No correlative evidence of costs of infection or immunity on leucocyte telomere length in a wild population of Soay sheep.

Telomere length (TL) is a biomarker hypothesized to capture evolutionarily and ecologically important physiological costs of reproduction, infection and immunity. Few studies have estimated the relationships among infection status, immunity, TL and fitness in natural systems. The hypothesis that short telomeres predict reduced survival because they reflect costly consequences of infection and immune investment remains largely untested. Using longitudinal data from a free-living Soay sheep population, we tested whether leucocyte TL was predicted by infection with nematode parasites and antibody levels against those parasites. Helminth parasite burdens were positively associated with leucocyte TL in both lambs and adults, which is not consistent with TL reflecting infection costs. We found no association between TL and helminth-specific IgG levels in either young or old individuals which suggests TL does not reflect costs of an activated immune response or immunosenescence. Furthermore, we found no support for TL acting as a mediator of trade-offs between infection, immunity and subsequent survival in the wild. Our results suggest that while variation in TL could reflect short-term variation in resource investment or environmental conditions, it does not capture costs of infection and immunity, nor does it behave like a marker of an individual's helminth-specific antibody immune response.

Purifying and balancing selection on embryonic semi-lethal haplotypes in a wild mammal.

Embryonic lethal mutations are arguably the earliest and most severe manifestation of inbreeding depression, but their impact on wild populations is not well understood. Here, we combined genomic, fitness, and life-history data from 5,925 wild Soay sheep sampled over nearly three decades to explore the impact of embryonic lethal mutations and their evolutionary dynamics. We searched for haplotypes that in their homozygous state are unusually rare in the offspring of known carrier parents and found three putatively semi-lethal haplotypes with 27%-46% fewer homozygous offspring than expected. Two of these haplotypes are decreasing in frequency, and gene-dropping simulations through the pedigree suggest that this is partially due to purifying selection. In contrast, the frequency of the third semi-lethal haplotype remains relatively stable over time. We show that the haplotype could be maintained by balancing selection because it is also associated with increased postnatal survival and body weight and because its cumulative frequency change is lower than in most drift-only simulations. Our study highlights embryonic mutations as a largely neglected contributor to inbreeding depression and provides a rare example of how harmful genetic variation can be maintained through balancing selection in a wild mammal population.

Trait-dependent associations between early- and late-life reproduction in a wild mammal.

Early- versus late-life trade-offs are a central prediction of life-history theory that are expected to shape the evolution of ageing. While ageing is widely observed in wild vertebrates, evidence that early-late trade-offs influence ageing rates remains limited. Vertebrate reproduction is a complex, multi-stage process, yet few studies have examined how different aspects of early-life reproductive allocation shape late-life performance and ageing. Here, we use longitudinal data from a 36-year study of wild Soay sheep to show that early-life reproduction predicts late-life reproductive performance in a trait-dependent manner. Females that started breeding earlier showed more rapid declines in annual breeding probability with age, consistent with a trade-off. However, age-related declines in offspring first-year survival and birth weight were not associated with early-life reproduction. Selective disappearance was evident in all three late-life reproductive measures, with longer-lived females having higher average performance. Our results provide mixed support for early-late reproductive trade-offs and show that the way early-life reproduction shapes late-life performance and ageing can differ among reproductive traits.

Age-specific impacts of vegetation functional traits on gastrointestinal nematode parasite burdens in a large herbivore.

Gastrointestinal nematode (GIN) parasites play an important role in the ecological dynamics of many animal populations. Recent studies suggest that fine-scale spatial variation in GIN infection dynamics is important in wildlife systems, but the environmental drivers underlying this variation remain poorly understood. We used data from over two decades of GIN parasite egg counts, host space use, and spatial vegetation data from a long-term study of Soay sheep on St Kilda to test how spatial autocorrelation and vegetation in an individual's home range predict parasite burden across three age groups. We developed a novel approach to quantify the plant functional traits present in a home range to describe the quality of vegetation present. Effects of vegetation and space varied between age classes. In immature lambs, strongyle parasite faecal egg counts (FEC) were spatially structured, being highest in the north and south of our study area. Independent of host body weight and spatial autocorrelation, plant functional traits predicted parasite egg counts. Higher egg counts were associated with more digestible and preferred plant functional traits, suggesting the association could be driven by host density and habitat preference. In contrast, we found no evidence that parasite FEC were related to plant functional traits in the host home range in yearlings or adult sheep. Adult FEC were spatially structured, with highest burdens in the north-east of our study area, while yearling FEC showed no evidence of spatial structuring. Parasite burdens in immature individuals appear more readily influenced by fine-scale spatial variation in the environment, highlighting the importance of such heterogeneity for our understanding of wildlife epidemiology and health. Our findings support the importance of fine-scale environmental variation for wildlife disease ecology and provides new evidence that such effects may vary across demographic groups within a population.

