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The examination of genetic structure in the deep-ocean hadal zone has focused on divergence between tectonic trenches to understand how environment and geography may drive species divergence and promote endemism. There has been little attempt to examine localized genetic structure within trenches, partly because of logistical challenges associated with sampling at an appropriate scale, and the large effective population sizes of species that can be sampled adequately may mask underlying genetic structure. Here we examine genetic structure in the superabundant amphipod Hirondellea gigas in the Mariana Trench at depths of 8126-10,545 m. RAD sequencing was used to identify 3182 loci containing 43,408 single nucleotide polymorphisms (SNPs) across individuals after stringent pruning of loci to prevent paralogous multicopy genomic regions being erroneously merged. Principal components analysis of SNP genotypes resolved no genetic structure between sampling locations, consistent with a signature of panmixia. However, discriminant analysis of principal components identified divergence between all sites driven by 301 outlier SNPs in 169 loci and significantly associated with latitude and depth. Functional annotation of loci identified differences between singleton loci used in analysis and paralogous loci pruned from the data set and also between outlier and nonoutlier loci, all consistent with hypotheses explaining the role of transposable elements driving genome dynamics. This study challenges the traditional perspective that highly abundant amphipods within a trench form a single panmictic population. We discuss the findings in relation to eco-evolutionary and ontogenetic processes operating in the deep sea, and highlight key challenges associated with population genetic analysis in nonmodel systems with inherent large effective population sizes and genomes., (© 2023 The Authors. Molecular Ecology published by John Wiley & Sons Ltd.)

The Qinghai-Tibetan Plateau, with low precipitation, low oxygen partial pressure, and temperatures routinely dropping below -30 °C in winter, presents several physiological challenges to its fauna. Yet it is home to many endemic mammalian species, including the plateau pika ( Ochotona curzoniae ). How these small animals that are incapable of hibernation survive the winter is an enigma. Measurements of daily energy expenditure (DEE) using the doubly labeled water method show that pikas suppress their DEE during winter. At the same body weight, pikas in winter expend 29.7% less than in summer, despite ambient temperatures being approximately 25 °C lower. Combined with resting metabolic rates (RMRs), this gives them an exceptionally low metabolic scope in winter (DEE/RMRt = 1.60 ± 0.30; RMRt is resting metabolic rate at thermoneutrality). Using implanted body temperature loggers and filming in the wild, we show that this is achieved by reducing body temperature and physical activity. Thyroid hormone (T 3 and T 4 ) measurements indicate this metabolic suppression is probably mediated via the thyroid axis. Winter activity was lower at sites where domestic yak ( Bos grunniens ) densities were higher. Pikas supplement their food intake at these sites by eating yak feces, demonstrated by direct observation, identification of yak DNA in pika stomach contents, and greater convergence in the yak/pika microbiotas in winter. This interspecific coprophagy allows pikas to thrive where yak are abundant and partially explains why pika densities are higher where domestic yak, their supposed direct competitors for food, are more abundant., Competing Interests: The authors declare no competing interest.

Phenotypic plasticity is predicted to evolve in more variable environments, conferring an advantage on individual lifetime fitness. It is less clear what the potential consequences of that plasticity will have on ecological population dynamics. Here, we use an invertebrate model system to examine the effects of environmental variation (resource availability) on the evolution of phenotypic plasticity in two life history traits-age and size at maturation-in long-running, experimental density-dependent environments. Specifically, we then explore the feedback from evolution of life history plasticity to subsequent ecological dynamics in novel conditions. Plasticity in both traits initially declined in all microcosm environments, but then evolved increased plasticity for age-at-maturation, significantly so in more environmentally variable environments. We also demonstrate how plasticity affects ecological dynamics by creating founder populations of different plastic phenotypes into new microcosms that had either familiar or novel environments. Populations originating from periodically variable environments that had evolved greatest plasticity had lowest variability in population size when introduced to novel environments than those from constant or random environments. This suggests that while plasticity may be costly it can confer benefits by reducing the likelihood that offspring will experience low survival through competitive bottlenecks in variable environments. In this study, we demonstrate how plasticity evolves in response to environmental variation and can alter population dynamics-demonstrating an eco-evolutionary feedback loop in a complex animal moderated by plasticity in growth., Competing Interests: None declared., (© 2021 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

While there is now an established recognition of microplastic pollution in the oceans, and the detrimental effects this may have on marine animals, the ocean depth at which such contamination is ingested by organisms has still not been established. Here, we detect the presence of ingested microplastics in the hindguts of Lysianassoidea amphipod populations, in six deep ocean trenches from around the Pacific Rim (Japan, Izu-Bonin, Mariana, Kermadec, New Hebrides and the Peru-Chile trenches), at depths ranging from 7000 m to 10 890 m. This illustrates that microplastic contaminants occur in the very deepest reaches of the oceans. Over 72% of individuals examined (65 of 90) contained at least one microparticle. The number of microparticles ingested per individual across all trenches ranged from 1 to 8. The mean and standard error of microparticles varied per trench, from 0.9 ± 0.4 (New Hebrides Trench) to 3.3 ± 0.7 (Mariana Trench). A subsample of microfibres and fragments analysed using FTIR were found to be a collection of plastic and synthetic materials (Nylon, polyethylene, polyamide, polyvinyl alcohol, polyvinylchloride, often with inorganic filler material), semi-synthetic (rayon and lyocell) and natural fibre (ramie). Notwithstanding, this study reports the deepest record of microplastic ingestion, indicating that anthropogenic debris is bioavailable to organisms at some of the deepest locations in the Earth's oceans., Competing Interests: The authors declare no competing interests.

The increasingly recognised effects of microbiomes on the eco-evolutionary dynamics of their hosts are promoting a view of the "hologenome" as an integral host-symbiont evolutionary entity. For example, sex-ratio distorting reproductive parasites such as Wolbachia are well-studied pivotal drivers of invertebrate reproductive processes, and more recent work is highlighting novel effects of microbiome assemblages on host mating behaviour and developmental incompatibilities that underpin or reinforce reproductive isolation processes. However, examining the hologenome and its eco-evolutionary effects in natural populations is challenging because microbiome composition is considerably influenced by environmental factors. Here we illustrate these challenges in a sympatric species complex of intertidal isopods (Jaera albifrons spp.) with pervasive sex-ratio distortion and ecological and behavioural reproductive isolation mechanisms. We deep-sequence the bacterial 16S rRNA gene among males and females collected in spring and summer from two coasts in north-east Scotland, and examine microbiome composition with a particular focus on reproductive parasites. Microbiomes of all species were diverse (overall 3,317 unique sequences among 3.8 million reads) and comprised mainly Proteobacteria and Bacteroidetes taxa typical of the marine intertidal zone, in particular Vibrio spp. However, we found little evidence of the reproductive parasites Wolbachia, Rickettsia, Spiroplasma and Cardinium, suggesting alternative causes of sex-ratio distortion. Notwithstanding, a significant proportion of the variance in microbiome composition among samples was explained by sex (14.1 %), nested within geographic (26.9 %) and seasonal (39.6 %) variance components. The functional relevance of this sex signal was difficult to ascertain given the absence of reproductive parasites, the ephemeral nature of the species assemblages and substantial environmental variability. These results establish the Jaera albifrons species complex as an intriguing system for examining the effects of microbiomes on reproductive processes and speciation, and highlight the difficulties associated with snapshot assays of microbiome composition in dynamic and complex environments., Competing Interests: The authors have declared that no competing interests exist.

