Molecular Ecology

Evolutionary and demographic processes such as selection, drift and migration shape the genetic variation of populations. Genetic diversity is often lower in populations toward higher latitudes. This decrease potentially threatens their survival as several factors are putting more pressure on the populations, including the spread of infectious diseases. In this study, we combined whole-genome re-sequencing with MHC class II genotyping and skin microbiome profiling in Bufo bufo and B. spinosus, two closely related European toad species. We investigated the underlying immunogenetic and microbial variation resulting from different demographic histories and environmental conditions to identify their potential impact on infection outcomes in these two species. We found lower immunogenetic diversity in B. bufo compared to B. spinosus, with highly significant differences in genes related to adaptive and innate immunity. We found lower overall MHC class II diversity and skin microbiome diversity at the species level in B. bufo, compared with B. spinosus. In contrast, at the individual level, B. bufo showed higher MHC allelic diversity and greater diversity in the core skin microbiota than B. spinosus. Together, our findings suggest that divergence in immunogenetic background and host-associated microbial communities may underlie differences in susceptibility to emerging infectious diseases. This integrative framework provides new insight into how host genetics and microbial communities jointly influence disease outcomes across environmental gradients.

Infection can substantially reduce host fitness and influence population dynamics, yet it is often difficult to detect and quantify in wild animal populations. Molecular tools offer a valuable means of identifying cryptic infection in natural systems. Using whole-genome bisulfite sequencing, we examined whether infection with the parasitic leech Ozobranchus margoi is associated with DNA methylation variation in loggerhead sea turtles (Caretta caretta), while also assessing the potential value of this variation as a biomarker of parasite infection. In nesting females, we identified infection-associated differentially methylated CpG sites associated with genes implicated in immune signalling and cellular regulation. Offspring of infected females also showed infection-associated methylation patterns, despite not being directly exposed to the parasite themselves. Differential methylation analyses identified genes involved in immunity, neurodevelopment and metabolic activity, with limited overlap in associated genes and no overlap in differentially methylated sites between generations. Maternal and offspring genome-wide methylation levels showed a non-linear association that differed subtly with maternal infection status, indicating that infection modifies intergenerational methylation associations. Finally, methylation profiles showed strong discriminatory power for maternal infection status in both maternal and hatchling samples using machine learning models, supporting their potential as candidate biomarkers of cryptic infection. Together, these results show that parasite infection is associated with distinct, generation-specific DNA methylation signatures, and highlight the potential value of epigenetic data for monitoring cryptic infection states in conservation-relevant systems.

Carotenoid- and melanin-based plumage coloration traits are key signals in avian communication and sexual selection as they are often thought to provide "honest" information about individual condition and fitness. These traits arise through distinct but interconnected physiological and genetic pathways. Recent work suggests that there may be a link between host-associated gut microbiota and the functional pathways leading to pigment-based plumage coloration, but this remains largely unexplored in wild populations. To address this gap, we tested whether variation in plumage coloration, as well as breeding condition, is associated with gut microbiome variation in wild populations of male Common Yellowthroats (Parulidae: Geothlypis trichas). We quantified multiple plumage coloration traits and characterized gut microbiome bacterial diversity using 16S rRNA metabarcoding. Through a comprehensive modeling framework, we found that individuals with brighter, more orange-tinted breast feathers and smaller cloacal protuberances (a proxy for breeding condition) exhibited higher gut microbiome diversity. At the taxonomic level, Methylobacterium-Methylorubrum, a carotenoid-producing bacteria, showed strong associations with multiple plumage traits, including mask area, breast feather hue, and saturation. Our results demonstrate that gut microbiome diversity is associated with variation in carotenoid-based coloration traits and breeding condition in Common Yellowthroats. More broadly, these results highlight the potential for host-microbiome interactions to shape phenotypic variation through physiological pathways in wild animal populations.

Landscape fragmentation, one of the leading drivers of biodiversity loss, can reshape both the genetics and microbiomes of wild populations. Although fragmentation is generally expected to limit gene flow and erode genetic diversity, and to disrupt host-associated microbial communities, these responses arise via different pathways and may therefore diverge within the same population. To understand how fragmentation simultaneously shapes population genetics and gut microbiomes, we analyzed fecal-derived host genomic and microbiome data from endangered, arboreal black howler monkeys (Alouatta pigra) across a fragmentation gradient. We then integrated these data with measures of ecological connectivity, habitat quality, and demography to identify the drivers of genetic and microbiome variation and structure. Multivariate analyses indicated that genetic patterns were shaped by both connectivity and habitat quality, whereas microbiome variation was driven mainly by habitat quality. Contrary to expectations under reduced realized connectivity with increasing isolation, monkeys showed the strongest gene flow signal in the most fragmented region, and higher genetic diversity and lower inbreeding than monkeys in continuous forest. Relatedness and isolation-by-distance patterns suggested that fragmentation has sex-specific effects on movement, disrupting the usual pattern of short-range male dispersal in the most fragmented region. Gut microbiomes, however, showed predicted negative responses to fragmentation: individuals in highly fragmented habitat had lower microbial diversity and compositional shifts consistent with lower-quality diets and increased exposure to disturbed environments. These results show contrasting biological responses to fragmentation within a single population, with genetic patterns likely resulting from compensatory behavioral flexibility and microbiome patterns reflecting local habitat degradation. Our findings underscore the need for conservation assessments that integrate multiple dimensions of population health rather than relying on any single indicator of fragmentation impact.

