Prepping for climate change by introgressive hybridization.

Species and populations may adapt to climate change by microevolutionary processes. However, standing genetic variation can be insufficient for this to occur. An interesting new study of a system of rainbowfish species shows that intraspecific hybridization enriches gene pools with adaptive variation that may allow persistence in a changing climate.

Genetic responses in sexual diploid and unisexual triploid goldfish (Carassius auratus) introduced into a high-altitude environment.

Anthropogenic biological invasions represent major concerns but enable us to investigate rapid evolutionary changes and adaptation to novel environments. The goldfish Carassius auratus with sexual diploids and unisexual triploids coexisting in natural waters is one of the most widespread invasive fishes in Tibet, providing an ideal model to study evolutionary processes during invasion in different reproductive forms from the same vertebrate. Here, using whole-genome resequencing data of 151 C. auratus individuals from invasive and native ranges, we found different patterns of genomic responses between diploid and triploid populations during their invasion of Tibet. For diploids, although invasive individuals derived from two different genetically distinct sources had a relative higher diversity (π) at the population level, their individual genetic diversity (genome-wide observed heterozygosity) was significantly lower (21.4%) than that of source individuals. Population structure analysis revealed that the invasive individuals formed a specific genetic cluster distinct from the source populations. Runs of homozygosity analysis showed low inbreeding only in invasive individuals, and only the invasive population experienced a recent decline in effective population size reflecting founder events. For triploids, however, invasive populations showed no loss of individual genetic diversity and no genetic differentiation relative to source populations. Regions of putative selective sweeps between invasive and source populations of diploids mainly involved genes associated with mannosidase activity and embryo development. Our results suggest that invasive diploids deriving from distinct sources still lost individual genetic diversity resulting from recent inbreeding and founder events and selective sweeps, and invasive triploids experienced no change in genetic diversity owing to their reproduction mode of gynogenesis that precludes inbreeding and founder effects and may make them more powerful invaders.

Genome-wide methylation in the panmictic European eel (Anguilla anguilla).

The role of methylation in adaptive, developmental and speciation processes has attracted considerable interest, but interpretation of results is complicated by diffuse boundaries between genetic and non-genetic variation. We studied whole genome genetic and methylation variation in the European eel, distributed from subarctic to subtropical environments, but with panmixia precluding genetically based local adaptation beyond single-generation responses. Overall methylation was 70.9%, with hypomethylation predominantly found in promoters and first exons. Redundancy analyses involving juvenile glass eels showed 0.06% and 0.03% of the variance at SNPs to be explained by localities and environmental variables, respectively, with GO terms of genes associated with outliers primarily involving neural system functioning. For CpGs 2.98% and 1.36% of variance was explained by localities and environmental variables. Differentially methylated regions particularly included genes involved in developmental processes, with Hox clusters featuring prominently. Life stage (adult versus glass eels) was the most important source of inter-individual variation in methylation, probably reflecting both ageing and developmental processes. Demethylation of transposable elements relative to pure European eel was observed in European X American eel hybrids, possibly representing postzygotic barriers in this system characterized by prolonged speciation and ongoing gene flow. Whereas the genetic data are consistent with a role of single-generation selective responses, the methylation results underpin the importance of epigenetics in the life cycle of eels and suggest interactions between local environments, development and phenotypic variation mediated by methylation variation. Eels are remarkable by having retained eight Hox clusters, and the results suggest important roles of methylation at Hox genes for adaptive processes.

Combining population genomics with demographic analyses highlights habitat patchiness and larval dispersal as determinants of connectivity in coastal fish species.

