Feral Horse Populations Research Articles

Methanogenic patterns in the gut microbiome are associated with survival in a population of feral horses

Gut microbiomes are widely hypothesised to influence host fitness and have been experimentally shown to affect host health and phenotypes under laboratory conditions. However, the extent to which they do so in free-living animal populations and the proximate mechanisms involved remain open questions. In this study, using long-term, individual-based life history and shallow shotgun metagenomic sequencing data (2394 fecal samples from 794 individuals collected between 2013–2019), we quantify relationships between gut microbiome variation and survival in a feral population of horses under natural food limitation (Sable Island, Canada), and test metagenome-derived predictions using short-chain fatty acid data. We report detailed evidence that variation in the gut microbiome is associated with a host fitness proxy in nature and outline hypotheses of pathogenesis and methanogenesis as key causal mechanisms which may underlie such patterns in feral horses, and perhaps, wild herbivores more generally.

Comparing methods to estimate feral burro abundance

AbstractObtaining precise and unbiased estimates of feral burro (Equus asinus) abundance in the western United States is challenging due to their cryptic pelage and the rugged terrain they inhabit. Management agencies employ helicopter‐based, simultaneous double‐observer sightability surveys (hereafter denoted as DOS) to estimate abundance of burros; but the DOS method routinely produces negatively biased estimates due to residual heterogeneity in detection probability. Consequently, testing alternative methods to improve upon current procedures is warranted. Residual heterogeneity in DOS surveys can be minimized by including radio‐collared individuals in the population. Alternatively, if distance measurements are recorded, residual heterogeneity can also be reduced via a mark‐recapture distance sampling (MRDS) approach. Aerial infrared (IR) surveys offer a safer alternative than helicopter‐based surveys because they can be flown at a higher altitude and require fewer observers in the aircraft. Further, IR surveys using a distance sampling approach have been shown to generate accurate and precise estimates of feral horse (E. caballus) populations. Accordingly, we compared results of surveys using aerial IR distance sampling, the standard DOS survey, a DOS survey incorporating detections of radio‐collared individuals, and an MRDS analysis of a feral burro population with a known minimum population size in central Utah, winter 2015–2016 and spring 2016. The minimum number of burros known alive during the winter and spring surveys were 236 and 136, respectively. The average detection probability of IR surveys was P = 0.88 (SE = 0.16) and distance models produced estimates of 127 burros (95% CIs = 99–175) for the winter survey, and 94 burros (CIs = 72–134) for the spring survey. Mean detection probability of the standard DOS surveys was P = 0.78 (SE = 0.09), and model‐generated abundance estimates were 155 burros (CIs = 133–227) in winter, and 92 burros (CIs = 79–139) in spring. Incorporating detections of radio‐collared individuals in the DOS survey resulted in a decreased detection probability (P = 0.46; SE = 0.06) and increased abundance estimates to 267 (CIs = 169–571) and 155 (CIs = 128–263) for winter and spring, respectively. Mark‐recapture distance sampling produced a mean detection probability of P = 0.48 (SE = 0.12) and resulted in estimates of 282 (CIs = 178–385) and 169 (CIs = 73–310) burros in winter and spring, respectively. Our study demonstrated that aerial IR surveys conducted using standard distance sampling can produce precise estimates of burro population sizes; however, estimates were negatively biased relative to the known population size. Small sample size limits generalization of our results, but the IR‐based distance approach did not improve upon DOS surveys. Accounting for residual heterogeneity through use of radio‐collars and mark‐recapture distance sampling eliminated the negative bias from the standard DOS survey but decreased survey precision. Managers will need to decide whether unbiased but less precise abundance estimates are preferable compared to a more precise, but biased, estimate.

Carbon emissions from Australian Sphagnum peatlands increase with feral horse (Equus caballus) presence

Peatlands are globally significant carbon sinks, but when disturbed, have the potential to release carbon back to the atmosphere as greenhouse gases. Feral horse populations in the Australian Alps degrade Sphagnum peatlands, which are highly sensitive to disturbance. However, the link between this degradation and peatland carbon cycling is not understood. Here, we compared the autumn daytime carbon dioxide (CO2) and methane (CH4) fluxes of 12 alpine and subalpine Sphagnum peatlands in Kosciuszko National Park, Australia. The presence of feral horses at these sites was correlated with higher carbon loss: sites with horses were losing carbon to the atmosphere (4.83 and 8.18 g CO2-e m−2 d−1 in areas of Sphagnum moss and bare soil, respectively), whereas sites without horses were removing carbon from the atmosphere (−6.39 g CO2-e m−2 d−1). Sites with feral horses also had higher soil bulk density, temperature, and electrical conductivity (EC), and higher water pH, EC, and turbidity, than sites without horses. Our findings suggest that excluding feral horses from peatland areas could reduce rates of carbon loss to the atmosphere, in addition to improving overall site condition, peat soil condition, and water quality. We discuss potential management applications, further research, and restoration opportunities arising from these results.