Long-term persistence of managed species will depend, in part, on whether the species harbors the physiological or genetic potential to adjust to warming temperatures, and whether relevant genetic variation is modified by management practices. The critically endangered Delta Smelt (Hypomesus transpacificus) is intensively managed, but little is known about the presence of genetic variation for resistance to elevated temperature, which will be important to maintain for their persistence in a rapidly warming future. Using a pedigree and whole genome sequencing data, we characterized the genetic variation and genomic architecture for CTMax (as a metric of upper thermal tolerance) across control and elevated rearing temperatures, alongside covarying traits (body size, degree of hatchery ancestry). Warmer rearing temperatures increased CTMax through acclimation but also resulted in reduced additive genetic variation for the trait, which could constrain adaptation under thermal stress. We found that larger fish had reduced CTMax, although this effect was diminished at elevated temperatures. We observed modest heritability for CTMax at rearing temperatures of 15{degrees}C and 18{degrees}C (0.26 and 0.16, respectively), but only a limited number of loci were identified that had consistent effects on CTMax across rearing temperatures. Instead, the genomic basis of thermal tolerance was highly dependent on rearing temperature (many loci detected with a GxE effect). The influence of domestication selection was indicated by changes in allele frequency, and divergence in upper thermal tolerance and plasticity, between low and high hatchery ancestry groups. Minimal overlap between loci associated with domestication and CTMax suggests that these traits possess separate genetic underpinnings. Knowledge of genetic variation supporting ecologically relevant physiological variation may be useful for refuge management and may inform supplementation in an ever-warming environment.

The coast redwood (Sequoia sempervirens) is a long-lived, hexaploid conifer of high ecological, cultural, and economic value whose range has been greatly reduced by historical logging. Effective restoration and conservation depend on understanding patterns of genetic differentiation across the redwood range to delineate populations for management prioritization. Yet, past range-wide studies provided only a partial picture of population structure in coast redwood as they relied on a limited set of genetic markers or limited sampling, as sequencing was done on the same range-wide provenance collection. Here, we analyze 334,029 SNPs from a new range-wide set of 224 individuals using a dosage-based approach that accounts for polyploidy. Principal coordinates and neighbor-joining analyses reveal clear latitudinal genetic differentiation, with a distinct break south of San Francisco Bay. Outlier SNP analysis indicates new candidate loci involved in salinity tolerance, climate stress response, and nutrient uptake, suggesting potential local adaptation. These results point to the central role of geography in shaping genetic variation in coast redwood and give scientific basis for designing new conservation strategies and future experiments, including assisted migration, provenance trials, and restoration planning aimed at preserving the species into the future.

Genomic forecasting approaches based on genotype-environment associations (GEAs) are increasingly used to estimate genomic offsets (GOs), which predict population maladaptation and extinction risk under current or future climatic conditions. Despite their widespread use, only a subset of studies have evaluated how accurately GOs predict (mal)adaptation, limiting their interpretation and application in policy and management. Here, we used GEA analyses to estimate GOs for past, present, and future climates in Lycaeides butterflies, focusing on the causes of variation in GOs among populations and their relationships with demographic parameters inferred from population genomic data. Using multivariate linear regression and genotyping-by-sequencing data from 42 Lycaeides populations (922 butterflies), we found that mean annual temperature, cumulative annual precipitation, and hybridization history together explained 47.6% of variation in genome-wide allele frequencies. Genomic offsets differed substantially among populations and across past, present, and future climates, with evidence for increasing maladaptation under more distant future climate scenarios. We found no relationship between GOs for present climates and contemporary effective population size. In contrast, genetic diversity, which reflects long-term effective population size, and local rates of gene flow together explained 27.3% of variation in contemporary GOs. Populations with higher genetic diversity and more gene flow exhibited lower GOs, consistent with the hypothesis that genetic diversity enhances adaptive capacity and that gene flow may introduce adaptive alleles. Overall, our results support the utility of GO predictions, particularly when validated with independent measures of adaptation, while cautioning against simplistic interpretations of GO as a direct measure of maladaptation in conservation and management contexts.