Gene flow shapes spatial genetic structure and the potential for local adaptation. Among marine animals with nonmigratory adults, the presence or absence of a pelagic larval stage is thought to be a key determinant in shaping gene flow and the genetic structure of populations. In addition, the spatial distribution of suitable habitats is expected to influence the distribution of biological populations and their connectivity patterns. We used whole genome sequencing to study demographic history and reduced representation (double-digest restriction associated DNA) sequencing data to analyse spatial genetic structure in broadnosed pipefish (Syngnathus typhle). Its main habitat is eelgrass beds, which are patchily distributed along the study area in southern Norway. Demographic connectivity among populations was inferred from long-term (~30-year) population counts that uncovered a rapid decline in spatial correlations in abundance with distance as short as ~2 km. These findings were contrasted with data for two other fish species that have a pelagic larval stage (corkwing wrasse, Symphodus melops; black goby, Gobius niger). For these latter species, we found wider spatial scales of connectivity and weaker genetic isolation-by-distance patterns, except where both species experienced a strong barrier to gene flow, seemingly due to lack of suitable habitat. Our findings verify expectations that a fragmented habitat and absence of a pelagic larval stage promote genetic structure, while presence of a pelagic larvae stage increases demographic connectivity and gene flow, except perhaps over extensive habitat gaps.

Long-term monitoring of a brown trout (Salmo trutta) population reveals kin-associated migration patterns and contributions by resident trout to the anadromous run.

BACKGROUND: In species showing partial migration, as is the case for many salmonid fishes, it is important to assess how anthropogenic pressure experienced by migrating individuals affects the total population. We focused on brown trout (Salmo trutta) from the Guddal River in the Norwegian Hardanger Fjord system, which encompasses both resident and anadromous individuals. Aquaculture has led to increased anthropogenic pressure on brown trout during the marine phase in this region. Fish traps in the Guddal River allow for sampling all ascending anadromous spawners and descending smolts. We analyzed microsatellite DNA markers from all individuals ascending in 2006-2016, along with all emigrating smolts in 2017. We investigated (1) if there was evidence for declines in census numbers and effective population size during that period, (2) if there was association between kinship and migration timing in smolts and anadromous adults, and (3) to what extent resident trout were parents of outmigrating smolts. RESULTS: Census counts of anadromous spawners showed no evidence for a decline from 2006 to 2016, but were lower than in 2000-2005. Estimates of effective population size also showed no trends of declines during the study period. Sibship reconstruction of the 2017 smolt run showed significant association between kinship and migration timing, and a similar association was indicated in anadromous spawners. Parentage assignment of 2017 smolts with ascending anadromous trout as candidate parents, and assuming that unknown parents represented resident trout, showed that 70% of smolts had at least one resident parent and 24% had two resident parents. CONCLUSIONS: The results bear evidence of a population that after an initial decline has stabilized at a lower number of anadromous spawners. The significant association between kinship and migration timing in smolts suggests that specific episodes of elevated mortality in the sea could disproportionally affect some families and reduce overall effective population size. Finally, the results based on parentage assignment demonstrate a strong buffering effect of resident trout in case of elevated marine mortality affecting anadromous trout, but also highlight that increased mortality of anadromous trout, most of which are females, may lower overall production in the system.

Speciation history of European (Anguilla anguilla) and American eel (A. rostrata), analysed using genomic data.

Speciation in the ocean could differ from terrestrial environments due to fewer barriers to gene flow. Hence, sympatric speciation might be common, with American and European eel being candidates for exemplifying this. They show disjunct continental distributions on both sides of the Atlantic, but spawn in overlapping regions of the Sargasso Sea from where juveniles are advected to North American, European and North African coasts. Hybridization and introgression are known to occur, with hybrids almost exclusively observed in Iceland. Different speciation scenarios have been suggested, involving either vicariance or sympatric ecological speciation. Using RAD sequencing and whole-genome sequencing data from parental species and F1 hybrids, we analysed speciation history based on the joint allele frequency spectrum (JAFS) and pairwise sequentially Markovian coalescent (PSMC) plots. JAFS supported a model involving a split without gene flow 150,000-160,000 generations ago, followed by secondary contact 87,000-92,000 generations ago, with 64% of the genome experiencing restricted gene flow. This supports vicariance rather than sympatric speciation, likely associated with Pleistocene glaciation cycles and ocean current changes. Whole-genome PSMC analysis of F1 hybrids from Iceland suggested divergence 200,000 generations ago and indicated subsequent gene flow rather than strict isolation. Finally, simulations showed that results from both approaches (JAFS and PSMC) were congruent. Hence, there is strong evidence against sympatric speciation in North Atlantic eels. These results reiterate the need for careful consideration of cases of possible sympatric speciation, as even in seemingly barrier-free oceanic environments palaeoceanographic factors may have promoted vicariance and allopatric speciation.