Selective pressure from pathogens is considered a key selective force driving the evolution of components of the immune system. Since single components of the immune system may interact with many pathogens, and single pathogens may be recognized by multiple components of the immune system, gaining a better understanding of the mechanisms of parasite-driven selection requires the study of multiple genes and pathogens. Toll-like receptors (TLRs) are a large gene family that code for antigen-presenting components of the innate immune response. In the present paper we characterize polymorphism and signatures of selection in seven TLRs in free-living bank voles Myodes glareolus. We report the first evidence of balancing selection in several TLR genes, supported by positive values of Fu and Li's D* in TLR2 and TLR5, and positive values of Tajima's D in LRR regions within TLR1 and TLR2. We further found significant associations between amino-acid alleles of TLR1 and TLR5 and susceptibility to infection with the blood pathogen Bartonella. Interestingly, selection patterns in TLRs presenting virus-derived motifs (TLR7 and TLR9) differed considerably from those interacting with bacterial PAMPs. In contrast to the highly variable TLRs presenting bacterial motifs, TLR7 and TLR9 had low polymorphism and displayed signatures of directional selection. These findings suggest different functional responses across the TLR gene family and highlight the complexity of parasite-driven selection.

Genome size varies considerably across taxa, and extensive research effort has gone into understanding whether variation can be explained by differences in key ecological and life-history traits among species. The extreme environmental conditions that characterize the deep sea have been hypothesized to promote large genome sizes in eukaryotes. Here we test this supposition by examining genome sizes among 13 species of deep-sea amphipods from the Mariana, Kermadec and New Hebrides trenches. Genome sizes were estimated using flow cytometry and found to vary nine-fold, ranging from 4.06 pg (4.04 Gb) in Paralicella caperesca to 34.79 pg (34.02 Gb) in Alicella gigantea . Phylogenetic independent contrast analysis identified a relationship between genome size and maximum body size, though this was largely driven by those species that display size gigantism. There was a distinct shift in the genome size trait diversification rate in the supergiant amphipod A. gigantea relative to the rest of the group. The variation in genome size observed is striking and argues against genome size being driven by a common evolutionary history, ecological niche and life-history strategy in deep-sea amphipods., Competing Interests: We have no competing interests.

A population's effective size (N e ) is a key parameter that shapes rates of inbreeding and loss of genetic diversity, thereby influencing evolutionary processes and population viability. However, estimating N e , and identifying key demographic mechanisms that underlie the N e to census population size (N) ratio, remains challenging, especially for small populations with overlapping generations and substantial environmental and demographic stochasticity and hence dynamic age-structure. A sophisticated demographic method of estimating N e /N, which uses Fisher's reproductive value to account for dynamic age-structure, has been formulated. However, this method requires detailed individual- and population-level data on sex- and age-specific reproduction and survival, and has rarely been implemented. Here, we use the reproductive value method and detailed demographic data to estimate N e /N for a small and apparently isolated red-billed chough (Pyrrhocorax pyrrhocorax) population of high conservation concern. We additionally calculated two single-sample molecular genetic estimates of N e to corroborate the demographic estimate and examine evidence for unobserved immigration and gene flow. The demographic estimate of N e /N was 0.21, reflecting a high total demographic variance (σ2dg) of 0.71. Females and males made similar overall contributions to σ2dg. However, contributions varied among sex-age classes, with greater contributions from 3 year-old females than males, but greater contributions from ≥5 year-old males than females. The demographic estimate of N e was ~30, suggesting that rates of increase of inbreeding and loss of genetic variation per generation will be relatively high. Molecular genetic estimates of N e computed from linkage disequilibrium and approximate Bayesian computation were approximately 50 and 30, respectively, providing no evidence of substantial unobserved immigration which could bias demographic estimates of N e . Our analyses identify key sex-age classes contributing to demographic variance and thus decreasing N e /N in a small age-structured population inhabiting a variable environment. They thereby demonstrate how assessments of N e can incorporate stochastic sex- and age-specific demography and elucidate key demographic processes affecting a population's evolutionary trajectory and viability. Furthermore, our analyses show that N e for the focal chough population is critically small, implying that management to re-establish genetic connectivity may be required to ensure population viability., (© 2017 The Authors. Journal of Animal Ecology © 2017 British Ecological Society.)

The legacy and reach of anthropogenic influence is most clearly evidenced by its impact on the most remote and inaccessible habitats on Earth. Here we identify extraordinary levels of persistent organic pollutants in the endemic amphipod fauna from two of the deepest ocean trenches (>10,000 metres). Contaminant levels were considerably higher than documented for nearby regions of heavy industrialization, indicating bioaccumulation of anthropogenic contamination and inferring that these pollutants are pervasive across the world's oceans and to full ocean depth.

Deleterious recessive alleles that are masked in outbred populations are predicted to be expressed in small, inbred populations, reducing both individual fitness and population viability. However, there are few definitive examples of phenotypic expression of lethal recessive alleles under inbreeding conditions in wild populations. Studies that demonstrate the action of such alleles, and infer their distribution and dynamics, are required to understand their potential impact on population viability and inform management responses. The Scottish population of red-billed choughs (Pyrrhocorax pyrrhocorax), which currently totals <60 breeding pairs and is of major conservation concern, has recently been affected by lethal blindness in nestlings. We used family data to show that the pattern of occurrence of blindness within and across affected families that produced blind nestlings was exactly 0·25, matching that expected given a single-locus autosomal lethal recessive allele. Furthermore, the observed distribution of blind nestlings within affected families did not differ from that expected given Mendelian inheritance of such an allele. Relatedness estimates showed that individuals from affected families were not more closely related to each other than they were to individuals from unaffected families that did not produce blind nestlings. Blind individuals tended to be less heterozygous than non-blind individuals, as expected if blindness was caused by the expression of a recessive allele under inbreeding. However, there was no difference in the variance in heterozygosity estimates, suggesting that some blind individuals were relatively outbred. These results suggest carriers of the blindness allele may be widely distributed across contemporary families rather than restricted to a single family lineage, implying that the allele has persisted across multiple generations. Blindness occurred at low frequency (affecting 1·6% of observed nestlings since 1981). However, affected families had larger initial brood sizes than unaffected families. Such high fecundity of carriers of a lethal recessive allele might reflect overdominance, potentially reducing purging and increasing allele persistence probability. We thereby demonstrate the phenotypic expression of a lethal recessive allele in a wild population of conservation concern, and provide a general framework for inferring allele distribution and persistence and informing management responses., (© 2016 The Authors. Journal of Animal Ecology published by John Wiley & Sons Ltd on behalf of British Ecological Society.)