PopEquus: A predictive modeling tool to support management decisions for free‐roaming horse populations

AbstractFeral horse (Equus caballus) population management is a challenging problem around the world because populations often exhibit density‐independent growth, can exert negative ecological effects on ecosystems, and require great cost to be managed. However, strong value‐based connections between people and horses cause contention around management decisions. To help make informed decisions, natural resource managers might benefit from more detailed understanding of how horse management alternatives, including combinations of removals and fertility control methods, could achieve objectives of sustainable, multiple‐use ecosystems while minimizing overall horse handling and fiscal costs. Here, we describe a modeling tool that simulates horse management alternatives and estimates trade‐offs in predicted metrics related to population size, animal handling, and direct costs of management. The model considers six management actions for populations (removals for adoption or long‐term holding; fertility control treatment with three vaccines, intrauterine devices, and mare sterilization), used alone or in combination. We simulated 19 alternative management scenarios at 2‐, 3‐, and 4‐year management return intervals and identified efficiency frontiers among alternatives for trade‐offs between predicted population size and six management metrics. Our analysis identified multiple alternatives that could maintain populations within target population size ranges, but some alternatives (e.g., removal and mare sterilization, removal and GonaCon treatment) performed better at minimizing overall animal handling requirements and management costs. Cost savings increased under alternatives with more effective, longer lasting fertility control techniques over longer management intervals compared with alternatives with less‐effective, shorter lasting fertility control techniques. We built a user‐friendly website application, PopEquus, that decision makers and interested individuals can use to simulate management alternatives and evaluate trade‐offs among management and cost metrics. Our results and website application provide quantitative trade‐off tools for horse population management decisions and can help support value‐based management decisions for wild or feral horse populations and ecosystems at local and regional scales around the world.

Hair Cortisol Concentrations in Feral Horses and the Influence of Physiological and Social Factors

Cortisol is a glucocorticoid hormone produced during activation of the hypothalamic–pituitary–adrenal axis (HPA) in response to psychological or physiological demands. High amounts of circulating cortisol can be found in individuals experiencing energetically demanding physiological events, such as pregnancy, lactation, injury, or starvation, but, also, in individuals who may have less obvious HPA activation from social situations. The feral horse population on Sable Island (Nova Scotia, Canada) provides an opportunity to look at hair cortisol concentration (HCC) as a proxy for circulating cortisol concentration to better understand physiological correlates. The horse’s complex social structure also allows us to look at how the population and group structure may influence HPA activation. Hair samples (n = 282) were analyzed from 113 females and 135 males. Females with dependent offspring (foals) had higher HCC than those females without dependent offspring (p = 0.005). Horses in poor body condition were also more likely to have higher HCC (females: p < 0.001, males: p = 0.028); females had greater variation in the body condition index (BCI), which also correlated with foal production. In general, the top-ranked models describing female cortisol levels included age, BCI, presence of a foal, as well as social measures such as harem size and the number of bachelors in the vicinity. The top model describing male cortisol levels included age, BCI, and year of collection only, and the number of bachelors in the home range appeared in subsequent, though still high-ranked, models. Among the variables not of direct interest, we found some significant results relating to hair color and hair texture. Differences in HCC patterns between feral and domestically kept horses (e.g., age and sex) are likely linked to periods of resource limitations, particularly for individuals experiencing energetically demanding processes such as reproduction, illness/parasitism, or related to experiencing the full range of social and reproductive behaviors.