Correlates of early reproduction and apparent fitness consequences in male Soay sheep.

Life history trade-offs are ubiquitous across species and place constraints on the timing of life history events, including the optimal age at first reproduction. However, studies on lifetime breeding success of male mammals are rare due to sex-biased dispersal and the requirement for genetic paternity inferences. We studied the correlates and apparent fitness consequences of early life reproduction among males in a free-living population of Soay sheep (Ovis aries) on St Kilda, Scotland. We investigated the factors associated with early breeding success and the apparent consequences of early success for survival and future reproduction. We used genetic paternity inferences, population data, and individual morphology measurements collected over 30 years. We found that individuals born in years with low-density population size had the highest early life breeding success and singletons were more likely to be successful than twins. Individuals that bred successfully at 7 months were more likely to survive their first winter. For individuals that survived their first winter, early breeding success was not associated with later breeding success. As individual heterogeneity affects breeding success, we believe that variation in individual quality masks the costs of early reproduction in this population. Our findings provide no evidence for selection for delayed age at reproduction in male Soay sheep.

Having a better home range does not reduce the cost of reproduction in Soay sheep.

A cost of reproduction may not be observable in the presence of environmental or individual heterogeneity because they affect the resources available to individuals. Individual space use is critical in determining both the resources available to individuals and the exposure to factors that mediate the value of these resources (e.g. competition and parasitism). Despite this, there has, to our knowledge, been little research to understand how between-individual differences in resource acquisition, caused by variation in space use, interact with environmental variation occurring at the population scale to influence estimates of the cost of reproduction in natural populations. We used long-term data from the St. Kilda Soay sheep population to understand how differences in age, relative home range quality, and average adult body mass, interacted with annual variation in population density and winter North Atlantic Oscillation index to influence over-winter survival and reproduction in the subsequent year, for females that had invested into reproduction to varying degrees. Our results suggest that Soay sheep females experience costs both in terms of future survival and future reproduction. However, we found little evidence that estimated costs of reproduction vary depending on relative home range quality. There are several possible causes for the lack of a relationship between relative home range quality and our estimate of the costs experienced by females. These include the potential for a correlation between relative home range quality and reproductive allocation to mask a relationship between home range quality and reproductive costs, as well as the potential for the benefit of higher quality home ranges being offset by higher densities. Nevertheless, our results raise questions regarding the presence or context-dependence of relationships between resource access and the estimated cost of reproduction.

Long-term temporal trends in gastrointestinal parasite infection in wild Soay sheep.

Monitoring the prevalence and abundance of parasites over time is important for addressing their potential impact on host life histories, immunological profiles and their influence as a selective force. Only long-term ecological studies have the potential to shed light on both the temporal trends in infection prevalence and abundance and the drivers of such trends, because of their ability to dissect drivers that may be confounded over shorter time scales. Despite this, only a relatively small number of such studies exist. Here, we analysed changes in the prevalence and abundance of gastrointestinal parasites in the wild Soay sheep population of St. Kilda across 31 years. The host population density (PD) has increased across the study, and PD is known to increase parasite transmission, but we found that PD and year explained temporal variation in parasite prevalence and abundance independently. Prevalence of both strongyle nematodes and coccidian microparasites increased during the study, and this effect varied between lambs, yearlings and adults. Meanwhile, abundance of strongyles was more strongly linked to host PD than to temporal (yearly) dynamics, while abundance of coccidia showed a strong temporal trend without any influence of PD. Strikingly, coccidian abundance increased 3-fold across the course of the study in lambs, while increases in yearlings and adults were negligible. Our decades-long, intensive, individual-based study will enable the role of environmental change and selection pressures in driving these dynamics to be determined, potentially providing unparalleled insight into the drivers of temporal variation in parasite dynamics in the wild.

Longitudinal dynamics of co-infecting gastrointestinal parasites in a wild sheep population.