Here, we describe the development of 16 polymorphic microsatellite markers using an Illumina MiSeq sequencing approach in the deep-sea amphipod Paralicella tenuipes A total of 25 577 844 DNA sequences were filtered for microsatellite motifs of which 197 873 sequences were identified. From these sequences, 64 had sufficient flanking regions for primer design and 16 of these loci were polymorphic. Between 5 and 30 alleles were detected per locus, with an average of 13.63 alleles per locus, across a total of 120 individuals from 5 separate deep sea trenches from the Pacific Ocean. For the 16 loci, observed and expected heterozygosity values ranged from 0.116 to 0.414 and 0.422 to 0.820, respectively, with one locus displaying significant deviation from Hardy-Weinberg equilibrium. The microsatellite loci that have been isolated and described here are the first molecular markers developed for deep sea amphipods and will be invaluable for elucidating the genetic population structure and the extent of connectivity between deep ocean trenches., (© The American Genetic Association. 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The interaction between environmental variation and population dynamics is of major importance, particularly for managed and economically important species, and especially given contemporary changes in climate variability. Recent analyses of exploited animal populations contested whether exploitation or environmental variation has the greatest influence on the stability of population dynamics, with consequences for variation in yield and extinction risk. Theoretical studies however have shown that harvesting can increase or decrease population variability depending on environmental variation, and requested controlled empirical studies to test predictions. Here, we use an invertebrate model species in experimental microcosms to explore the interaction between selective harvesting and environmental variation in food availability in affecting the variability of stage-structured animal populations over 20 generations. In a constant food environment, harvesting adults had negligible impact on population variability or population size, but in the variable food environments, harvesting adults increased population variability and reduced its size. The impact of harvesting on population variability differed between proportional and threshold harvesting, between randomly and periodically varying environments, and at different points of the time series. Our study suggests that predicting the responses to selective harvesting is sensitive to the demographic structures and processes that emerge in environments with different patterns of environmental variation.

Landscape genomics promises to provide novel insights into how neutral and adaptive processes shape genome-wide variation within and among populations. However, there has been little emphasis on examining whether individual-based phenotype-genotype relationships derived from approaches such as genome-wide association (GWAS) manifest themselves as a population-level signature of selection in a landscape context. The two may prove irreconcilable as individual-level patterns become diluted by high levels of gene flow and complex phenotypic or environmental heterogeneity. We illustrate this issue with a case study that examines the role of the highly prevalent gastrointestinal nematode Trichostrongylus tenuis in shaping genomic signatures of selection in red grouse (Lagopus lagopus scotica). Individual-level GWAS involving 384 SNPs has previously identified five SNPs that explain variation in T. tenuis burden. Here, we examine whether these same SNPs display population-level relationships between T. tenuis burden and genetic structure across a small-scale landscape of 21 sites with heterogeneous parasite pressure. Moreover, we identify adaptive SNPs showing signatures of directional selection using F(ST) outlier analysis and relate population- and individual-level patterns of multilocus neutral and adaptive genetic structure to T. tenuis burden. The five candidate SNPs for parasite-driven selection were neither associated with T. tenuis burden on a population level, nor under directional selection. Similarly, there was no evidence of parasite-driven selection in SNPs identified as candidates for directional selection. We discuss these results in the context of red grouse ecology and highlight the broader consequences for the utility of landscape genomics approaches for identifying signatures of selection., (© 2015 John Wiley & Sons Ltd.)

Attempts to mitigate the impact of invasive species on native ecosystems increasingly target large land masses where control, rather than eradication, is the management objective. Depressing numbers of invasive species to a level where their impact on native biodiversity is tolerable requires overcoming the impact of compensatory immigration from non-controlled portions of the landscape. Because of the expected scale-dependency of dispersal, the overall size of invasive species management areas relative to the dispersal ability of the controlled species will determine the size of any effectively conserved core area unaffected by immigration from surrounding areas. However, when dispersal is male-biased, as in many mammalian invasive carnivores, males may be overrepresented amongst immigrants, reducing the potential growth rate of invasive species populations in re-invaded areas. Using data collected from a project that gradually imposed spatially comprehensive control on invasive American mink ( Neovison vison ) over a 10,000 km 2 area of NE Scotland, we show that mink captures were reduced to almost zero in 3 years, whilst there was a threefold increase in the proportion of male immigrants. Dispersal was often long distance and linking adjacent river catchments, asymptoting at 38 and 31 km for males and females respectively. Breeding and dispersal were spatially heterogeneous, with 40 % of river sections accounting for most captures of juvenile (85 %), adult female (65 %) and immigrant (57 %) mink. Concentrating control effort on such areas, so as to turn them into "attractive dispersal sinks" could make a disproportionate contribution to the management of recurrent re-invasion of mainland invasive species management areas., (© The Author(s) 2016.)

Identifying the genetic architecture underlying complex phenotypes is a notoriously difficult problem that often impedes progress in understanding adaptive eco-evolutionary processes in natural populations. Host-parasite interactions are fundamentally important drivers of evolutionary processes, but a lack of understanding of the genes involved in the host's response to chronic parasite insult makes it particularly difficult to understand the mechanisms of host life history trade-offs and the adaptive dynamics involved. Here, we examine the genetic basis of gastrointestinal nematode (Trichostrongylus tenuis) burden in 695 red grouse (Lagopus lagopus scotica) individuals genotyped at 384 genome-wide SNPs. We first use genome-wide association to identify individual SNPs associated with nematode burden. We then partition genome-wide heritability to identify chromosomes with greater heritability than expected from gene content, due to harbouring a multitude of additive SNPs with individually undetectable effects. We identified five SNPs on five chromosomes that accounted for differences of up to 556 worms per bird, but together explained at best 4.9% of the phenotypic variance. These SNPs were closely linked to genes representing a range of physiological processes including the immune system, protein degradation and energy metabolism. Genome partitioning indicated genome-wide heritability of up to 29% and three chromosomes with excess heritability of up to 4.3% (total 8.9%). These results implicate SNPs and novel genomic regions underlying nematode burden in this system and suggest that this phenotype is somewhere between being based on few large-effect genes (oligogenic) and based on a large number of genes with small individual but large combined effects (polygenic)., (© 2015 John Wiley & Sons Ltd.)

The extent to which genotypic variation at a priori identified candidate genes can explain variation in complex phenotypes is a major debate in evolutionary biology. Whereas some high-profile genes such as the MHC or MC1R clearly do account for variation in ecologically relevant characters, many complex phenotypes such as response to parasite infection may well be underpinned by a large number of genes, each of small and effectively undetectable effect. Here, we characterize a suite of novel candidate genes for variation in gastrointestinal nematode (Trichostrongylus tenuis) burden among red grouse (Lagopus lagopus scotica) individuals across a network of moors in north-east Scotland. We test for associations between parasite load and genotypic variation in twelve genes previously identified to be differentially expressed in experimentally infected red grouse or genetically differentiated among red grouse populations with overall different parasite loads. These genes are associated with a broad physiological response including immune system processes. Based on individual-level generalized linear models, genotypic variants in nine genes were significantly associated with parasite load, with effect sizes accounting for differences of 514-666 worms per bird. All but one of these variants were synonymous or untranslated, suggesting that these may be linked to protein-coding variants or affect regulatory processes. In contrast, population-level analyses revealed few and inconsistent associations with parasite load, and little evidence of signatures of natural selection. We discuss the broader significance of these contrasting results in the context of the utility of population genomics and landscape genomics approaches in detecting adaptive genomic signatures., (© 2015 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2015 European Society For Evolutionary Biology.)

