Adult dispersal occurs in some species, is often facultative, and is poorly understood, but has important management implications, particularly with respect to disease spread. Although the role of adult dispersal in spreading disease has been documented, the potential influence of disease on dispersal has received little attention. African buffalo (Syncerus caffer) are wide-ranging and harbor many pathogens that can affect nearby livestock. Dispersal of adult buffalo has been described, but ecological and social drivers of buffalo dispersal are poorly understood. We investigated drivers of adult buffalo dispersal during a 4-year longitudinal study at Kruger National Park, South Africa. We monitored the spatial movement of 304 female buffalo in two focal areas using satellite and radio collars, capturing each buffalo every 6 months to assess animal traits and disease status. We used generalized linear mixed models to determine whether likelihood of dispersal for individual female buffalo was influenced by animal traits, herd identity, environmental variables, gastrointestinal parasites, or microparasite infections. The likelihood and drivers of buffalo dispersal varied by herd, area, and year. In the Lower Sabie herd, where resources were abundant, younger individuals were more likely to disperse, with most dispersal occurring in the early wet season and during an unusually dry year, 2009. In the resource-poor Crocodile Bridge area, buffalo in poor condition were most likely to disperse. Our findings suggest that dispersal of female buffalo is driven by either seasonal (Lower Sabie) or perhaps social (Crocodile Bridge) resource restriction, indicating resource limitation and dispersal decisions are tightly linked for this social ungulate. We found no direct effects of infections on buffalo dispersal, assuaging fears that highly infectious individuals might be more prone to dispersing, which could accelerate the spatial spread of infectious diseases.

Large-scale ruminant genome sequencing provides insights into their evolution and distinct traits.

The ruminants are one of the most successful mammalian lineages, exhibiting morphological and habitat diversity and containing several key livestock species. To better understand their evolution, we generated and analyzed de novo assembled genomes of 44 ruminant species, representing all six Ruminantia families. We used these genomes to create a time-calibrated phylogeny to resolve topological controversies, overcoming the challenges of incomplete lineage sorting. Population dynamic analyses show that population declines commenced between 100,000 and 50,000 years ago, which is concomitant with expansion in human populations. We also reveal genes and regulatory elements that possibly contribute to the evolution of the digestive system, cranial appendages, immune system, metabolism, body size, cursorial locomotion, and dentition of the ruminants.

Identification of a contact zone and hybridization for two subspecies of the American pika (Ochotona princeps) within a single protected area.

Genetic variation is the basis upon which natural selection acts to yield evolutionary change. In a rapidly changing environment, increasing genetic variation should increase evolutionary potential, particularly for small, isolated populations. However, the introduction of new alleles, either through natural or human-mediated processes, may have unpredictable consequences such as outbreeding depression. In this study, we identified a contact zone and limited gene flow between historically separated genetic lineages of American pikas (Ochotona princeps), representing the northern and southern Rocky Mountain subspecies, within Rocky Mountain National Park. The limited spatial extent of gene flow observed may be the result of geographic barriers to dispersal, selection against hybrid individuals, or both. Our fine-scale population genetic analysis suggests gene flow is limited but not completely obstructed by extreme topography such as glacial valleys, as well as streams including the Colorado River. The discovery of two subspecies within this single protected area has implications for monitoring and management, particularly in the light of recent analyses suggesting that the pikas in this park are vulnerable to fragmentation and local extinction under future projected climates. Future research should focus on the fitness consequences of introgression among distinct genetic lineages in this location and elsewhere, as well as within the context of genetic rescue as a conservation and management strategy for a climate sensitive species.

Assessing changes in functional connectivity in a desert bighorn sheep metapopulation after two generations.

Determining how species move across complex and fragmented landscapes and interact with human-made barriers is a major research focus in conservation. Studies estimating functional connectivity from movement, dispersal or gene flow usually rely on a single study period and rarely consider variation over time. We contrasted genetic structure and gene flow across barriers for a metapopulation of desert bighorn sheep (Ovis canadensis nelsoni) using genotypes collected 2000-2003 and 2013-2015. Based on the recently observed but unexpected spread of a respiratory pathogen across an interstate highway previously identified as a barrier to gene flow, we hypothesized that bighorn sheep changed how they interacted with that barrier, and that shifts in metapopulation structure influenced gene flow, genetic diversity and connectivity. Population assignment tests, genetic structure and genetic recapture demonstrated that bighorn sheep crossed the interstate highway in at least one location in 2013-2015, sharply reducing genetic structure between two populations, but supported conclusions of an earlier study that such crossings were very infrequent or unknown in 2000-2003. A recently expanded population established new links and caused decreases in genetic structure among multiple populations. Genetic diversity showed only slight increases in populations linked by new connections. Genetic structure and assignments revealed other previously undetected changes in movements and distribution, but much was consistent. Thus, we observed changes in both structural and functional connectivity over just two generations, but only in specific locations. Movement patterns of species should be revisited periodically to enable informed management, particularly in dynamic and fragmented systems.

Detection of bacterial-reactive natural IgM antibodies in desert bighorn sheep populations.

Ecoimmunology is a burgeoning field of ecology which studies immune responses in wildlife by utilizing general immune assays such as the detection of natural antibody. Unlike adaptive antibodies, natural antibodies are important in innate immune responses and often recognized conserved epitopes present in pathogens. Here, we describe a procedure for measuring natural antibodies reactive to bacterial antigens that may be applicable to a variety of organisms. IgM from desert bighorn sheep plasma samples was tested for reactivity to outer membrane proteins from Vibrio coralliilyticus, a marine bacterium to which sheep would have not been exposed. Immunoblotting demonstrated bighorn sheep IgM could bind to a variety of bacterial cell envelope proteins while ELISA analysis allowed for rapid determination of natural antibody levels in hundreds of individual animals. Natural antibody levels were correlated with the ability of plasma to kill laboratory strains of E. coli bacteria. Finally, we demonstrate that natural antibody levels varied in two distinct populations of desert bighorn sheep. These data demonstrate a novel and specific measure of natural antibody function and show that this varies in ecologically relevant ways.

Simulating the spread of selection-driven genotypes using landscape resistance models for desert bighorn sheep.

Landscape genetic studies based on neutral genetic markers have contributed to our understanding of the influence of landscape composition and configuration on gene flow and genetic variation. However, the potential for species to adapt to changing landscapes will depend on how natural selection influences adaptive genetic variation. We demonstrate how landscape resistance models can be combined with genetic simulations incorporating natural selection to explore how the spread of adaptive variation is affected by landscape characteristics, using desert bighorn sheep (Ovis canadensis nelsoni) in three differing regions of the southwestern United States as an example. We conducted genetic sampling and least-cost path modeling to optimize landscape resistance models independently for each region, and then simulated the spread of an adaptive allele favored by selection across each region. Optimized landscape resistance models differed between regions with respect to landscape variables included and their relationships to resistance, but the slope of terrain and the presence of water barriers and major roads had the greatest impacts on gene flow. Genetic simulations showed that differences among landscapes strongly influenced spread of adaptive genetic variation, with faster spread (1) in landscapes with more continuously distributed habitat and (2) when a pre-existing allele (i.e., standing genetic variation) rather than a novel allele (i.e., mutation) served as the source of adaptive genetic variation. The combination of landscape resistance models and genetic simulations has broad conservation applications and can facilitate comparisons of adaptive potential within and between landscapes.