Demographic history has shaped the strongly differentiated corkwing wrasse populations in Northern Europe.

Understanding the biological processes involved in genetic differentiation and divergence between populations within species is a pivotal aim in evolutionary biology. One particular phenomenon that requires clarification is the maintenance of genetic barriers despite the high potential for gene flow in the marine environment. Such patterns have been attributed to limited dispersal or local adaptation, and to a lesser extent to the demographic history of the species. The corkwing wrasse (Symphodus melops) is an example of a marine fish species where regions of particular strong divergence are observed. One such genetic break occurred at a surprisingly small spatial scale (FST ~0.1), over a short coastline (<60 km) in the North Sea-Skagerrak transition area in southwestern Norway. Here, we investigate the observed divergence and purported reproductive isolation using genome resequencing. Our results suggest that historical events during the post-glacial recolonization route can explain the present population structure of the corkwing wrasse in the northeast Atlantic. While the divergence across the break is strong, we detected ongoing gene flow between populations over the break suggesting recent contact or negative selection against hybrids. Moreover, we found few outlier loci and no clear genomic regions potentially being under selection. We concluded that neutral processes and random genetic drift e.g., due to founder events during colonization have shaped the population structure in this species in Northern Europe. Our findings underline the need to take into account the demographic process in studies of divergence processes.

Interbreeding between local and translocated populations of a cleaner fish in an experimental mesocosm predicts risk of disrupted local adaptation.

Translocation of organisms within or outside its native range carries the risk of modifying the community of the recipient ecosystems and induces gene flow between locally adapted populations or closely related species. In this study, we evaluated the genetic consequences of large-scale translocation of cleaner wrasses that has become a common practice within the salmon aquaculture industry in northern Europe to combat sea lice infestation. A major concern with this practice is the potential for hybridization of escaped organisms with the local, recipient wrasse population, and thus potentially introduce exogenous alleles and breaking down coadapted gene complexes in local populations. We investigated the potential threat for such genetic introgressions in a large seminatural mesocosm basin. The experimental setting represented a simulated translocation of corkwing wrasse (Symphodus melops) that occurs on a large scale in the Norwegian salmon industry. Parentage assignment analysis of mesocosm's offspring revealed 30% (195 out of 651 offspring) interbreeding between the two populations, despite their being genetically (F ST = 0.094, p < 0.05) and phenotypically differentiated. Moreover, our results suggest that reproductive fitness of the translocated western population doubled that of the local southern population. Our results confirm that human translocations may overcome the impediments imposed by natural habitat discontinuities and urge for immediate action to manage the genetic resources of these small benthic wrasses.

Heat tolerance and gene expression responses to heat stress in threespine sticklebacks from ecologically divergent environments.

Ectothermic animals like fishes are extremely dependent on temperature, as they are not able to change body temperature physiologically. When populations are found in isolated water bodies such as small lakes they will have to respond to stressful high temperatures by behavioral avoidance, phenotypic plasticity or microevolutionary change. We analyzed threespine sticklebacks from two large and two small lakes, representing different isolated populations. We determined maximum critical thermal limits (CTmax) and the associated gene expression responses in three heat shock (hsp60, hsp70, hsp90) and two key metabolic (idh2, fbp2) genes at ecologically relevant moderate heat stress (26 °C) as well as at the critical thermal limit (CTmax). CTmax showed slight variation across populations with no strong indication of local adaptation. Likewise, there was no strong evidence for local adaptation at the level of gene expression. The expression of the metabolic genes indicated a shift from aerobic towards anaerobic energy production with extreme heat stress. We conclude that threespine sticklebacks do not show severe stress during the warmest temperatures they are likely to encounter during current temperature regimes in Denmark, and following this show no sign of local adaptation even in small, isolated water bodies.

Genomic parallelism and lack thereof in contrasting systems of three-spined sticklebacks.

