Molecular Ecology

Thermal extremes are among the most immediate environmental challenges faced by animals. Coping with these conditions across life is complex because growth changes body size, heat exchange and thermoregulatory demands. Consequently, adaptive responses in young, small individuals may be maladaptive, or require adjustment, later in life. However, in endotherms we know very little about how thermoregulatory responses to hot and cold temperatures change during development, or about the molecular mechanisms regulating responses. We examined the molecular responses to acute heat (40{degrees}C), cold (12{degrees}C) and control (23{degrees}C) conditions in 1- and 8-week-old ostrich chicks (Struthio camelus), a rapidly growing species exposed to strong daily and seasonal temperature variation. We found that in 1-week-old chicks, 32% of temperature-related genes were involved in both heat and cold responses, indicating that responses to opposing temperatures involve overlapping molecular pathways. However, the response of 75% of these genes changed during development. For example, some genes that increased with heat when young decreased with heat later in development, and vice versa. Such opposing selection pressures may maintain genetic variation in thermoregulatory pathways. Consistent with this prediction, comparisons between ostrich subspecies adapted to different thermal environments revealed patterns of genomic variation compatible with balancing selection in differentially expressed genes. Our results show that hot and cold temperatures trigger overlapping molecular responses that change during development, which shape genetic variation in the thermoregulatory system.

Climate change is causing rapid increases in temperature which drives genomic changes tied to adaptation. However, predicting the outcomes of climate change presents challenges as the anticipated conditions have yet to be experienced by natural populations. Modelling and lab experiments suggest that natural populations will experience shifts in life history, physiology, phenology, and ecology, but the underlying genomic mechanisms involved are unknown. However, some contemporary natural populations experience habitat warming through geothermal activity and can provide valuable insights into evolutionary responses. Geothermally warmed habitats should impose strong selection on ectotherms compared to ambient habitats as they increase metabolic demands, alter developmental processes, and offer novel ecological conditions. We leveraged Icelandic threespine sticklebacks (Gasterosteus aculeatus) from populations that have adaptively diverged along a geothermal/ambient habitat axis. We obtained 173,485 single nucleotide polymorphisms (SNPs) across four independent instances of population divergence using whole genome sequencing. While the majority of genomic differentiation between geothermal/ambient ecotypes was non-parallel, the MAPK signalling pathway appeared across all ecotype pairs. We also identified a putative inversion located on chromosome XXI which appears to drive parallel genomic differentiation between geothermal and ambient ecotypes. Candidate genes within the putative inversion correspond to metabolic adaptations, including regulation of appetite and fat content. Appetite level showed strong heritable divergence between ecotypes, while the rate of weight loss during starvation and fat levels differed between ecotypes. Overall, both polygenic adaptation and parallel structural variation appeared to be key genomic mechanisms for adaptation to geothermally warmed environments. While allelic divergence was largely unique across populations, it resulted in similar functional phenotypic outcomes. Thus, structural and allelic variation both operate to facilitate adaptation to warming environments. Therefore, while management from a genomic perspective will play a role in mitigating the effects of climate change, this study suggests that consideration of functional molecular pathways will be key to conservation but with precise changes being difficult to predict due to the highly polygenic nature of thermal adaptation.

The formation of ecotypes is shaped by mechanisms that reduce gene flow through complex interactions between ecological, historical, and genomic factors. In the European anchovy (Engraulis encrasicolus), marine and coastal ecotypes have been identified in the North-East Atlantic and Mediterranean Sea, yet the genomic basis of their divergence remains unclear. Here, we present the first genome-scale analysis of this species complex, integrating whole-genome sequencing (WGS) and RAD-seq data from populations across its distribution range. In addition to the marine and coastal ecotypes, we identify a previously undetected lineage which is present in southern Morocco, the Canary Islands and even in South Africa. This southern Atlantic lineage exhibits a gradient of admixture with northern populations near the Atlantic-Mediterranean transition zone. Genomic differentiation landscapes reveal large regions of high linkage disequilibrium, likely corresponding to thirteen structural variants (SVs) segregating within or between the lineages. Notably, three of the six SVs contributing to the gene flow barrier between northern ecotypes originated in the southern lineage, supporting a partially shared evolutionary history between the coastal ecotype and the southern lineage. Our findings suggest that anchovy ecotype divergence has been shaped by a combination of ancient structural variation, admixture, and local adaptation. This study highlights how SVs that arose between geographically isolated lineages can act as key genetic elements in ecotype formation, reinforcing reproductive isolation through distinct evolutionary pathways.