Epigenetic modification of cytosine methylation states can be elicited by environmental stresses and may be a key process affecting phenotypic plasticity and adaptation. Parasites are potent stressors with profound physiological and ecological effects on their host, but there is little understanding in how parasites may influence host methylation states. Here, we estimate epigenetic diversity and differentiation among 21 populations of red grouse (Lagopus lagopus scotica) in north-east Scotland and test for association of gastrointestinal parasite load (caecal nematode Trichostrongylus tenuis) with hepatic genome-wide and locus-specific methylation states. Following methylation-sensitive AFLP (MSAP), 129 bands, representing 73 methylation-susceptible and 56 nonmethylated epiloci, were scored across 234 individuals. The populations differed significantly in genome-wide methylation levels and were also significantly epigenetically (F(SC) = 0.0227; P < 0.001) and genetically (F(SC) = 0.0058; P < 0.001) differentiated. Parasite load was not associated with either genome-wide methylation levels or epigenetic differentiation. Instead, we found eight disproportionately differentiated epilocus-specific methylation states (F(ST) outliers) using bayescan software and significant positive and negative association of 35 methylation states with parasite load from bespoke generalized estimating equations (GEE), simple logistic regression (sam) and Bayesian environmental analysis (bayenv2). Following Sanger sequencing, genome mapping and geneontology (go) annotation, some of these epiloci were linked to genes involved in regulation of cell cycle, signalling, metabolism, immune system and notably rRNA methylation, histone acetylation and small RNAs. These findings demonstrate an epigenetic signature of parasite load in populations of a wild bird and suggest intriguing physiological effects of parasite-associated cytosine methylation., (© 2014 John Wiley & Sons Ltd.)

A perceived consequence of a population bottleneck is the erosion of genetic diversity and concomitant reduction in individual fitness and evolutionary potential. Although reduced genetic variation associated with demographic perturbation has been amply demonstrated for neutral molecular markers, the effective management of genetic resources in natural populations is hindered by a lack of understanding of how adaptive genetic variation will respond to population fluctuations, given these are affected by selection as well as drift. Here, we demonstrate that selection counters drift to maintain polymorphism at a major histocompatibility complex (MHC) locus through a population bottleneck in an inbred island population of water voles. Before and after the bottleneck, MHC allele frequencies were close to balancing selection equilibrium but became skewed by drift when the population size was critically low. MHC heterozygosity generally conformed to Hardy-Weinberg expectations except in one generation during the population recovery where there was a significant excess of heterozygous genotypes, which simulations ascribed to strong differential MHC-dependent survival. Low allelic diversity and highly skewed frequency distributions at microsatellite loci indicated potent genetic drift due to a strong founder affect and/or previous population bottlenecks. This study is a real-time examination of the predictions of fundamental evolutionary theory in low genetic diversity situations. The findings highlight that conservation efforts to maintain the genetic health and evolutionary potential of natural populations should consider the genetic basis for fitness-related traits, and how such adaptive genetic diversity will vary in response to both the demographic fluctuations and the effects of selection.

Parasitic nematodes are found in almost all wild vertebrate populations but few studies have investigated these host-parasite relationships in the wild. For parasites with free-living stages, the external environment has a major influence on life-history traits, and development and survival is generally low at sub-zero temperatures. For reindeer that inhabit the high Arctic archipelago of Svalbard, parasite transmission is expected to occur in the summer, due to the extreme environmental conditions and the reduced food intake by the host in winter. Here we show experimentally that, contrary to most parasitic nematodes, Marshallagia marshalli of Svalbard reindeer is transmitted during the Arctic winter. Winter transmission was demonstrated by removing parasites in the autumn, using a novel delayed-release anthelmintic bolus, and estimating re-infection rates in reindeer sampled in October, February and April. Larval stages of nematodes were identified using molecular tools, whereas adult stages were identified using microscopy. The abundance of M. marshalli adult worms and L4s increased significantly from October to April, indicating that reindeer were being infected with L3s from the pasture throughout the winter. To our knowledge, this study is the first to experimentally demonstrate over-winter transmission of a gastro-intestinal nematode parasite in a wild animal. Potential mechanisms associated with this unusual transmission strategy are discussed in light of our knowledge of the life-history traits of this parasite., (Copyright © 2012 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.)

Culicoides biting midges (Diptera: Ceratopogonidae) vector a wide variety of internationally important arboviral pathogens of livestock and represent a widespread biting nuisance. This study investigated the influence of landscape, host and remotely-sensed climate factors on local abundance of livestock-associated species in Scotland, within a hierarchical generalized linear model framework. The Culicoides obsoletus group and the Culicoides pulicaris group accounted for 56% and 41%, respectively, of adult females trapped. Culicoides impunctatus Goetghebuer and C. pulicaris s.s. Linnaeus were the most abundant and widespread species in the C. pulicaris group (accounting for 29% and 10%, respectively, of females trapped). Abundance models performed well for C. impunctatus, Culicoides deltus Edwards and Culicoides punctatus Meigen (adjusted R(2) : 0.59-0.70), but not for C. pulicaris s.s. (adjusted R(2) : 0.36) and the C. obsoletus group (adjusted R(2) : 0.08). Local-scale abundance patterns were best explained by models combining host, landscape and climate factors. The abundance of C. impunctatus was negatively associated with cattle density, but positively associated with pasture cover, consistent with this species' preference in the larval stage for lightly grazed, wet rush pasture. Predicted abundances of this species varied widely among farms even over short distances (less than a few km). Modelling approaches that may facilitate the more accurate prediction of local abundance patterns for a wider range of Culicoides species are discussed., (© 2011 The Authors. Medical and Veterinary Entomology © 2011 The Royal Entomological Society.)

The identification of genes involved in a host's response to parasite infection provides both a means for understanding the pathways involved in immune defence and a target for examining host-parasite co-evolution. Most studies rely on a candidate gene approach derived from model systems to identify gene targets of interest, and there have been a dearth of studies geared towards providing a holistic overview of immune response from natural populations. We carried out an experiment in a natural population of red grouse (Lagopus lagopus scoticus) to manipulate levels of Trichostrongylus tenuis parasite infection. The transcriptomic response of individuals was examined from standard cDNA and suppressive subtractive hybridization (SSH) libraries produced from gut, liver and spleen, enriching for genes expressed in response to T. tenuis infection. A total of 2209 and 3716 unique transcript sequences were identified from the cDNA and SSH libraries, respectively. Forty-five of these had Gene Ontology annotation associated with immune response. Some of these genes have previously been reported from laboratory-based studies of model species as important in immune response to gastrointestinal parasite infection; however, multiple novel genes were also identified. These may reveal novel pathways involved in the host response of grouse to T. tenuis and provide a resource that can be utilized as candidate genes in other species. All sequences described have been deposited in GenBank (accession numbers GW698221-GW706922), (© 2010 Blackwell Publishing Ltd.)