Parallel evolution and the extent to which it involves gene reuse have attracted much interest. Whereas it has theoretically been predicted under which circumstances gene reuse is expected, empirical studies that directly compare systems showing high and low parallelism are rare. Three-spined stickleback (Gasterosteus aculeatus), where freshwater populations have been independently founded by ancestral marine populations, represent prime examples of phenotypic and genomic parallelism, but cases exist where parallelism is low. Based on RAD (restriction site associated DNA) sequencing, we analysed SNPs and chromosome inversions in populations in Denmark and Greenland showing low and high parallelism, respectively. We identified parallelism across freshwater populations in Greenland at genomic regions previously identified to be associated with marine-freshwater divergence. These same markers also separated Danish marine and freshwater sticklebacks, albeit to a weaker extent. Hence, parallelism was not absent in Denmark but possibly constrained by spatially and temporally varying selection. Divergence time estimates found one Danish freshwater population to be much older than the others. It also deviated strongly with respect to parallelism and may represent earlier postglacial colonization based on a different pool of standing variation and eliciting different adaptive responses to freshwater conditions. These findings provide empirical support to previous suggestions that the time since replicate populations had access to a common pool of standing variation is a major factor determining gene reuse. At last, based on the observed parallelism in the Greenlandic system we discuss the predictability of adaptive responses in newly established populations.

A continuous genome assembly of the corkwing wrasse (Symphodus melops).

The wrasses (Labridae) are one of the most successful and species-rich families of the Perciformes order of teleost fish. Its members display great morphological diversity, and occupy distinct trophic levels in coastal waters and coral reefs. The cleaning behaviour displayed by some wrasses, such as corkwing wrasse (Symphodus melops), is of particular interest for the salmon aquaculture industry to combat and control sea lice infestation as an alternative to chemicals and pharmaceuticals. There are still few genome assemblies available within this fish family for comparative and functional studies, despite the rapid increase in genome resources generated during the past years. Here, we present a highly continuous genome assembly of the corkwing wrasse using PacBio SMRT sequencing (x28.8) followed by error correction with paired-end Illumina data (x132.9). The present genome assembly consists of 5040 contigs (N50 = 461,652 bp) and a total size of 614 Mbp, of which 8.5% of the genome sequence encode known repeated elements. The genome assembly covers 94.21% of highly conserved genes across ray-finned fish species. We find evidence for increased copy numbers specific for corkwing wrasse possibly highlighting diversification and adaptive processes in gene families including N-linked glycosylation (ST8SIA6) and stress response kinases (HIPK1). By comparative analyses, we discover that de novo repeats, often not properly investigated during genome annotation, encode hundreds of immune-related genes. This new genomic resource, together with the ballan wrasse (Labrus bergylta), will allow for in-depth comparative genomics as well as population genetic analyses for the understudied wrasses.

Low genetic and phenotypic divergence in a contact zone between freshwater and marine sticklebacks: gene flow constrains adaptation.

BACKGROUND: Distinct hybrid zones and phenotypic and genomic divergence is often observed between marine and freshwater threespine sticklebacks (Gasterosteus aculeatus). Nevertheless, cases also exist where marine-freshwater divergence is diffuse despite seemingly similar environmental settings. In order to assess what characterizes these highly different outcomes, we focused on the latter kind of system in the Odder River, Denmark. Here, a previous study based on RAD (Restriction site Associated DNA) sequencing found non-significant genome-wide differentiation between marine and freshwater sticklebacks. In the present study, we analyzed samples on a finer geographical scale. We assessed if the system should be regarded as panmictic, or if fine-scale genetic structure and local selection was present but dominated by strong migration. We also asked if specific population components, that is the two sexes and different lateral plate morphs, contributed disproportionally more to dispersal. RESULTS: We assessed variation at 96 SNPs and the Eda gene that affects lateral plate number, conducted molecular sex identification, and analyzed morphological traits. Genetic differentiation estimated by FST was non-significant throughout the system. Nevertheless, spatial autocorrelation analysis suggested fine scale genetic structure with a genetic patch size of 770 m. There was no evidence for sex-biased dispersal, but full-plated individuals showed higher dispersal than low- and partial-plated individuals. The system was dominated by full-plated morphs characteristic of marine sticklebacks, but in the upstream part of the river body shape and frequency of low-plated morphs changed in the direction expected for freshwater sticklebacks. Five markers including Eda were under possible diversifying selection. However, only subtle clinal patterns were observed for traits and markers. CONCLUSIONS: We suggest that gene flow from marine sticklebacks overwhelms adaptation to freshwater conditions, but the short genetic patch size means that the effect of gene flow on the most upstream region must be indirect and occurs over generations. The occurrence of both weak unimodal and strong bimodal hybrid zones within the same species is striking. We suggest environmental and demographic factors that could determine these outcomes, but also highlight the possibility that long-term population history and the presence or absence of genomic incompatibilities could be a contributing factor.