Use of density‐impact functions to inform and improve the environmental outcomes of feral horse management

The available science often demonstrates the need for feral horse population control but not the degree of control required to achieve environmental conservation objectives. To better manage the influence of feral horses, we must first understand the relationship between feral horse density and environmental impact. We recorded vegetation and soil disturbance, and the sign of potential causes of this impact in two parts of the Australian Alps, the Bogong high plains (BHP) and the Eastern Victorian Alps (EVA). We calculated density‐impact functions to assist managers with determining feral horse density targets for control programmes. Minimal sign of feral horse impact was detected on the BHP, with no impact of feral horses observed along 99% of the length of transects. In contrast, impacts assigned to feral horses were significantly higher in the EVA, where a larger, higher‐density population of feral horses existed. However, greater than 83% of the walked transect length was still undisturbed by feral horses in the EVA. We detected a threshold of horse impact at ~ 250 horse faecal piles per ha. Above this threshold, a slight increase in horse density resulted in a disproportionately large increase in impact. In this context, a relatively small population control effort may substantially reduce direct horse impact. But where horse densities exist below this threshold, considerably more expense and control effort (resulting from the difficulties related to control at low density) is likely to make very little difference to an already low level of direct impact. The combined impacts associated with the sign of deer, feral pigs, fire and humans were large compared to that of feral horses. Management of feral horses to reduce their direct impact is unlikely to be beneficial without complementary management to reduce the effects of these other agents of impact.

Proximate factors affecting mortality and maternal abandonment of young free-roaming feral horse foals

The burgeoning population of feral horses in the American west is due to high population growth, resulting from low adult mortality and high foal survival. In two populations of feral horses in western Utah, USA only 15 foals died (5%; mean age <1 month) over a 4-year period. Seven additional foals (age <70 days) were observed separated from their dam, with no return to suckling or associating with the dam (i.e., abandoned). Factors affecting fate of foals were examined by comparing dead and separated foals with siblings (n = 19 dams, n = 32 siblings). Foals becoming separated or dying were observed in all years of the study, were unrelated to horse density, environmental effects, or gather events. There was no effect of dam body condition, parity, or age on foal survival or separation, and no effect of length of time the dam was in a group, whether the foal was born into the same group as conceived, and number of group changes made by the dam while pregnant. Dams of foals that died or were separated were more likely to change groups within 2 months after the foal was first seen, mostly after foal death or separation. Separated foals were near their dam less often, but there was no difference in frequency of social interactions. Separation of foals and dams is a natural occurrence in feral horses and survival likelihood for these foals is high – all separated foals that remained on the range in this study survived.

Evaluation of recurrent laryngeal neuropathy in domestic and feral horse populations in Australia using histologic and immunohistochemical analysis: A pilot study.

Little is known about potential differences in the left recurrent laryngeal nerve (Lrln) and left cricoarytenoideus dorsalis (LCAD) muscle between domestic and feral horse populations. If a difference exists, feral horses may provide a useful control population for research related to recurrent laryngeal neuropathy (RLN) and increase our understanding of potential population pressures influencing the incidence RLN. The objective of this study was to compare the Lrln and LCAD of domestic and feral horses using histological and immunohistochemical techniques (IHC). Sixteen horses, domestic (n = 8) and feral (n = 8), without clinical or ancillary examinations that were processed at an abattoir had the Lrln and LCAD muscle harvested immediately following death. Carcass weights were recorded. Subjective and morphometric histologic assessment were performed on Lrln sections. The LCAD was assessed for myosin heavy chain (fibre type proportion, diameter and grouping using IHC. Fibre-type grouping consistent with RLN was seen in both groups. Regenerating fibre clusters were more common in domestic compared to feral horses (p = 0.04). No other histologic differences occurred between groups. Muscle fibre typing demonstrated a lower mean percentage of type IIX fibres in the feral group compared to the domestic group (p = 0.03). There was no difference in type I or IIA proportions or mean diameter of any fibre type between the groups. The domestic population showed evidence of nerve regeneration suggesting RLN in this group, yet this was not supported by the higher proportion of type IIX muscle fibres compared to the feral population. Further evaluation to clarify the significance and wider occurrence of the differences is indicated.