Spawning patterns in female brown trout Salmo trutta were examined by documenting the construction of nests in a small stream and later excavating them to recover progeny. The maternal provenance of nests was determined by genetic typing of embryos using microsatellite markers. Seventy-two nests, for which position and date of construction were known, were made by 59 individuals. Position and date of construction were known for a further 35 nests, comprising 11 Atlantic salmon Salmo salar nests and 24 nests which contained few or no progeny. Salmo trutta showed a behavioural preference for spawning near (≤ 1 m) prior nests; nests made by different individuals tended to accumulate in a spatial sequence that progressed upstream. The directionality of the association between prior and new nests suggests that later spawners use the residual depressions created by previous spawners as the first element of their own nests., (© 2011 Crown Marine Scotland. Journal of Fish Biology © 2011 The Fisheries Society of the British Isles.)

A central issue in ecology is in understanding the relative influences of intrinsic and extrinsic effects on population regulation. Previous studies on the cyclic population dynamics of red grouse (Lagopus lagopus scoticus) have emphasized the destabilizing effects of either nematode parasites or territorial behaviour and aggression. The potential interacting effects of these processes, mediated through density-dependent, environmentally induced alterations of host immunocompetence influencing susceptibility to parasites have not been considered. Male red grouse at high density are more aggressive, associated with increased testosterone, which potentially could lead to reduced immunocompetence at a stage when parasites are most prevalent. This could depress individual condition, breeding performance and survival and thus drive or contribute to overall reductions in population size. Here, we characterize the transcriptomic response of grouse to nematode parasite infection and investigate how this is subsequently affected by testosterone, using a microarray approach contrasting red grouse with high and low parasite load at both high and low testosterone titre. A suite of 52 transcripts showed a significant level of up-regulation to either chronic parasite load or experimental parasite infection. Of these, 51 (98%) showed a reduced level of expression under conditions of high parasite load and high testosterone. The genes up-regulated by parasites and then down-regulated at high testosterone titre were not necessarily associated with immune response, as might be intuitively expected. The results are discussed in relation to the fitness and condition of individual red grouse and factors influencing the regulation of abundance in natural populations., (© 2010 Blackwell Publishing Ltd.)

Population genetic structure of North Atlantic killer whale samples was resolved from differences in allele frequencies of 17 microsatellite loci, mtDNA control region haplotype frequencies and for a subset of samples, using complete mitogenome sequences. Three significantly differentiated populations were identified. Differentiation based on microsatellite allele frequencies was greater between the two allopatric populations than between the two pairs of partially sympatric populations. Spatial clustering of individuals within each of these populations overlaps with the distribution of particular prey resources: herring, mackerel and tuna, which each population has been seen predating. Phylogenetic analyses using complete mitogenomes suggested two populations could have resulted from single founding events and subsequent matrilineal expansion. The third population, which was sampled at lower latitudes and lower density, consisted of maternal lineages from three highly divergent clades. Pairwise population differentiation was greater for estimates based on mtDNA control region haplotype frequencies than for estimates based on microsatellite allele frequencies, and there were no mitogenome haplotypes shared among populations. This suggests low or no female migration and that gene flow was primarily male mediated when populations spatially and temporally overlap. These results demonstrate that genetic differentiation can arise through resource specialization in the absence of physical barriers to gene flow., (© 2010 Blackwell Publishing Ltd.)

Testosterone underlies the expression of most secondary sexual traits, playing a key role in sexual selection. However, high levels might be associated with physiological costs, such as immunosuppression. Immunostimulant carotenoids underpin the expression of many red-yellow ornaments, but are regulated by testosterone and constrained by parasites. We manipulated testosterone and nematode burdens in red grouse (Lagopus lagopus scoticus) in two populations to tease apart their effects on carotenoid levels, ornament size and colouration in three time-step periods. We found no evidence for interactive effects of testosterone and parasites on ornament size and colouration. We showed that ornament colouration was testosterone-driven. However, parasites decreased comb size with a time delay and testosterone increased carotenoid levels in one of the populations. This suggests that environmental context plays a key role in determining how individuals resolve the trade-off between allocating carotenoids for ornamental coloration or for self-maintenance needs. Our study advocates that adequately testing the mechanisms behind the production or maintenance of secondary sexual characters has to take into account the dynamics of sexual trait expression and their environmental context.

Over the past two decades the fields of molecular ecology and population genetics have been dominated by the use of putatively neutral DNA markers, primarily to resolve spatio-temporal patterns of genetic variation to inform our understanding of population structure, gene flow and pedigree. Recent emphasis in comparative functional genomics, however, has fuelled a resurgence of interest in functionally important genetic variation that underpins phenotypic traits of adaptive or ecological significance. It may prove a major challenge to transfer genomics information from classical model species to examine functional diversity in non-model species in natural populations, but already multiple gene-targeted candidate loci with major effect on phenotype and fitness have been identified. Here we briefly describe some of the research strategies used for isolating and characterising functional genetic diversity at candidate gene-targeted loci, and illustrate the efficacy of some of these approaches using our own studies on red grouse (Lagopus lagopus scoticus). We then review how candidate gene markers have been used to: (1) quantify genetic diversity among populations to identify those depauperate in genetic diversity and requiring specific management action; (2) identify the strength and mode of selection operating on individuals within natural populations; and (3) understand direct mechanistic links between allelic variation at single genes and variance in individual fitness.

Vertebrates commonly use carotenoid-based traits as social signals. These can reliably advertise current nutritional status and health because carotenoids must be acquired through the diet and their allocation to ornaments is traded-off against other self-maintenance needs. We propose that the coloration more generally reveals an individual's ability to cope with stressful conditions. We tested this idea by manipulating the nematode parasite infection in free-living red grouse (Lagopus lagopus scoticus) and examining the effects on body mass, carotenoid-based coloration of a main social signal and the amount of corticosterone deposited in feathers grown during the experiment. We show that parasites increase stress and reduce carotenoid-based coloration, and that the impact of parasites on coloration was associated with changes in corticosterone, more than changes in body mass. Carotenoid-based coloration appears linked to physiological stress and could therefore reveal an individual's ability to cope with stressors.

Oxidative stress, the physiological condition whereby the production of reactive oxygen and nitrogen species overwhelms the capacity of antioxidant defences, causes damage to key bio-molecules. It has been implicated in many diseases, and is proposed as a reliable currency in the trade-off between individual health and ornamentation. Whether oxidative stress mediates the expression of carotenoid-based signals, which are among the commonest signals of many birds, fish and reptiles, remains controversial. In the present study, we explored interactions between parasites, oxidative stress and the carotenoid-based ornamentation of red grouse Lagopus lagopus scoticus. We tested whether removing nematode parasites influenced both oxidative balance (levels of oxidative damage and circulating antioxidant defences) and carotenoid-based ornamentation. At the treatment group level, parasite purging enhanced the size and colouration of ornaments but did not significantly affect circulating carotenoids, antioxidant defences or oxidative damage. However, relative changes in these traits among individuals indicated that males with a greater number of parasites prior to treatment (parasite purging) showed a greater increase in the levels of circulating carotenoids and antioxidants, and a greater decrease in oxidative damage, than those with initially fewer parasites. At the individual level, a greater increase in carotenoid pigmentation was associated with a greater reduction in oxidative damage. Therefore, an individual's ability to express a carotenoid-based ornament appeared to be linked to its current oxidative balance and susceptibility to oxidative stress. Our experimental results suggest that oxidative stress can mediate the impact of parasites on carotenoid-based signals, and we discuss possible mechanisms linking carotenoid-based ornaments to oxidative stress.