PSMC (pairwise sequentially Markovian coalescent) analysis of RAD (restriction site associated DNA) sequencing data.

The pairwise sequentially Markovian coalescent (PSMC) method uses the genome sequence of a single individual to estimate demographic history covering a time span of thousands of generations. Although originally designed for whole-genome data, we here use simulations to investigate its applicability to reference genome-aligned restriction site associated DNA (RAD) data. We find that RAD data can potentially be used for PSMC analysis, but at present with limitations. The key factor is the proportion (p) of the genome that the RAD data covers. In our simulations, a proportion of 10% can still retain a substantial amount of coalescent information, whereas for 1% estimation becomes unreliable. The performance depends strongly on mutation rate (µ) and recombination rate (r) and is proportional to µ*p/r. When the value of this term is low, increasing the amount of data and number of iterations helps restoring the power of the estimation. We subsequently analyse one whole-genome-sequenced and 17 RAD-sequenced three-spined sticklebacks (Gasterosteus aculeatus) from a lake in Greenland. The whole-genome sequence suggests a relatively recent expansion and decline within ca. 4000-40 000 generations ago, possibly reflecting postglacial expansion and founding of the lake population. RAD data, where chromosomes from 10 individuals are combined, identify a similar pattern. Our study provides guidance about the use of PSMC analysis and suggests measures that can improve its utility for RAD data. Finally, the study shows that RAD loci in general contain coalescent information that can be used for developing more targeted methods.

Region-wide and ecotype-specific differences in demographic histories of threespine stickleback populations, estimated from whole genome sequences.

We analysed 81 whole genome sequences of threespine sticklebacks from Pacific North America, Greenland and Northern Europe, representing 16 populations. Principal component analysis of nuclear SNPs grouped populations according to geographical location, with Pacific populations being more divergent from each other relative to European and Greenlandic populations. Analysis of mitogenome sequences showed Northern European populations to represent a single phylogeographical lineage, whereas Greenlandic and particularly Pacific populations showed admixture between lineages. We estimated demographic history using a genomewide coalescence with recombination approach. The Pacific populations showed gradual population expansion starting >100 Kya, possibly reflecting persistence in cryptic refuges near the present distributional range, although we do not rule out possible influence of ancient admixture. Sharp population declines ca. 14-15 Kya were suggested to reflect founding of freshwater populations by marine ancestors. In Greenland and Northern Europe, demographic expansion started ca. 20-25 Kya coinciding with the end of the Last Glacial Maximum. In both regions, marine and freshwater populations started to show different demographic trajectories ca. 8-9 Kya, suggesting that this was the time of recolonization. In Northern Europe, this estimate was surprisingly late, but found support in subfossil evidence for presence of several freshwater fish species but not sticklebacks 12 Kya. The results demonstrate distinctly different demographic histories across geographical regions with potential consequences for adaptive processes. They also provide empirical support for previous assumptions about freshwater populations being founded independently from large, coherent marine populations, a key element in the Transporter Hypothesis invoked to explain the widespread occurrence of parallel evolution across freshwater stickleback populations.

The impact of selection, gene flow and demographic history on heterogeneous genomic divergence: three-spine sticklebacks in divergent environments.