Long-term serological surveillance for West Nile and Usutu virus in horses in south-West Spain

West Nile virus (WNV) is a re-emerging zoonotic pathogen with increasing incidence in Europe, producing a recent outbreak in 2020 in Spain with 77 human cases and eight fatalities. However, the factors explaining the observed changes in the incidence of WNV in Europe are not completely understood. Longitudinal monitoring of WNV in wild animals across Europe is a useful approach to understand the eco-epidemiology of WNV in the wild and the risk of spillover into humans. However, such studies are very scarce up to now. Here, we analysed the occurrence of WNV and Usutu virus (USUV) antibodies in 2102 samples collected between 2005 and 2020 from a population of feral horses in Doñana National Park. The prevalence of WNV antibodies varied between years, with a mean seroprevalence of 8.1% (range 0%–25%) and seasonally. Climate conditions including mean minimum annual temperatures and mean rainy days per year were positively correlated with WNV seroprevalence, while the annual rainfall was negatively. We also detected the highest incidence of seroconversions in 2020 coinciding with the human outbreak in southern Spain. Usutu virus-specific antibodies were detected in the horse population since 2011. The WNV outbreak in humans was preceded by a long period of increasing circulation of WNV among horses with a very high exposure in the year of the outbreak. These results highlight the utility of One Health approaches to better understand the transmission dynamics of zoonotics pathogens.

39 An in-Vitro Model of the Equine Fecal Microbiome to Assess How Horse Population Affects Fermentation Following a Starch Challenge

Abstract The use of an in-vitro model of the equine microbiome is beneficial to assess how fermentation patterns differ based on a horse populations’ habitual diet. The purpose of this study was to determine the in-vitro fermentation patterns of the microbial community within different horse populations before and following a starch challenge. Fecal samples were taken from three different populations of horses: horses from the Shackleford Banks, a North Carolina feral horse population living on native grasses; horses from the NCSU Equine Educational Unit that are predominantly kept on cool season mixed pastures, though may be supplemented with hay and concentrates when warranted; and privately owned horses that were fed mixed diets consisting of grass hay, concentrates and some pasture. Horses were monitored and fecal samples were collected immediately following a void and were stored on dry ice and frozen until analysis. Fecal samples from individual horses were pooled to form a representative sample for each population and mixed with an anaerobic medium to prepare an inoculum. The inoculum was placed into bottles containing either a treatment substrate of alfalfa (A) or of alfalfa and starch (AS). Bottles were capped, purged with CO2 and placed in a water bath at 39°C to incubate for 0, 2, 4 or 24 hours. Culture samples were processed to measure methane and short chain fatty acids (SCFAs; acetate, propionate, butyrate and other isoacids) using gas chromatography. Results were analyzed using the Proc Mixed procedure in SAS to compare the effects of horse population, time and treatment. Methane was significantly greater after 24 hours within all populations with AS, as the inoculum compared with A alone (P = 0.03). Propionate was greater for AS (molar percentage, mean ± standard deviation; 2.21 ± 4.97%) versus the A treatment (11.95 ± 4.97%, P = 0.02). Acetate concentrations were significantly (P &amp;lt; 0.001) greater within the A treatment in the Shackleford and NCSU horses (60.31 ± 10.3% and 62.18 ± 11.47%, respectively) compared with the AS treatment (59.09 ± 11.21% and 59.77 ± 8.85%, respectively). Privately owned horses showed similar values of acetate concentration when comparing the treatment of A (63.42 ± 10.13%) versus AS (63.55 ± 10.77%). Butyrate concentrations were greater in the Shackleford Banks and NCSU horses (13.16 ± 1.92% and 2.64 ± 4.22%, respectively) compared with the privately owned horses (10.96 ± 2%). Isoacids were greater in the Shackleford Banks horses (10.19 ± 4.15%) than the NCSU horses (9.87 ± 3.98%) and the privately owned horses (9.52 ± 3.53%; P&amp;lt; 0.0001). It appears that fermentation of starch differs between these horse populations, likely due to their habitual diet.