O_LIDisentangling the relative influence of abiotic and biotic factors on plant population differentiation is a major challenge. Orchids often occur in patchily distributed populations, and all orchids depend on orchid mycorrhizal fungi (OrM) for seed germination. Hence, local abiotic conditions together with OrM may influence population differentiation and adaptation. C_LIO_LIBased on 316,952 polymorphic SNPs sampled from 21 populations throughout the range of the Mediterranean orchid, Orchis italica (Poir.), we performed a suite of analyses to test how population differentiation and potential adaptation is influenced by the interplay between abiotic factors and OrM. C_LIO_LIWe found strong differentiation at the regional level, while loci under selection were associated with temperature, precipitation regime, soil texture, and overall OrM abundance. Outlier SNP functions were associated with stress responses and metabolic processes in the presence of OrM. C_LIO_LIAbiotic conditions and OrM combined determines differentiation in O. italica. Identifying selective pressures underlying differentiation and adaptive variation is critical for understanding plant responses to ongoing environmental change. C_LI

Species vary in their ability to adapt to rapid changes, with the presence of genetic variation often facilitating long-term evolutionary responses. Given the impending threat of climate change, it is critical to investigate how genetic variation facilitates persistence and possible range expansion in animals. Here, we combine genomic and climatic data to characterize the drivers of local adaptation in the widely distributed, social wasp Mischocyttarus mexicanus cubicola. Using whole genome sequence data, we show that adaptive genomic variation is linked to a climatic gradient across the broad distribution of this species. We found strong population structure, dividing the species into two genetic clusters that follow subtropical and temperate regions. Patterns of gene flow across the range deviate from those expected by isolation by distance alone with climatic differences resulting in reduced gene flow even between adjacent populations. Importantly, genotype-environment analyses reveal candidate single nucleotide polymorphism (SNPs) associated with temperature and rainfall, suggesting adaptation for thermal and desiccation tolerance. In particular, candidate SNPs in or near mitochondrial genes ND5, CO1, and COIII are linked to cold tolerance and metabolism. Similarly, the Gld nuclear gene shown to mediate cold hardiness and cuticle formation, shows two candidate SNPs with non-synonymous mutations unique to temperate populations. Together, our results reveal candidate SNPs consistent with local adaptation to distinct climatic conditions. Thus, the integration of genomic and climatic data can be a powerful approach to predict vulnerability and persistence of species under rapid climate change.