Heterogeneous genomic divergence between populations may reflect selection, but should also be seen in conjunction with gene flow and drift, particularly population bottlenecks. Marine and freshwater three-spine stickleback (Gasterosteus aculeatus) populations often exhibit different lateral armour plate morphs. Moreover, strikingly parallel genomic footprints across different marine-freshwater population pairs are interpreted as parallel evolution and gene reuse. Nevertheless, in some geographic regions like the North Sea and Baltic Sea, different patterns are observed. Freshwater populations in coastal regions are often dominated by marine morphs, suggesting that gene flow overwhelms selection, and genomic parallelism may also be less pronounced. We used RAD sequencing for analysing 28 888 SNPs in two marine and seven freshwater populations in Denmark, Europe. Freshwater populations represented a variety of environments: river populations accessible to gene flow from marine sticklebacks and large and small isolated lakes with and without fish predators. Sticklebacks in an accessible river environment showed minimal morphological and genomewide divergence from marine populations, supporting the hypothesis of gene flow overriding selection. Allele frequency spectra suggested bottlenecks in all freshwater populations, and particularly two small lake populations. However, genomic footprints ascribed to selection could nevertheless be identified. No genomic regions were consistent freshwater-marine outliers, and parallelism was much lower than in other comparable studies. Two genomic regions previously described to be under divergent selection in freshwater and marine populations were outliers between different freshwater populations. We ascribe these patterns to stronger environmental heterogeneity among freshwater populations in our study as compared to most other studies, although the demographic history involving bottlenecks should also be considered in the interpretation of results.

Comparative analysis of complete mitochondrial genomes suggests that relaxed purifying selection is driving high nonsynonymous evolutionary rate of the NADH2 gene in whitefish (Coregonus ssp.).

Several studies have recently reported evidence for positive selection acting on the mitochondrial genome (mitogenome), emphasizing its potential role in adaptive divergence and speciation. In this study we searched 107 full mitogenomes of recently diverged species and lineages of whitefish (Coregonus ssp.) for signals of positive selection. These salmonids show several distinct morphological and ecological differences that may be associated with energetics and therefore potentially positive selection at the mitogenome level. We found that purifying selection and genetic drift were the predominant evolutionary forces acting on the analyzed mitogenomes. However, the NADH dehydrogenase 2 gene (ND2) showed a highly elevated dN/dS ratio compared to the other mitochondrial genes, which was significantly higher in whitefish compared to other salmonids. We therefore further examined nonsynonymous evolution in ND2 by (i) mapping amino acid changes to a protein model structure which showed that they were located away from key functional residues of the protein, (ii) locating them in the sequences of other species of fish (Salmonidae, Anguillidae, Scombridae and Percidae) only to find pronounced overlap of nonsynonymous regions. We thus conclude that relaxed purifying selection is driving the evolution of ND2 by affecting mostly regions that have lower functional relevance.

Human effects on ecological connectivity in aquatic ecosystems: Integrating scientific approaches to support management and mitigation.

Understanding the drivers and implications of anthropogenic disturbance of ecological connectivity is a key concern for the conservation of biodiversity and ecosystem processes. Here, we review human activities that affect the movements and dispersal of aquatic organisms, including damming of rivers, river regulation, habitat loss and alteration, human-assisted dispersal of organisms and climate change. Using a series of case studies, we show that the insight needed to understand the nature and implications of connectivity, and to underpin conservation and management, is best achieved via data synthesis from multiple analytical approaches. We identify four key knowledge requirements for progressing our understanding of the effects of anthropogenic impacts on ecological connectivity: autecology; population structure; movement characteristics; and environmental tolerance/phenotypic plasticity. Structuring empirical research around these four broad data requirements, and using this information to parameterise appropriate models and develop management approaches, will allow for mitigation of the effects of anthropogenic disturbance on ecological connectivity in aquatic ecosystems.

Relationship between amino acid changes in mitochondrial ATP6 and life-history variation in anguillid eels.

Mitochondrial genes are part of the oxidative phosphorylation pathway and important for energy production. Although evidence for positive selection at the mitochondrial level exists, few studies have investigated the link between amino acid changes and phenotype. Here we test the hypothesis that differences in two life-history related traits, migratory distance between spawning and foraging areas and larval phase duration, are associated with divergent selection within the mitochondrial ATP6 gene in anguillid eels. We compare amino acid changes among 18 species with the sequence of the putative ancestral species, believed to have shown short migratory distance and larval phase duration. We find positive correlations between both life-history related traits and (i) the number of amino acid changes and (ii) the strength of the combined physico-chemical and structural changes at positions previously identified as candidates for positive selection. This supports a link between genotype and phenotype driven by positive selection at ATP6.