Ecological divergence has a central role in speciation and is therefore an important source of biodiversity. Studying the micro-evolutionary processes of ecological diversification at its early stages provides an opportunity for investigating the causative mechanisms and ecological conditions promoting divergence. Here we use morphological traits, nitrogen stable isotope ratios and tooth wear to characterize two disparate types of North Atlantic killer whale. We find a highly specialist type, which reaches up to 8.5 m in length and a generalist type which reaches up to 6.6 m in length. There is a single fixed genetic difference in the mtDNA control region between these types, indicating integrity of groupings and a shallow divergence. Phylogenetic analysis indicates this divergence is independent of similar ecological divergences in the Pacific and Antarctic. Niche-width in the generalist type is more strongly influenced by between-individual variation rather than within-individual variation in the composition of the diet. This first step to divergent specialization on different ecological resources provides a rare example of the ecological conditions at the early stages of adaptive radiation.

Analysis of DNA sequence data has proven invaluable for defining the relationships among taxa, as well as resolving their evolutionary histories. Here, we analyzed DNA sequence variation of one mitochondrial gene (COI) and two nuclear regions (ITSI and II) to clarify the phylogenetic position of Culicoides dewulfi, a midge species widely spread in Europe and a suspected vector for bluetongue virus. Various authors have described C. dewulfi either as part of the Culicoides obsoletus sensu lato complex or as a separate taxonomic group. A maximum likelihood phylogeny, based upon an optimal model of sequence evolution, placed C. dewulfi outwith the C. obsoletus s.l. complex. Shimodaira-Hasegawa test highlighted that this topology was significantly more likely than any topology that placed C. dewulfi anywhere else in the phylogeny. As such, C. dewulfi should not be considered part of the C. obsoletus s.l. complex and instead be treated as a separate group, phylogenetically close to the classical Old World vector C. imicola.

The recent spread of Bluetongue disease in northwestern Europe has indicated the ability of Palaearctic Culicoides species to vector the disease. Because the different midge species vary in their ability to harbor and transmit the Bluetongue virus, quick and reliable identification is necessary to resolve the species composition of midge communities, both adult and larval, at any place at any given time point. Given that morphological identification of Culicoides species is problematic, we developed three multiplex polymerase chain reaction (PCR) assays that facilitate high-throughput analysis of midge specimens. One assay distinguishes between species of the so-called Culicoides obsoletus s.l. complex (including C. dewulfi), whereas two assays facilitate differentiation of species of the Culicoides pulicaris s.l. complex. These assays yield two PCR products: one species-specific and one generic band. We show the application of the assays in the analysis of Culicoides larvae from three different farms in northeast Scotland.

To be effective, management programmes geared towards halting or reversing the spread of invasive species must focus on defined and defensible areas. This requires knowledge of the dispersal of non-native species targeted for control to better understand invasion and recolonisation scenarios. We investigated the genetic structure of invasive American mink (Neovison vison) in Scotland, and incorporated landscape genetic approaches to examine resultant patterns in relation to geographical features that may influence dispersal. Populations of mink sampled from 10 sites in two regions (Argyll and Northeast Scotland) show a distinct genetic structure. First, the majority of pairwise population comparisons yielded F(ST) values that were significantly greater than zero. Second, AMOVA revealed that most of the genetic variance was attributable to differences among regions. Assignment tests placed 89 or more of individuals into their sampled region. Bayesian clustering methods grouped samples into two clusters according to their region of origin. Wombling approach identified the Cairngorms Mountains as a major impediment to gene flow between the regions. Mantel pairwise correlations between genetic and geographical distances estimated as least-cost distance assuming a linear increase in the cost of movement with increasing elevation were higher than Euclidean distances or distance along waterways. Spatial autocorrelation analyses revealed stronger spatial structuring for females than for males. These results suggest that gene flow by American mink is restricted by landscape features (mountain ranges) and that eradication attempt should in the first instance break down the connectivity between management units separated by mountains.

The fundamental role of the major histocompatibility complex (MHC) in immune recognition has led to a general consensus that the characteristically high levels of functional polymorphism at MHC genes is maintained by balancing selection operating through host-parasite coevolution. However, the actual mechanism by which selection operates is unclear. Two hypotheses have been proposed: overdominance (or heterozygote superiority) and negative frequency-dependent selection. Evidence for these hypotheses was evaluated by examining MHC-parasite relationships in an island population of water voles (Arvicola terrestris). Generalized linear mixed models were used to examine whether individual variation at an MHC class II DRB locus explained variation in the individual burdens of five different parasites. MHC genotype explained a significant amount of variation in the burden of gamasid mites, fleas (Megabothris walkeri) and nymphs of sheep ticks (Ixodes ricinus). Additionally, MHC heterozygotes were simultaneously co-infected by fewer parasite types than homozygotes. In each case where an MHC-dependent effect on parasite burden was resolved, the heterozygote genotype was associated with fewer parasites, and the heterozygote outperformed each homozygote in two of three cases, suggesting an overall superiority against parasitism for MHC heterozygote genotypes. This is the first demonstration of MHC heterozygote superiority against multiple parasites in a natural population, a mechanism that could help maintain high levels of functional MHC genetic diversity in natural populations.

Extravagant ornaments evolved to advertise their bearers' quality, the honesty of the signal being ensured by the cost paid to produce or maintain it. The oxidation handicap hypothesis (OHH) proposes that a main cost of testosterone-dependent ornamentation is oxidative stress, a condition whereby the production of reactive oxygen and nitrogen species (ROS/RNS) overwhelms the capacity of antioxidant defences. ROS/RNS are unstable, very reactive by-products of normal metabolic processes that can cause extensive damage to key biomolecules (cellular proteins, lipids and DNA). Oxidative stress has been implicated in the aetiology of many diseases and could link ornamentation and genetic variation in fitness-related traits. We tested the OHH in a free-living bird, the red grouse. We show that elevated testosterone enhanced ornamentation and increased circulating antioxidant levels, but caused oxidative damage. Males with smaller ornaments suffered more oxidative damage than those with larger ornaments when forced to increase testosterone levels, consistent with a handicap mechanism. Parasites depleted antioxidant defences, caused oxidative damage and reduced ornament expression. Oxidative damage extent and the ability of males to increase antioxidant defences also explained the impacts of testosterone and parasites on ornamentation within treatment groups. Because oxidative stress is intimately linked to immune function, parasite resistance and fitness, it provides a reliable currency in the trade-off between individual health and ornamentation. The costs induced by oxidative stress can apply to a wide range of signals, which are testosterone-dependent or coloured by pigments with antioxidant properties.