138 Immunization against oocyte growth factors in feral mares

Feral horse populations continue to grow, and management is focused on removal both at a substantial cost in the process and their long-term care. Fertility control has been identified as a means to provide a solution. Two EPA registered immunocontraceptives (Gonacon and ZonaStat-H) are available for equids, but both require an initial dose and at least one booster ranging from weeks to years later making this relatively impractical. Our aim was to develop a single dose long-lasting contraceptive targeting 2 oocyte-specific growth factors (OGF) GDF-9 and BMP-15. In our proof-of-concept study, GDF-9 antigen representing 14 amino acids of the mature protein, and a BMP-15 antigen representing 24 amino acids in the mature sequence were produced. Both were conjugated to keyhole limpet hemocyanin (KLH), and the experimental vaccine was manufactured using Pet-Gel A as an adjuvant. We reported that domestic mares were treated with an initial dose and 4 boosters at 6 week intervals. All treated mares developed antibodies and demonstrated infertility for an entire year, and 88% of the mares remained infertile for the following year. The current investigation was conducted to examine the effects of a single dose with a potentially long-lasting immunostimulant. We enrolled 32 mares and 4 stallions with proven fertility that had been removed from the Nevada range that year. All mares had produced foalsthe year before. In May, 16 of these mares were treated with a new OGF vaccine formulation using the same antigens conjugated to KLH and AdjuVac adjuvant containing liposomes. Another 16 mares received a sham injection of adjuvant alone. Serum samples were collected monthly for titer analysis via enzyme-linked immunosorbent assay and progesterone assays via radioimmunoassay. Pregnancy determination was also conducted monthly via transrectal ultrasonography. All mares were then housed with fertile stallions in August. Four treated and 4 control mares were housed in each of 4 pens with a single stallion. All (16/16) of control mares became pregnant. One treated mare was euthanized for reasons unrelated to treatment. Of the remaining treated mares 67% (10/15) became pregnant within 60 d while 20% (3/15) became pregnant much later (180–300 d). Two mares (13%) remained anovulatory for 2 years. The mares with delayed and inhibited fertility demonstrated the greatest (P < 0.001) and longest lasting (P < 0.0001) anti-OGF titers. While this specific vaccine attenuated ovulation it failed to sufficiently alter fertility as pregnancy rates were not different. Our goal is to now find an adjuvant that will result in a single long-lasting vaccine.

3 A comparison of the equine fecal microbiome within different horse populations

The equine fecal microbiome may vary across horse populations due to the diversity of the habitual diet. The purpose of this study was to assess and compare the microbial population of different horse populations, specifically the differences between feral versus domesticated populations. Samples were collected from 3 different populations of horses: horses from the Shackleford Banks (n = 24), a feral horse population living on the Outer Banks of North Carolina who eat native grasses such as Spartina marsh and island grasses; horses from the NCSU Equine Educational Unit (n = 18) that are predominantly kept on cool season mixed pastures and may be supplemented with hay and concentrates from time to time; and finally, privately owned horses (n = 36) that are fed mixed diets consisting of pasture, hay and concentrates. Horses were monitored and samples were collected immediately following a void by swabbing the middle of the void. Swabs were placed in a tube containing 500 uL DNA/RNA shield (Zymo Research, Irvine, CA) and were sent to the Emerging Technology Center (Purina Animal Nutrition, Gray Summit, MO) where they were stored at −80°C and then the V3 and V4 regions of the 16S rRNA gene were sequenced following the Illumina 16S Protocol (San Diego, CA). Samples were processed, filtered and trimmed through DADA2 using the QIIME2 pipeline. Statistical analysis was performed in R(Version 4.1.1) and a P-value of ≤0.05 was considered significant. After processing to eliminate samples with low sampling depth (<20,085), 78 total samples across the 3 populations were analyzed. For the results, when testing α diversity with Shannon's Index, a Kruskal-Wallis rank sum test revealed a significant difference between all populations (P = 0.01). There was a visual distinction between the Shackleford Banks population compared with the others when utilizing Bray-Curtis to assess β diversity. Additionally, an apparent significant difference between all populations using the PERMANOVA UniFrac test (P < 0.001) was observed. The 3 most predominant bacterial phylum seen across all populations were Firmicutes, Bacteroidetes and Spirochaetes. The top 5 phyla observed in the Shackleford Banks population were Firmicutes, Bacteroidetes, Spirochaetes, Kiritimatiellaeota and Fibrobacteres. Based on the results, there is a distinctive separation in microbial diversity between these horse populations, specifically between the Shackleford Banks horses versus the NCSU and privately owned horses. This separation is likely due to the habitual diet of these specific horse populations influencing the composition of their microbiome within the hindgut.