As heatwaves increase in intensity and frequency, more birds are exposed to sublethal heat, which can affect many elements of the phenotype, from growth to cognition to reproduction. These widespread performance-related effects of heat, coupled with the rapid declines seen in many bird populations in recent decades, underscore the urgency of detecting recent heat exposure and its downstream physiological effects in the wild. To develop minimally invasive biomarkers of past heat, we experimentally elevated nest temperatures for free-living nestling Tree Swallows (Tachycineta bicolor) for four hours on their twelfth day of life. Twenty-four hours later, we returned to collect a small blood sample and quantify carryover effects of prior sublethal heat on the blood transcriptome. By comparing these carryover effects to those that occur in the immediate aftermath of heat, we identify biomarkers of heat that reflect distinct and time-dependent processes. Candidate biomarkers include four upregulated genes with connections to stress and disease (LAMA3, ATP1B1, RASGEF1A, TMEM181) and two additional down-regulated genes. By incorporating the sex of each nestling into our analyses, we also unveiled marked sexual dimorphism in the blood transcriptome, even among autosomal genes and including pathways that imply inherent sex differences in heat tolerance. When these sex differences are controlled, we see that the sexes respond to heat with overwhelming similarity, further grounding the utility of our suggested transcriptomic biomarkers. Though these biomarkers will require further validation to be used across bird species, our collective results uncover temporally calibrated targets can be measured with just one drop of blood, improving our understanding of climate impacts on wild birds. Lay summaryO_LIAs global temperatures rise, many birds experience bouts of heat stress, but we do not have simple biomarkers that reliably reflect this past exposure in the wild. C_LIO_LIWe tested whether a small blood sample could reveal recent heat stress through changes in gene activity. C_LIO_LIOur experiment exposed nestling Tree Swallows to a non-lethal heat stressor and measured how their gene activity changed during and after the heat event. C_LIO_LISome genes reacted quickly but returned to normal within a day, while others showed longer-lasting effects. C_LIO_LIMales and females responded to heat in similar ways, even though their baseline gene activity differed substantially. C_LIO_LISix genes responded consistently across the sexes and in relation to temperature, making them promising biomarkers of past heat. C_LIO_LIThese results can help scientists better track heat exposure in wild birds and improve predictions on how populations respond to continued climate change. C_LI

Free-living nematodes are among the most abundant animals on Earth and play critical ecological roles in soil ecosystems. However, the global population structure and evolutionary history of most species remain poorly understood. Here, we analyzed genome-wide variation in Oscheius tipulae using whole-genome sequence data from 31 isolates, including 28 publicly available genomes and three newly collected strains from Korea. Population structure analyses, phylogenomic inference, and ancestry estimation consistently identified three deeply divergent lineages. These analyses did not detect admixture among lineages and collectively supported a predominantly tree-like evolutionary history. Notably, the lineages were structured by latitude rather than geographic proximity. Isolates from similar latitudinal zones clustered together regardless of continental origin, forming three major groups: northern mid-latitude (NML), low-latitude (LL), and southern mid-latitude (SML). This pattern indicates that the lineages have maintained largely independent evolutionary trajectories over extended timescales despite the potential for long-distance dispersal. Furthermore, environmentally associated variants showed significant differentiation among lineages, indicating that environmental selection may contribute to the maintenance of this latitudinally structured diversity. Our results reveal unexpectedly deep global divergence within O. tipulae, and highlight the importance of ecological divergence and long-term lineage retention in shaping the global diversity of this group.

Host-parasite co-evolution drives the diversification of host immune genes involved in the recognition of pathogen antigens and molecular patterns. In contrast, the immune genes involved in self-recognition and inhibition of immune responses against self-cells (missing-self immunity) are expected to be evolutionarily constrained. However, many pathogens, such as the Lyme disease agent Borrelia, hijack these genes to evade the immune system and may therefore select for their diversification. How these contrasting but concurrent selective forces shape the evolution of missing-self regulators is not clearly understood. To fill this gap, we investigated polymorphism and molecular signatures of selection acting on a missing-self regulator, the Complement Factor H (CFH), in bank vole populations, which are an important wild reservoir for Borrelia. We then compared the geographic structuring in the CFH domain interacting with Borrelia (CCP 20) against a genomic background represented by RAD-seq markers. We found signals of positive and diversifying selection at CCP 20, suggesting that CFH evolved in response to pressures from pathogens. Additionally, we found other innate immunity genes within the alternative complement pathway, which is regulated by CFH, under diversifying selection, highlighting its involvement in host-parasite coevolution. This study demonstrates that an innate missing-self sensor in a wild vertebrate is under diversifying selection, likely driven by pathogens.