Within-year variation in infection is a ubiquitous feature of natural populations, but is determined by a complex interplay of environmental, parasitological and host factors. At the same time, co-infection is the norm in the wild. Longitudinal dynamics of co-infecting parasites may therefore be further complicated by covariation across multiple parasites. Here, we used fecal parasite egg and oocyst counts collected repeatedly from individually marked wild Soay sheep to investigate seasonal dynamics of six gastrointestinal parasite groups. Prevalence and abundance tended to be higher in spring and summer, and abundance was higher in lambs compared to adults. We found that within-year variation in highly prevalent strongyle nematode counts was dependent on adult reproductive status, where reproductive ewes had distinct dynamics compared to males and barren ewes. For similarly prevalent coccidia we found an overall peak in oocyst counts in spring but no differences among males, barren and pregnant ewes. Using multivariate mixed-effects models, we further show that apparent positive correlation between strongyle and coccidia counts was driven by short-term within-individual changes in both counts rather than long-term among-individual covariation. Overall, these results demonstrate that seasonality varies across demographic and parasite groups and highlight the value of investigating co-infection dynamics over time.

Functionally distinct T-helper cell phenotypes predict resistance to different types of parasites in a wild mammal.

The adaptive immune system is critical to an effective response to infection in vertebrates, with T-helper (Th) cells pivotal in orchestrating these responses. In natural populations where co-infections are the norm, different Th responses are likely to play an important role in maintaining host health and fitness, a relationship which remains poorly understood in wild animals. In this study, we characterised variation in functionally distinct Th responses in a wild population of Soay sheep by enumerating cells expressing Th-subset specific transcription factors and quantifying Th-associated cytokines. We tested the prediction that raised Th1 and Th2 responses should predict reduced apicomplexan and helminth parasite burdens, respectively. All measures of Th-associated cytokine production increased with age, while Th17- and regulatory Th-associated cytokine production increased more rapidly with age in males than females. Independent of age, sex, and each other, IL-4 and Gata3 negatively predicted gastro-intestinal nematode faecal egg count, while IFN-γ negatively predicted coccidian faecal oocyst count. Our results provide important support from outside the laboratory that Th1 and Th2 responses predict resistance to different kinds of parasites, and illustrate how harnessing specific reagents and tools from laboratory immunology will illuminate our understanding of host-parasite interactions in the wild.

Vitamin D status is heritable and under environment-dependent selection in the wild.

Vitamin D has a well-established role in skeletal health and is increasingly linked to chronic disease and mortality in humans and companion animals. Despite the clear significance of vitamin D for health and obvious implications for fitness under natural conditions, no longitudinal study has tested whether the circulating concentration of vitamin D is under natural selection in the wild. Here, we show that concentrations of dietary-derived vitamin D2 and endogenously produced vitamin D3  metabolites are heritable and largely polygenic in a wild population of Soay sheep (Ovis aries). Vitamin D2  status was positively associated with female adult survival, and vitamin D3  status predicted female fecundity in particular, good environment years when sheep density and competition for resources was low. Our study provides evidence that vitamin D status has the potential to respond to selection, and also provides new insights into how vitamin D metabolism is associated with fitness in the wild.

Genomic prediction in the wild: A case study in Soay sheep.

Genomic prediction, the technique whereby an individual's genetic component of their phenotype is estimated from its genome, has revolutionised animal and plant breeding and medical genetics. However, despite being first introduced nearly two decades ago, it has hardly been adopted by the evolutionary genetics community studying wild organisms. Here, genomic prediction is performed on eight traits in a wild population of Soay sheep. The population has been the focus of a >30 year evolutionary ecology study and there is already considerable understanding of the genetic architecture of the focal Mendelian and quantitative traits. We show that the accuracy of genomic prediction is high for all traits, but especially those with loci of large effect segregating. Five different methods are compared, and the two methods that can accommodate zero-effect and large-effect loci in the same model tend to perform best. If the accuracy of genomic prediction is similar in other wild populations, then there is a real opportunity for pedigree-free molecular quantitative genetics research to be enabled in many more wild populations; currently the literature is dominated by studies that have required decades of field data collection to generate sufficiently deep pedigrees. Finally, some of the potential applications of genomic prediction in wild populations are discussed.

Associations between MHC class II variation and phenotypic traits in a free-living sheep population.

Pathogen-mediated selection (PMS) is thought to maintain the high level of allelic diversity observed in the major histocompatibility complex (MHC) class II genes. A comprehensive way to demonstrate contemporary selection is to examine associations between MHC variation and individual fitness. As individual fitness is hard to measure, many studies examine associations between MHC variation and phenotypic traits, including direct or indirect measures of adaptive immunity thought to contribute to fitness. Here, we tested associations between MHC class II variation and five phenotypic traits measured in free-living sheep captured in August: weight, strongyle faecal egg count, and plasma IgA, IgE and IgG immunoglobulin titres against the gastrointestinal nematode parasite Teladorsagia circumcincta. We found no association between MHC class II variation and weight or strongyle faecal egg count. We did, however, find associations between MHC class II variation and immunoglobulin levels which varied with isotype, age and sex. Our results suggest associations between MHC and phenotypic traits are more likely to be found for traits more closely associated with pathogen defence than integrative traits such as bodyweight and highlight the association between MHC variation and antibodies in wild populations.