The Amelioration of Grazing through Physiological Integration by a Clonal Dune Plant

Rhizomatous growth and associated physiological integration can allow a clonal dune species to potentially compensate for the selective removal of leaves associated with herbivory. Hydrocotyle bonariensis is a rhizomatous clonal plant species that is abundant in the coastal dune environments of the southeastern United States that are inhabited by large feral horse populations. H. bonariensis has been shown to integrate resources among ramets within extensive clones as an adaptation to resource heterogeneity in sandy soils. In this study, we hypothesized that clonal integration is a mechanism that promotes H. bonariensis persistence in these communities, despite high levels of herbivory by feral horses. In a field experiment, we used exclosures to test for herbivory in H. bonariensis over a four-month period. We found that feral horses utilized H. bonariensis as a food species, and that while grazing will suppress clonal biomass, H. bonariensis is able to maintain populations in a high grazing regime with and without competition present. We then conducted an experiment in which portions of H. bonariensis clones were clipped to simulate different levels of grazing. Half of the clones were severed to eliminate the possibility of integration. We found that after 12 weeks, the mean number of leaves and ramets increased as the grazing level increased, for integrated clones. Integrated clones had significantly increased biomass production compared to the severed equivalents. Our research suggests that rhizomatous growth and physiological integration are traits that allow clonal plant species to maintain populations and to tolerate grazing in coastal dune environments.

Laissez-Faire Stallions? Males' Fecal Cortisol Metabolite Concentrations Do Not Vary with Increased Female Turnover in Feral Horses (Equus caballus).

Stress responses can be triggered by several physical and social factors, prompting physiological reactions including increases in glucocorticoid concentrations. In a population of feral horses (Equus caballus) on Shackleford Banks, North Carolina, females previously immunized with the immunocontraceptive agent porcine zona pellucida (PZP) change social groups (bands) more often than unimmunized females, disrupting the social stability within the population. We assessed the effects of increased female group changing behavior (or female turnover) on individual male stress by comparing fecal cortisol metabolite (FCM) concentrations among stallions experiencing varying amounts of female group changing behavior. FCM concentrations did not significantly correlate with female turnover. Similarly, FCM concentrations were not dependent upon the timing of female group changing behavior. These findings suggest that female turnover rate has little influence on physiological measures of stress in associated stallions. That said, Shackleford stallions experiencing increased female turnover do engage in behaviors typically associated with stress (increased vigilance, highly escalated male-male conflicts). Future work should compare FCM concentrations across time within populations and among populations managed under different strategies to better isolate factors influencing stallion stress physiology. Such studies are especially important if we are to determine how changes in female behavior related to immunocontraception impact physiological and behavioral indicators of stress for non-target animals. Finally, our study highlights the importance of considering both physiological and behavioral measures when investigating animal responses to potentially challenging situations.

Seasonal resource selection and movement ecology of free‐ranging horses in the western United States

AbstractUnderstanding factors driving resource selection and habitat use of different species is an important component of management and conservation. Feral horses (Equus caballus) are free ranging across various vegetation types in the western United States, yet few studies have quantified their resource selection and seasonal use. We conducted a study to determine effects of vegetation community, distance to water, and topographic variables on seasonal resource selection in 2 feral horse populations in Great Basin sagebrush (Artemisia spp.) ecosystems of west‐central Utah, USA: Conger Herd Management Area (HMA) and Frisco HMA. We deployed global positioning system (GPS) radio‐collars on 38 female horses and GPS‐transmitters braided and glued into the tail hair of 14 males, collecting locations every 2 hours for 1–4 years between 2016 and 2020. We calculated home range size and core use area of social groups (harems) and bachelor males using auto‐correlated kernel density estimators for each biologically defined season (breeding, fall, and winter) per study year. We examined seasonal home range size and overlap of harem groups and bachelor males and compared movement speed of bachelors and harems among seasons. We determined seasonal resource selection in a use‐availability framework using resource selection functions. We hypothesized that horses would select for areas of high herbaceous vegetation, that water would be a key variable in resource selection models like other equids, and home range size in winter would be largest because horses can eat snow for hydration and could therefore roam farther from surface water. Mean annual home range size was 103.12 ± 37.38 km2 (SD) for Conger harems and 117.47 ± 32.75 km2 for Frisco harems. At Conger there was no difference in home range size between harem groups and bachelor males, but home range size was smaller in winter than other seasons, whereas winter home range size at Frisco was larger than other seasons. Bachelor males moved at higher speeds than harems during all seasons, and harem groups from both populations had lower movement speeds in winter. Harem groups had distinct winter ranges with little overlap on breeding season ranges. In both populations, all horses selected for herbaceous vegetation types and avoided forest relative to shrubland throughout the year. Harems at Frisco were consistently located closer to water sources, whereas selection for water sources by Conger harems varied seasonally, with winter having the lowest selection. Harem groups at Conger had an average of 10.6% of their home ranges outside the HMA boundary and Frisco harems had up to 66.8% outside, likely because of the horseshoe shape of Frisco HMA in which shrub meadows (foraging areas) comprise the horseshoe center, which is outside the HMA. Our results highlight the importance of water sources, which were a key predictor of horse movement patterns in our study. We emphasize the utility of telemetry devices to understand resource selection of feral horses at a fine scale, enabling management to be more targeted and facilitate planning.