Post-glacial recolonization of Fennoscandia created secondary contact zones in many species, offering opportunities to study how gene flow and selection contribute to their establishment and maintenance. Here, we analyse genomic data from three boreal tree species--Picea abies, Betula pendula, and Pinus sylvestris--sampled along a latitudinal gradient in Sweden. Despite differences in colonization timing and dispersal ecology, all three species exhibit north-south genetic structuring aligned with environmental gradients. Most notably, the two main genetic clusters within each species overlap in a shared contact zone, corresponding to the climatic transition between Swedens two major environmental zones. The extent and structure of the contact zone differ among species: P. abies shows stronger genetic structure and moderate gene flow, B. pendula exhibits intermediate differentiation and gene flow, and P. sylvestris displays the weakest structure with stronger gene flow. All three species also show genomic signatures of local adaptation, with distinct underlying architectures. In P. abies, adaptive loci are broadly distributed across the genome, while, strikingly, they are mostly found within an inversion on chromosome 1 in B. pendula. In P. sylvestris, local adaptation likely relies on subtle allele frequency shifts across many loci with weak signals. These patterns align with theoretical expectations for polygenic local adaptation under varying migration regimes. Our comparative approach demonstrates how gene flow and selection jointly shape genomic landscapes in shared environments and contributes to understanding local adaptation in forest trees, with implications for predicting species responses to climate change.

Hybridisation is a major source of evolutionary innovation. However, several prezygotic and postzygotic factors influence its likelihood and evolutionary outcomes. Differences in mating systems can have a major effect on the extent and direction of hybridisation and introgression. In plants, epigenetic mechanisms help to stabilize hybrid genomes and contribute to reproductive isolation, but the relationship between genetic and epigenetic changes in animal hybrids is unclear. We analysed the extent of a unique case of natural hybridisation between two genetically distant mangrove killifish species with different mating systems, Kryptolebias hermaphroditus (self-fertilising) and K. ocellatus (outcrossing), and the methylation patterns of their hybrids. Hybridisation rate between the species ranged between 14% and 26%. Although co-existing parental species displayed highly distinct genetic (microsatellites and SNPs) and methylation patterns (37,000 differentially methylated cytosines), our results indicate that F1 hybrids are viable and able to backcross with parental species. Hybrids had predominantly intermediate methylation patterns (88.5% of the sites) suggesting additive effects, as expected from hybridisation between genetically distant species. Differentially methylated cytosines between hybrids and both parental species (5,800) suggest that introgressive hybridisation may play a role in generating novel genetic and epigenetic variation which could lead to species diversification. We also found a small percentage of non-additive epigenetic effects which might act as an evolutionary bet-hedging strategy and increase fitness under environmental change.

Environmental change, such as increased rates of urbanization, can induce shifts in phenotypic plasticity with some individuals adapting to city life while others are displaced. A key trait that can facilitate adaptation is the degree at which animals respond to stress. This stress response has a heritable component and exhibits intra- and inter-individual variation. However, the mechanisms behind this variability and whether they might be responsible for adaptation to different environments are not known. Variation in DNA methylation can be a potential mechanism that mediates environmental effects on the stress response. We used an inter- and intra-environmental cross-foster experiment to analyze the contribution of DNA methylation to early-life phenotypic variation. We found that at hatching, urban house wren (Troglodytes aedon) offspring had increased methylation as compared to their rural counterparts, and observed plasticity in methylation as offspring aged, indicating developmental effects of the rearing environment on methylation. Differential methylation analyses showed that cellular respiration genes were differentially expressed at hatching and behavioral and metabolism genes were differentially expressed at fledgling. Lastly, hyper-methylation of a single gene (CNTNAP2) is associated with increased glucocorticoid levels. These differential methylation patterns linked to a specific physiological phenotype suggest that DNA methylation may be a mechanism by which individuals adapt to novel environments. Characterizing genetic and environmental influences on methylation is critical for understanding the role of epigenetic mechanisms in evolutionary adaptation.