The association between female reproductive performance and leukocyte telomere length in wild Soay sheep.

Telomere length (TL), typically measured across a sample of blood cells, has emerged as an exciting potential marker of physiological state and of the costs of investment in growth and reproduction within evolutionary ecology. While there is mounting evidence from studies of wild vertebrates that short TL predicts raised subsequent mortality risk, the relationship between reproductive investment and TL is less clear cut, and previous studies report both negative and positive associations. In this study, we examined the relationship between TL and different aspects of maternal reproductive performance in a free-living population of Soay sheep. We find evidence for shorter TL in females that bred, and thus paid any costs of gestation, compared to females that did not breed. However, we found no evidence for any association between TL and litter size. Furthermore, females that invested in gestation and lactation actually had longer TL than females who only invested in gestation because their offspring died shortly after birth. We used multivariate models to decompose these associations into among- and within-individual effects, and discovered that within-individual effects were driving both the negative association between TL and gestation, and the positive association between TL and lactation. This suggests that telomere dynamics may reflect recent physiologically costly investment or variation in physiological condition, depending on the aspect of reproduction being investigated. Our results highlight the physiological complexity of vertebrate reproduction, and the need to better understand how and why different aspects of physiological variation underpinning life histories impact blood cell TL.

Mutation load decreases with haplotype age in wild Soay sheep.

Runs of homozygosity (ROH) are pervasive in diploid genomes and expose the effects of deleterious recessive mutations, but how exactly these regions contribute to variation in fitness remains unclear. Here, we combined empirical analyses and simulations to explore the deleterious effects of ROH with varying genetic map lengths in wild Soay sheep. Using a long-term dataset of 4879 individuals genotyped at 417K SNPs, we found that inbreeding depression increases with ROH length. A 1% genomic increase in long ROH (>12.5 cM) reduced the odds of first-year survival by 12.4% compared to only 7.7% for medium ROH (1.56-12.5 cM), whereas short ROH (<1.56 cM) had no effect on survival. We show by forward genetic simulations that this is predicted: compared to shorter ROH, long ROH will have higher densities of deleterious alleles, with larger average effects on fitness and lower population frequencies. Taken together, our results are consistent with the idea that the mutation load decreases in older haplotypes underlying shorter ROH, where purifying selection has had more time to purge deleterious mutations. Finally, our study demonstrates that strong inbreeding depression can persist despite ongoing purging in a historically small population.

Patterns of MHC-dependent sexual selection in a free-living population of sheep.

The MHC is one of the most polymorphic gene clusters in vertebrates and play an essential role in adaptive immunity. Apart from pathogen-mediated selection, sexual selection can also contribute to the maintenance of MHC diversity. MHC-dependent sexual selection could occur via several mechanisms but at present there is no consensus as to which of these mechanisms are involved and their importance. Previous studies have often suffered from limited genetic and behavioural data and small sample size, and were rarely able to examine all the mechanisms together, determine whether signatures of MHC-based non-random mating are independent of genomic effects or differentiate whether MHC-dependent sexual selection takes place at the pre- or post-copulatory stage. In this study, we use Monte Carlo simulation to investigate evidence for non-random MHC-dependent mating patterns by all three mechanisms in a free-living population of Soay sheep. Using 1710 sheep diplotyped at the MHC class IIa region and genome-wide SNPs, together with field observations of consorts, we found sexual selection against a particular haplotype in males at the pre-copulatory stage and sexual selection against female MHC heterozygosity during the rut. We also found MHC-dependent disassortative mating at the post-copulatory stage, along with strong evidence of inbreeding avoidance at both stages. However, results from generalized linear mixed models suggest that the pattern of MHC-dependent disassortative mating could be a by-product of inbreeding avoidance. Our results therefore suggest that while multiple apparent mechanisms of non-random mating with respect to the MHC may occur, some of them have alternative explanations.

Heritable variation in telomere length predicts mortality in Soay sheep.