Patterns of spatio-temporal genetic variation at a class II major histocompatibility complex (MHC) locus and multiple microsatellite loci were analysed within and between three water vole metapopulations in Scotland, UK. Comparisons of MHC and microsatellite spatial genetic differentiation, based on standardised tests between two demographically asynchronous zones within a metapopulation, suggested that spatial MHC variation was affected by balancing selection, directional selection and random genetic drift, but that the relative effects of these microevolutionary forces vary temporally. At the metapopulation level, between-year differentiation for MHC loci was significantly correlated with that of microsatellites, signifying that neutral factors such as migration and drift were primarily responsible for overall temporal genetic change at the metapopulation scale. Between metapopulations, patterns of genetic differentiation implied that, at large spatial scales, MHC variation was primarily affected by directional selection and drift. Levels of MHC heterozygosity in excess of Hardy-Weinberg expectations were consistent with overdominant balancing selection operating on MHC variation within metapopulations. However, this effect was not constant among all samples, indicating temporal variation in the strength of selection relative to other factors. The results highlight the benefit of contrasting variation at MHC with neutral markers to separate the effects of stochastic and deterministic microevolutionary forces, and add to a growing body of evidence showing that the mode and relative strength of selection acting on MHC diversity varies both spatially and temporally.

Background: Extravagant ornaments used as social signals evolved to advertise their bearers' quality. The Immunocompetence Handicap Hypothesis proposes that testosterone-dependent ornaments reliably signal health and parasite resistance; however, empirical studies have shown mixed support. Alternatively, immune function and parasite resistance may be indirectly or directly related to glucocorticoid stress hormones. We propose that an understanding of the interplay between the individual and its environment, particularly how they cope with stressors, is crucial for understanding the honesty of social signals., Methodology/principal Findings: We analyzed corticosterone deposited in growing feathers as an integrated measure of hypothalamic-pituitary-adrenal activity in a wild territorial bird, the red grouse Lagopus lagopus scoticus. We manipulated two key, interrelated components, parasites and testosterone, which influence both ornamentation and fitness. Birds were initially purged of parasites, and later challenged with parasites or not, while at the same time being given testosterone or control implants, using a factorial experimental design. At the treatment level, testosterone enhanced ornamentation, while parasites reduced it, but only in males not implanted with testosterone. Among individuals, the degree to which both parasites and testosterone had an effect was strongly dependent on the amount of corticosterone in the feather grown during the experiment. The more stressors birds had experienced (i.e., higher corticosterone), the more parasites developed, and the less testosterone enhanced ornamentation., Conclusions/significance: With this unique focus on the individual, and a novel, integrative, measure of response to stressors, we show that ornamentation is ultimately a product of the cumulative physiological response to environmental challenges. These findings lead toward a more realistic concept of honesty in signaling as well as a broader discussion of the concept of stress.

The bluetongue (BT) vector Culicoides imicola Kieffer (Diptera: Ceratopogonidae) has undergone widespread range expansion across most of the Mediterranean basin, concomitant with the largest BT epizootic outbreaks on record. Knowledge of the substructure of this vector expansion would be useful for identifying specific source-expansion systems. To this end we analysed the haplotype diversity of the mitochondrial cytochrome oxidase I gene in 273 C. imicola from 88 Mediterranean sites and outgroups. All the C. imicola haplotypes (n = 26) formed a single, distinct clade in comparison with haplotypes of four other species of the Imicola group from southern Africa, confirming C. imicola as a single phylospecies. Haplotype distribution showed extreme differentiation across the Mediterranean basin, with four common haplotypes each predominating in different areas. Eastern and western areas characterized by distinct BT incursions accounted for most of the molecular variance in haplotype composition. Shared common haplotypes identified one area of incursion and expansion encompassing the western half of the Mediterranean basin, with evidence of population growth, and another system encompassing Anatolian Turkey, the Aegean Islands and mainland Greece. A third area of range expansion was identified in the central Mediterranean, with a possible source in Algeria and unsampled parts of central North Africa. We conclude that the expansion of C. imicola in the Mediterranean basin consists of at least three incursions followed by expansions and that the western system experiences conditions promoting high population growth.

Daubenton's bat (Myotis daubentonii) is a known reservoir for European bat lyssavirus type 2 (EBLV-2). An appreciation of the potential for epidemiological spread and disease risk requires an understanding of the dispersal of the primary host, and any large-scale geographical barriers that may impede gene flow. The spatial pattern of microsatellite and mitochondrial DNA variation was examined to infer patterns of dispersal of bats among 35 populations across Scotland. DNA sequence variation at the mitochondrial control region and ND1 genes revealed two distinct phylogeographical clades, with generally nonoverlapping geographical distributions except for a small number of populations where both matrilines were found in sympatry. Such discontinuity suggests that Scotland was recolonized twice following the retreat of the Pleistocene ice sheet with little subsequent matrilineal introgression. However, eight microsatellite loci showed low levels of genetic divergence among populations, even between populations from the two distinct mitochondrial DNA clades. An overall, macrogeographical genetic isolation-by-distance pattern was observed, with high levels of gene flow among local populations. Apparently contrasting patterns of mitochondrial and microsatellite divergence at different scales could be explained by sex-specific differences in gene flow at large scales.

Populations of red grouse (Lagopus lagopus scoticus) undergo regular multiannual cycles in abundance. The 'kinship hypothesis' posits that such cycles are caused by changes in kin structure among territorial males producing delayed density-dependent changes in aggressiveness, which in turn influence recruitment and regulate density. The kinship hypothesis makes several specific predictions about the levels of kinship, aggressiveness and recruitment through a population cycle: (i) kin structure will build up during the increase phase of a cycle, but break down prior to peak density; (ii) kin structure influences aggressiveness, such that there will be a negative relationship between kinship and aggressiveness over the years; (iii) as aggressiveness regulates recruitment and density, there will be a negative relationship between aggressiveness in one year and both recruitment and density in the next; (iv) as kin structure influences recruitment via an affect on aggressiveness, there will be a positive relationship between kinship in one year and recruitment the next. Here we test these predictions through the course of an 8-year cycle in a natural population of red grouse in northeast Scotland, using microsatellite DNA markers to resolve changing patterns of kin structure, and supra-orbital comb height of grouse as an index of aggressiveness. Both kin structure and aggressiveness were dynamic through the course of the cycle, and changing patterns were entirely consistent with the expectations of the kinship hypothesis. Results are discussed in relation to potential drivers of population regulation and implications of dynamic kin structure for population genetics.

Biting midges of the Culicoides obsoletus Meigen and Culicoides pulicaris L. species complexes (Diptera: Ceratopogonidae) are increasingly implicated as vectors of bluetongue virus in Palearctic regions. However, predicting epidemiological risk and the spread of disease is hampered because whilst vector competence of Culicoides is expressed only in adult females, morphological identification of constituent species is only readily applicable to adult males and some species distinguishing traits have overlapping character states. Furthermore, adult males are typically rare in field collections, making characterisation of Culicoides communities impossible. Here we highlight the utility of mitochondrial cytochrome oxidase subunit I (COI) DNA sequences for taxonomic resolution and species identification of all species within C. obsoletus and C. pulicarus complexes. Culicoides were collected from 18 sites in the UK and Continental Europe, and identified to species level, or species complex level, based on morphological characters. The sample comprised four species from the C. obsoletus complex (n = 88) and five species from the C. pulicaris complex (n = 39). The DNA sequence of the 5' end of the COI gene was obtained from all individuals. Each member species formed a well-supported reciprocally monophyletic clade in a maximum likelihood phylogeny. Levels of DNA sequence divergence were sufficiently high between species to allow the design of species-specific PCR primers that can be used in PCR for identification of members of the C. pulicaris complex or in a multiplex PCR to identify members of the C. obsoletus complex. This approach provides a valuable diagnostic tool for monitoring species composition in mixed field collections of Culicoides.