Pleistocene climatic fluctuations have often driven range shifts and hybridization among related species, leaving present-day genomic footprints. In the Iberian Peninsula, Lepus timidus, after its post-deglaciation retreat, has left extensive mitochondrial DNA traces in three other hare species, but the genomic correlates and underlying biogeographic scenarios are still incompletely understood. This study focuses on Lepus castroviejoi, endemic to the Cantabrian region, using its non-Iberian sister species, L. corsicanus, for comparison. By analyzing coalescent patterns from 10 genomes, we estimate that these species remained isolated since their divergence, around 50,000 years ago, consistent with their current allopatry. Further analyses with 25 additional genomes indicate that small fractions of the L. castroviejoi genome originate from L. granatensis, L. timidus, and L. europaeus (0.72%, 0.08%, and 0.04%, respectively). Introgression dating based on tract lengths suggests L. granatensis was already admixed with L. timidus when it hybridized with L. castroviejoi, which could explain the granatensis-timidus ancestry tract junctions detected in L. castroviejoi. Genomic segments with such junctions contain genes enriched for cell signaling and olfactory receptor activity, possibly facilitating genetic exchange. This research demonstrates how genomic ancestry inferences can reveal complex multiway admixture histories and illuminate past biogeographic events.

1.Freshwater sponges fulfill critical ecological functions, including maintaining water quality, regulating nutrient dynamics, offering habitats for diverse taxa, and serving as a vital food source for various species. However, their patterns of dispersal and genetic connectivity remain inadequately understood, posing significant challenges to effective conservation assessments. We examined genetic connectivity and genetic adaptation to local environmental conditions in Ephydatia muelleri across its geographic range using ddRADseq-derived SNPs from 106 individuals collected from 11 localities spanning North America, Europe, and Asia. Analysis of 3,182 neutral SNPs revealed low connectivity and strong genetic structure among regions within two main genetic clusters of North America and Eurasia, while 115 SNPs identified to be under selection showed considerable evidence for differentiated, polygenic adaptation to light and temperature conditions across sampled locations, as well as selection on gene regulatory processes. These findings align with the "monopolization hypothesis", suggesting that historical climatic and geological conditions of the Last Glacial Maximum, including habitat expansion, contraction, and natural barriers, have contributed more to the current genetic structure of E. muelleri populations than contemporary gene flow, which is restricted by monopolistic habitat colonization by this species. Our results provide novel support for ecological theory on dispersal in aquatic invertebrates, as well as insights into the plasticity of E. muelleri in the face of varying environmental conditions that are fundamentally important for freshwater ecosystem conservation.

Early-life experiences can have profound and long-lasting effects on adult phenotype and thereby Darwinian fitness, though the mechanisms driving these effects remain poorly understood. Epigenetic alterations, especially DNA methylation which affects gene expression, potentially mediate developmental condition effects on adult phenotype. We tested for such effects using captive zebra finches that were reared in either small or large broods, a manipulation that is known to have pleiotropic phenotypic effects. Analyzing whole genome DNA methylation patterns in erythrocytes from 50 individuals sampled in adulthood, we found 0.8% of all CpG sites after filtering to be differentially methylated after correction for multiple testing. We identified 149 non-transiently differentially methylated sites (DMSs) where the DNA methylation difference between treatments was larger than 25%. These DMSs were located in 19 autosomal chromosomes, in or near genes involved in critical biological processes such as cell growth, division, and differentiation, regulation of immune response, muscle contraction, and neuronal signaling. These findings suggest that epigenetic modifications such as DNA methylation potentially mediate long-term effects of early-life adversity via differential gene expression, but follow-up studies are needed to identify the extent to which the observed DMSs are functionally related to the previously observed phenotypic effects.