Reconstruction of the Major Maternal and Paternal Lineages in the Feral New Zealand Kaimanawa Horses.

New Zealand has the fourth largest feral horse population in the world. The Kaimanawas (KHs) are feral horses descended from various domestic horse breeds released into the Kaimanawa ranges in the 19th and 20th centuries. Over time, the population size has fluctuated dramatically due to hunting, large-scale farming and forestry. Currently, the herd is managed by an annual round-up, limiting the number to 300 individuals to protect the native ecosystem. Here, we genotyped 96 KHs for uniparental markers (mitochondrial DNA, Y-chromosome) and assessed their genetic similarity with respect to other domestic horses. We show that at least six maternal and six paternal lineages contributed unequally to the KH gene pool, and today's KH population possibly represents two sub-populations. Our results indicate that three horse breeds, namely Welsh ponies, Thoroughbreds and Arabian horses had a major influence in the genetic-makeup of the extant KH population. We show that mitochondrial genetic diversity in KHs (π = 0.00687 ± 0.00355) is closer to that of the Sable Island horses (π = 0.0034 ± 0.00301), and less than other feral horse populations around the world. Our current findings, combined with ongoing genomic research, will provide insight into the population-specific genetic variation and inbreeding among KHs. This will largely advance equine research and improve the management of future breeding programs of these treasured New Zealand horse.

Shallow shotgun sequencing of the microbiome recapitulates 16S amplicon results and provides functional insights.

Prevailing 16S rRNA gene-amplicon methods for characterizing the bacterial microbiome of wildlife are economical, but result in coarse taxonomic classifications, are subject to primer and 16S copy number biases, and do not allow for direct estimation of microbiome functional potential. While deep shotgun metagenomic sequencing can overcome many of these limitations, it is prohibitively expensive for large sample sets. Here we evaluated the ability of shallow shotgun metagenomic sequencing to characterize taxonomic and functional patterns in the faecal microbiome of a model population of feral horses (Sable Island, Canada). Since 2007, this unmanaged population has been the subject of an individual-based, long-term ecological study. Using deep shotgun metagenomic sequencing, we determined the sequencing depth required to accurately characterize the horse microbiome. In comparing conventional vs. high-throughput shotgun metagenomic library preparation techniques, we validate the use of more cost-effective laboratory methods. Finally, we characterize similarities between 16S amplicon and shallow shotgun characterization of the microbiome, and demonstrate that the latter recapitulates biological patterns first described in a published amplicon data set. Unlike for amplicon data, we further demonstrate how shallow shotgun metagenomic data provide useful insights regarding microbiome functional potential which support previously hypothesized diet effects in this study system.

Features of reproduction in an isolated island population of the feral horses of the Lake Manych-Gudilo (Rostov Region, Russia)

The article describes the results of a pilot study of the reproduction features of a small population of feral horses (Equus ferus) isolated for about 70 years on the Vodny Island in the Lake Manych-Gudilo (Rostov Nature Reserve, Rostov Region, Russia). The population was characterized by a fairly stable social structure for more than 10 years, including a constant composition of adult members of harem bands, though the scarce cases of mares being covered by ‘outsider’ stallions were recorded, which could be considered as one of the variants of sneak mating. The main purposes of the study were to confirm by molecular genetic methods the possible participation in the reproduction of ‘outsider’ stallions. The study was based on a combination of the direct ethological observations and genetic analyses of the individual animals. The ethological observations conducted during more than ten years revealed that the harem bands remained relatively stable throughout the period of the study. Both direct observations and genetic data revealed that the stallions heading neighboring harem bands actually participated in the fertilization of the mares. The subdominant stallions in the multi-male family bands also occasionally participated in the reproduction. No cases of participation of the bachelor stallions in the reproduction were recorded, which can be explained by their low social rank. The participation of the ‘outsider’ stallions in reproduction increases the overall genetic diversity in the population. With this, several instances of mating by close relatives were also observed that might result at random or forcibly under conditions of the complete isolation of a rather small population. The levels of observed and estimated heterozygosity (HO = 0.581 and HE = 0.606) were estimated for the first time. Such ratio of HO to HE corresponds to the provisions of the Wahlund effect and seems to indicate a small impact of the reproduction of the ‘outsider’ stallions on the level of genetic diversity in the studied local feral horse population.