We applied a phenotypic QST (PST) vs. FST approach to study spatial variation in selection among great snipe (Gallinago media) populations in two regions of northern Europe. Morphological divergence between regions was high despite low differentiation in selectively neutral genetic markers, whereas populations within regions showed very little neutral divergence and trait differentiation. QST > FST was robust against altering assumptions about the additive genetic proportions of variance components. The homogenizing effect of gene flow (or a short time available for neutral divergence) has apparently been effectively counterbalanced by differential natural selection, although one trait showed some evidence of being under uniform stabilizing selection. Neutral markers can hence be misleading for identifying evolutionary significant units, and adopting the PST-FST approach might therefore be valuable when common garden experiments is not an option. We discuss the statistical difficulties of documenting uniform selection as opposed to divergent selection, and the need for estimating measurement error. Instead of only comparing overall QST and FST values, we advocate the use of partial matrix permutation tests to analyse pairwise QST differences among populations, while statistically controlling for neutral differentiation.

In so-called model species, such as human and mouse, genes of the major histocompatibility complex (MHC) are characterized by extremely high levels of polymorphism, and it is considered that such diversity is maintained by balancing selection. ;There is now a recognized need to expand studies into nonmodel species to examine whether high MHC diversity is mirrored in natural populations, and to determine the ecological, ethological, and evolutionary processes that underpin balancing selection. To address such issues, a necessary prerequisite is the ability to characterize diversity at a single, expressed, polymorphic MHC locus on which selection may be acting. Here, we provide the first description of allelic diversity at exon 2 of an MHC class II DRB locus in the European water vole (Arvicola terrestris), characterize variation across four natural populations, and test whether the patterns of variation are consistent with the effects of balancing selection. Using single-strand conformation polymorphism analysis and subsequent DNA sequencing of gel excisions, five DRB alleles were resolved, each with a unique amino acid sequence, among 100 individuals from four geographically distinct populations. Reverse transcription polymerase chain reaction confirmed that the alleles were products from an expressed locus. Intra-allelic amino acid differences were high (10.5-33.3%), and the nonsynonymous substitution rate exceeded the synonymous substitution rate for the functional peptide-binding region (d (N):d (S)=3.91 and P<0.005). Phylogenetic comparison of resolved alleles with closely related homologues indicated that each allele represented a unique lineage preserved across speciation events. These results indicate that balancing selection has maintained diversity of DRB allelic lineages and amino acid function over evolutionary time scales, but may be less effective at preserving alleles in contemporary populations where stochastic microevolutionary processes may dominate.

Theory predicts that the impact of gene flow on the genetic structure of populations in patchy habitats depends on its scale and the demographic attributes of demes (e.g. local colony sizes and timing of reproduction), but empirical evidence is scarce. We inferred the impact of gene flow on genetic structure among populations of water voles Arvicola terrestris that differed in average colony sizes, population turnover and degree of patchiness. Colonies typically consisted of few reproducing adults and several juveniles. Twelve polymorphic microsatellite DNA loci were examined. Levels of individual genetic variability in all areas were high (H(O) = 0.69-0.78). Assignments of juveniles to parents revealed frequent dispersal over long distances. The populations showed negative F(IS) values among juveniles, F(IS) values around zero among adults, high F(ST) values among colonies for juveniles, and moderate, often insignificant, F(ST) values for parents. We inferred that excess heterozygosity within colonies reflected the few individuals dispersing from a large area to form discrete breeding colonies. Thus pre-breeding dispersal followed by rapid reproduction results in a seasonal increase in differentiation due to local family groups. Genetic variation was as high in low-density populations in patchy habitats as in populations in continuous habitats used for comparison. In contrast to most theoretical predictions, we found that populations living in patchy habitats can maintain high levels of genetic variability when only a few adults contribute to breeding in each colony, when the variance of reproductive success among colonies is likely to be low, and when dispersal between colonies exceeds nearest-neighbour distances.

The major histocompatibility complex (MHC) has become a paradigm for how selection can act to maintain adaptively important genetic diversity in natural populations. Here, we review the contribution of studies on the MHC in non-model species to our understanding of how selection affects MHC diversity, emphasising how ecological and ethological processes influence the tempo and mode of evolution at the MHC, and conversely, how variability at the MHC affects individual fitness, population dynamics and viability. We focus on three main areas: the types of information that have been used to detect the action of selection on MHC genes; the relative contributions of parasite-mediated and sexual selection on the maintenance of MHC diversity; and possible future lines of research that may help resolve some of the unanswered issues associated with MHC evolution.

Testosterone often mediates trade-offs between reproduction and other life-history traits, which are usually investigated using testosterone implants. However, this approach does not distinguish between the physiological and behavioral effects of testosterone. We studied a wild game bird, the red grouse Lagopus lagopus scoticus, and took a new approach to investigate mechanisms linking elevated testosterone to increased parasite intensity. We caught males in autumn, removed their parasites, implanted them with the antiandrogen flutamide in combination with an aromatase inhibitor (FA males) or with empty implants (control males), and challenged them with parasites. The FA treatment increased testosterone concentration and physiological stress, but without enhancing testosterone-dependent behaviors, because testosterone receptors were blocked. FA males ended up with more parasites than the control males the following autumn, an effect similar to that of a testosterone treatment reported elsewhere. However, and unlike the testosterone treatment, the FA treatment did not affect home range, pairing, or breeding success. The results supported a physiological mechanism (increased susceptibility) linking elevated testosterone and increased parasite intensity. The FA treatment provided a new way of investigating testosterone-mediated trade-offs whereby testosterone concentration was increased while the effects on behavior were blocked, resulting in physiological costs without phenotypic benefits.

The phylogeographic pattern of mitochondrial DNA variation in water voles (Arvicola terrestris) from 57 localities across the United Kingdom and representative samples from Spain, France, Switzerland and Finland was determined from sequence variation in the central portion of the control region. Twenty-seven different haplotypes were resolved which formed two distinct phylogenetic clades. This major division separated haplotypes found in Scotland from those found in England and Wales. Nested clade analysis of haplotypes indicated that such a division was a consequence of allopatric fragmentation. The haplotypes found in Switzerland, France and Spain clustered with Scottish haplotypes, whereas the haplotype from Finland clustered with the English/Welsh haplotypes. These patterns indicate that contemporary Scottish populations are derived from an Iberian glacial refugium, whereas English and Welsh populations are derived from an eastern European refugium. As such, the postglacial recolonization of the United Kingdom must have involved two colonization events, either in different localities with no subsequent contact, or as two waves separated over time, with the second wave of colonizers displacing the first. An analysis of molecular variance (AMOVA) identified significant population genetic divergence within both the major clades, indicative of restricted gene flow and regional population isolation. The implications of both phylogeographical and population genetic structure are discussed in context with the conservation of water voles in Britain.