Gene expression, resulting from complex regulatory interactions, plays an important role in adaptation and speciation. While gene expression historically has been studied in the context of reproductive isolation in speciation research, a role for evolved differences in gene expression in adaptation to novel niches is increasingly appreciated. How gene expression evolves and enables divergent ecological adaptation, and how changes in gene expression relate to genomic architecture and genetic divergence are pressing questions in understanding the processes of adaptation and ecological speciation. Further, how plasticity in gene expression can both contribute to and be affected by the process of ecological adaptation is a crucial component in understanding gene expression evolution. To address these questions, we investigate the role of evolved and plastic gene expression differences in adaptation leveraging an established host plant shift in the peacock fly Tephritis conura. Using a cross-fostering design where larvae feed on either natal or alternate host plants, we uncover extensive evolved differences in gene expression between the ecotypes, strikingly in genes associated with processing of host plant chemicals. We find limited evidence for plasticity, with some indications of higher plasticity in the ancestral ecotype where the expression of three gene coexpression modules is altered when larvae are cross-fostered to the derived host plant. Interestingly, we find an enrichment of differentially expressed genes within a large, ecotype-specific inversion in the T. conura genome. This finding adds to evidence that inversions are important for enabling diversification in the face of gene flow and underscores that effects on gene expression may be key to understanding the role of inversions.

Even seemingly homogeneous on the surface, the oceans display high environmental heterogeneity across space and time. Indeed, different soft barriers structure the marine environment, which offers an appealing opportunity to study various evolutionary processes such as population differentiation and speciation. Here, we focus on Amphiprion clarkii (Actinopterygii; Perciformes), the most widespread of clownfishes that exhibits the highest colour polymorphism. Clownfishes can only disperse during a short pelagic larval phase before their sessile adult lifestyle, which might limit connectivity among populations, thus facilitating speciation events. Consequently, the taxonomic status of A. clarkii has been under debate. We used whole-genome resequencing data of 67 A. clarkii specimens spread across the Indian and Pacific Oceans to characterise the species population structure, demographic history, and colour polymorphism. We found that A. clarkii spread from the Indo-Pacific Ocean to the Pacific and Indian Oceans following a stepping-stone dispersal and that gene flow was pervasive throughout its demographic history. Moreover, edge populations exhibited more similar colouration patterns compared to central populations. However, we demonstrate that colour polymorphism is not associated with population structure, thus, colour phenotype is unreliable in assessing the taxonomic status of A. clarkii. Our study further highlights the power of whole-genome comparative studies to determine the taxonomy of geographically wide-ranging and phenotypically diverse species, supporting the status of A. clarkii as a single species.

Understanding how environmental variation interacts with gene flow to shape population genomic patterns is a central goal in evolutionary biology. We investigated how geographic and environmental differences impact genomic variation in the clouded sulfur butterfly (Colias philodice eriphyle) by conducting whole-genome resequencing across replicated transects consisting of paired high- and low-elevation sites on both sides of a major mountain range. Despite sampling across steep environmental gradients, we found no evidence of discrete population structure, indicating high connectivity across the region. Nonetheless, significant isolation by distance - strongest in eastern populations - revealed that geographic distance still imposes limits on gene flow, and genetic diversity was also elevated in the east. Genotype-environment association analyses identified more than 16,000 loci associated with elevation, precipitation, and solar radiation. Our redundancy analysis identified precipitation as the strongest predictor of adaptive genomic differentiation, and candidate genes included those linked to melanization and thermoregulation (e.g., TH and yellow). These results demonstrate that even in a largely panmictic population, environmental variation can maintain regional-scale signals of local adaptation. Because insects are declining globally and remain underrepresented in genomic monitoring, conducting whole-genome analyses in a widespread species provides valuable context for assessing how insects today persist across such diverse landscapes and their potential for withstanding future environmental change.