Runs of homozygosity in Sable Island feral horses reveal the genomic consequences of inbreeding and divergence from domestic breeds

BackgroundUnderstanding inbreeding and its impact on fitness and evolutionary potential is fundamental to species conservation and agriculture. Long stretches of homozygous genotypes, known as runs of homozygosity (ROH), result from inbreeding and their number and length can provide useful population-level information on inbreeding characteristics and locations of signatures of selection. However, the utility of ROH for conservation is limited for natural populations where baseline data and genomic tools are lacking. Comparing ROH metrics in recently feral vs. domestic populations of well understood species like the horse could provide information on the genetic health of those populations and offer insight into how such metrics compare between managed and unmanaged populations. Here we characterized ROH, inbreeding coefficients, and ROH islands in a feral horse population from Sable Island, Canada, using ~41 000 SNPs and contrasted results with those from 33 domestic breeds to assess the impacts of isolation on ROH abundance, length, distribution, and ROH islands.ResultsROH number, length, and ROH-based inbreeding coefficients (FROH) in Sable Island horses were generally greater than in domestic breeds. Short runs, which typically coalesce many generations prior, were more abundant than long runs in all populations, but run length distributions indicated more recent population bottlenecks in Sable Island horses. Nine ROH islands were detected in Sable Island horses, exhibiting very little overlap with those found in domestic breeds. Gene ontology (GO) enrichment analysis for Sable Island ROH islands revealed enrichment for genes associated with 3 clusters of biological pathways largely associated with metabolism and immune function.ConclusionsThis study indicates that Sable Island horses tend to be more inbred than their domestic counterparts and that most of this inbreeding is due to historical bottlenecks and founder effects rather than recent mating between close relatives. Unique ROH islands in the Sable Island population suggest adaptation to local selective pressures and/or strong genetic drift and highlight the value of this population as a reservoir of equine genetic variation. This research illustrates how ROH analyses can be applied to gain insights into the population history, genetic health, and divergence of wild or feral populations of conservation concern.

Examining the costs and benefits of male-male associations in a group-living equid

In feral horse (Equus caballus) populations, females can associate in year-round social groups with one male (single-stallion (SS) groups) or multiple males (multi-stallion (MS) groups). Given the expectation of intense male competition for mating opportunities, male-male associations raise questions regarding their supporting mechanisms and adaptive consequences. The function of MS groups has been highly debated which led to a series of conflicting hypotheses attempting to describe the relationship between the males, and the costs and benefits of this association in relation to SS groups. Based on previous studies, our goal was to compare the group size and stability, males’ defence behaviour against group outsiders and access to females, levels of male aggression to group members, and the activity budget and body condition of horses in MS and SS groups. Between the months of May to August (breeding season) from 2016 to 2021, we collected demographic and behavioural data from feral Garrano horses living in northern Portugal, where wolf (Canis lupus signatus) predation is an important factor. Contrary to mutualistic hypotheses, our results indicated that SS males had more breeding females than MS groups and group type did not influence female membership stability. However, dominant males in MS groups may be benefitting from the presence of another male via decreased effort in group defence and more resting time. We recorded less aggressive behaviours from dominant to subordinate males in MS groups when subordinates displayed more defensive behaviours against outsiders, but we found little evidence that subordinates were being rewarded with access to females, as proposed by reciprocal-altruism hypotheses. Females in MS groups did not suffer higher levels of male aggression when compared with SS females, and the number of males in the group had no significant influence on the time females spent grazing and resting, and ultimately on their body condition. Future research should explore how MS male associations shape male-female relationships, which could provide further insights to the existence and maintenance of subordinate males. This study is the first to report on predation impact by wolves in MS and SS groups, highlighting the need to further explore how different ecological pressures shape horse behaviour and social organization. Furthermore, our results provide information that can help improve the welfare of domestic horses and can make informed management decisions of feral and free-ranging horse populations, particularly in instances where males are targeted for removals. Considering the advantages, as well as costs, provided by extra males may be useful to managers and horse owners, who can then act accordingly and with the least disturbance to horse populations.