Telomere length (TL) is considered an important biomarker of whole-organism health and aging. Across humans and other vertebrates, short telomeres are associated with increased subsequent mortality risk, but the processes responsible for this correlation remain uncertain. A key unanswered question is whether TL-mortality associations arise due to positive effects of genes or early-life environment on both an individual's average lifetime TL and their longevity, or due to more immediate effects of environmental stressors on within-individual TL loss and increased mortality risk. Addressing this question requires longitudinal TL and life history data across the entire lifetimes of many individuals, which are difficult to obtain for long-lived species like humans. Using longitudinal data and samples collected over nearly two decades, as part of a long-term study of wild Soay sheep, we dissected an observed positive association between TL and subsequent survival using multivariate quantitative genetic models. We found no evidence that telomere attrition was associated with increased mortality risk, suggesting that TL is not an important marker of biological aging or exposure to environmental stress in our study system. Instead, we find that among-individual differences in average TL are associated with increased lifespan. Our analyses suggest that this correlation between an individual's average TL and lifespan has a genetic basis. This demonstrates that TL has the potential to evolve under natural conditions, and suggests an important role of genetics underlying the widespread observation that short telomeres predict mortality.

Maternally derived anti-helminth antibodies predict offspring survival in a wild mammal.

The transfer of antibodies from mother to offspring provides crucial protection against infection to offspring during early life in humans and domestic and laboratory animals. However, few studies have tested the consequences of variation in maternal antibody transfer for offspring fitness in the wild. Further, separating the immunoprotective effects of antibodies from their association with nutritional resources provided by mothers is difficult. Here, we measured plasma levels of total and parasite-specific antibodies in neonatal (less than 10 days old) wild Soay sheep over 25 years to quantify variation in maternal antibody transfer and test its association with offspring survival. Maternal antibody transfer was predicted by maternal age and previous antibody responses, and was consistent within mothers across years. Neonatal total IgG antibody levels were positively related to early growth, suggesting they reflected nutritional transfer. Neonatal parasite-specific IgG levels positively predicted first-year survival, independent of lamb weight, total IgG levels and subsequent lamb parasite-specific antibody levels. This relationship was partly mediated via an indirect negative association with parasite burden. We show that among-female variation in maternal antibody transfer can have long-term effects on offspring growth, parasite burden and fitness in the wild, and is likely to impact naturally occurring host-parasite dynamics.

The genetic architecture of helminth-specific immune responses in a wild population of Soay sheep (Ovis aries).

Much of our knowledge of the drivers of immune variation, and how these responses vary over time, comes from humans, domesticated livestock or laboratory organisms. While the genetic basis of variation in immune responses have been investigated in these systems, there is a poor understanding of how genetic variation influences immunity in natural, untreated populations living in complex environments. Here, we examine the genetic architecture of variation in immune traits in the Soay sheep of St Kilda, an unmanaged population of sheep infected with strongyle gastrointestinal nematodes. We assayed IgA, IgE and IgG antibodies against the prevalent nematode Teladorsagia circumcincta in the blood plasma of > 3,000 sheep collected over 26 years. Antibody levels were significantly heritable (h2 = 0.21 to 0.57) and highly stable over an individual's lifespan. IgA levels were strongly associated with a region on chromosome 24 explaining 21.1% and 24.5% of heritable variation in lambs and adults, respectively. This region was adjacent to two candidate loci, Class II Major Histocompatibility Complex Transactivator (CIITA) and C-Type Lectin Domain Containing 16A (CLEC16A). Lamb IgA levels were also associated with the immunoglobulin heavy constant loci (IGH) complex, and adult IgE levels and lamb IgA and IgG levels were associated with the major histocompatibility complex (MHC). This study provides evidence of high heritability of a complex immunological trait under natural conditions and provides the first evidence from a genome-wide study that large effect genes located outside the MHC region exist for immune traits in the wild.

Pedigree-Based Estimation of Reproductive Value.

How successful an individual or cohort is, in terms of their genetic contribution to the future population, is encapsulated in the concept of reproductive value, and is crucial for understanding selection and evolution. Long-term studies of pedigreed populations offer the opportunity to estimate reproductive values directly. However, the degree to which genetic contributions, as defined by a pedigree, may converge on their long-run values within the time frames of available data sets, such that they may be interpreted as estimates of reproductive value, is unclear. We develop a system for pedigree-based calculation of the expected genetic representation that both individuals and cohorts make to the population in the years following their birth. We apply this system to inference of individual and cohort reproductive values in Soay sheep (Ovis aries) from St Kilda, Outer Hebrides. We observe that these genetic contributions appear to become relatively stable within modest time frames. As such, it may be reasonable to consider pedigree-based calculations of genetic contributions to future generations as estimates of reproductive value. This approach and the knowledge that the estimates can stabilize within decades should offer new opportunities to analyze data from pedigreed wild populations, which will be of value to many fields within evolutionary biology and demography.