Whole-genome sequence data is proving to be highly informative about the past demography of free-living populations, and in the context of endangered species, it can provide a quantification of the genetic risk posed by reduced genetic diversity and inbreeding. Prior to 1920, the Florida scrub-jay (Aphelocoma coerulescens) had been widespread across Florida, but with the expansion of agriculture and human habitation, its population has declined by 95%, resulting in fragmentation into semi-isolated subpopulations. By sequencing 241 individuals sampled from five different regions and across two time points, this study quantifies a greater magnitude of loss of genetic diversity and greater levels of inbreeding in smaller and more isolated subpopulations. Consistent with population genetics theory, reduction in population size results in a dramatic loss of rare alleles, skewing the site frequency spectrum far from the expected equilibrium. Increased inbreeding in the smaller, more remote subpopulations is especially evident in the increased size and number of runs of homozygosity. The Florida scrub-jay displays limited dispersal, and habitat fragmentation has greatly reduced the magnitude of gene flow in the past 30 years, resulting in further decline of genetic diversity, especially in the peripheral populations. Analysis of these data is informative in guiding conservation efforts to retain genetic diversity and minimize the consequences of inbreeding in the Florida scrub-jay. HighlightsO_LIFive regional populations show distinct degrees of population isolation and decline. C_LIO_LIThere has been commensurate loss of genetic diversity, skewed site frequency spectra, reduced migration, and increased inbreeding (FROH). C_LIO_LIAs many state-wide populations decline, the smaller, more remote populations provide a glimpse into the future and a testbed for remediation approaches. C_LI

Anthropogenic climate change has caused unprecedented declines across a number of marine taxa. Coral reef ecosystems, which are formed by scleractinian corals, face widespread declines in ecosystem health and function due to co-occurring environmental stressors. Frequent exposure of corals to a variety of biotic and abiotic stressors makes these cnidarians a prime candidate for investigating the effects of multiple stressors on marine ecosystems. In recent decades, hyperthermic bleaching events and disease outbreaks have been prominent stressors on reefs. Disease outbreaks often follow hyperthermic bleaching events, yet the mechanisms driving the diffuse associations between bleaching and disease are poorly understood. Here we investigated the mechanisms linking sequential bleaching and disease using the model cnidarian Exaiptasia diaphana. We examined the transcriptomic responses of anemones to immune challenge during acute recovery from prior heat stress. We observed notable upregulation of apoptotic pathways and downregulation of autophagic pathways in previously heat-stressed anemones, while anemones maintained at ambient temperatures displayed an inverse pattern characterized by downregulation of apoptosis. Furthermore, network analyses suggest that disruption of host-Symbiodiniaceae nutrient exchange during bleaching recovery of previously heat-stressed anemones may contribute to observed immune suppression following heat stress. These results provide insight regarding the cellular mechanisms facilitating increased disease susceptibility during recovery from heat stress, highlighting the roles of immunological regulation and nutrient availability in these processes.

Living at high density and with low genetic diversity are factors that should both increase the susceptibility of organisms to disease. Therefore, group living organisms, especially those that are inbred, should be especially vulnerable to infection and therefore have particular strategies to cope with infection. Phenotypic plasticity, underpinned by epigenetic changes, could allow group living organisms to rapidly respond to infection challenges. To explore the potential role of epigenetic modifications in the immune response to a group-living species with low genetic diversity, we compared the genome-wide DNA methylation profiles of five colonies of social spiders (Stegodyphus dumicola) in their natural habitat in Namibia at the point just before they succumbed to infection to a point at least six months previously where they were presumably healthier. We found increases in genome- and chromosome-wide methylation levels in the CpG, CHG, and CHH contexts, although the genome-wide changes were not clearly different from zero. These changes were most prominent in the CHG context, especially at a narrow region of chromosome 13, hinting at an as-of-yet unsuspected role of this DNA methylation context in phenotypic plasticity. However, there were few clear patterns of differential methylation at the base level, and genes with a known immune function in spiders had mean methylation changes close to zero. Our results suggest that DNA methylation may change with infection at large genomic scales, but that this type of epigenetic change is not necessarily integral to the immune response of social spiders.