Invasive alien species (IAS) represent a major threat to biodiversity, human health, and economy. Yet the role of host-associated microbiomes in invasion processes remains poorly understood in natural systems. Here, we investigated the bacterial and fungal microbiomes of IAS and native Drosophila species collected along an anthropization gradient in French Guiana. Analyses of Drosophila communities revealed that IAS establishment is limited outside coastal habitats. Using bacterial 16S rRNA and fungal ITS metabarcoding of pooled individuals, we assessed microbial diversity, composition, and structure across host species, host category (invasive vs. native), and locality. Bacterial communities associated with IAS harboured reduced alpha diversity and distinct community structure compared to native Drosophila species, and host phylogenetic relatedness and locality also contributed to bacteriome variation. In contrast, fungal communities were mainly structured by locality and host species. We further identified core and exclusive taxa, compared bacteriome composition between native and IAS hosts using global reference datasets, and integrated information from the microbiome literature. This approach led to the identification of 45 bacterial and fungal candidate taxa potentially associated with host adaptation or competitiveness across coastal and rainforest environments. Overall, our results highlight microbiome variation as a potential component of invasion dynamics in Neotropical Drosophila.

Hybridisation between wild and domestic taxa can favour the spread of domestic alleles into wild populations through backcrossing. The complex interplay of random genetic drift, recombination, and selection can shape the fate of introgressed alleles. Maladaptive domestic variants are likely to be purged by natural selection, but others may persist across generations. It has long been known that the Apennine Italian wolf population, exposed to large numbers of free-ranging dogs, has experienced extensive introgression. The unusually high frequency of black wolves observed in Italy, compared to other European populations, may parallel patterns documented in North American wolves, where the melanistic KB allele at the CBD103 gene, of domestic origin, has spread over thousands of years of introgression. We tested whether the KB mutation entered the peninsular Italian wolf population via hybridisation and spread through adaptive introgression. Genome-wide analyses of black and wild-type (grey-coated) Apennine wolves showed no clear signatures of recent dog ancestry in most melanistic animals. Our ancestry reconstruction approaches identified two distinct KB haplogroups of domestic origin, suggesting multiple introgression events. Notably, we found molecular evidence consistent with balancing selection on the KB haplotypes, whose functional role, nonetheless, warrants further research. Therefore, the microevolutionary genomic and ecological consequences of wolf-dog hybridisation in Italy should be carefully investigated to inform appropriate science-based conservation management strategies.

The unprecedented rate of climate warming threatens many species, and assessing their vulnerability to climate change represents a critical challenge in conservation biology. The Apollo butterfly, an emblematic mountain species, is expected to be impacted by climate change. Here, we analysed thousands of SNPs from 101 localities across Apollo French distribution. We identified 93 SNPs strongly associated with climate variation using five genotype-environment association analyses. We forecasted future climate maladaptation of French Apollo populations using four genomic offset methods and integrated these results with neutral and adaptive genetic diversity, genetic structure and adaptive climatic niches to infer their vulnerability to climate change. Jura and Alps populations exhibited the lowest risk of vulnerability to climate change, with low genomic offsets, high genetic diversity and connectivity, whereas Auvergne populations showed the highest genomic offsets and lowest neutral and adaptive genetic diversity. Only a reduced percentage (<1%) of the current distribution is predicted to face climatic conditions outside the current range, suggesting that adaptive variability required to adapt to future climates may already be present, and that assisted gene flow could represent an effective conservation strategy. Finally, we discuss some of the main challenges of genomic forecasts, particularly for declining non-model species.

The potential for environmental change to compound the detrimental effects of inbreeding depression in small and isolated populations is a significant concern in conservation biology. Previous evidence syntheses suggested that environmental stress exacerbates inbreeding depression, but were based on limited data. Here, we comprehensively test the relationship between inbreeding depression and environmental stress in natural populations using Bayesian mixed-effects meta-analysis on a large, high-quality data set of 2127 inbreeding depression effect sizes from animals and plants. Our results show that inbreeding depression is significantly higher in benign than in stressful environments. Analyses of both inbreeding depression and stress-induced changes in genetic load supported a unimodal (humped) relationship between the costs of inbreeding and stress intensity, with a peak at intermediate stress. At the highest levels of stress there was, on average, a significantly greater inbreeding load in benign than in stressful environments. We suggest that the lower cost of inbreeding associated with extreme stress results from constraints on the expression of inbreeding depression as fitness and phenotypes decline towards zero. Our findings help to resolve long-standing uncertainty around how inbreeding and environmental change interact, revealing that inbreeding responses vary non-linearly with environmental stress intensity, but showing that stress does not generally amplify inbreeding depression. As such, they will inform both the management of populations of conservation concern and predictions of species responses to global environmental change.

True lemmings (genus Lemmus) underwent substantial range shifts during the Late Pleistocene and the Pleistocene-Holocene transition, but the impact of these events on present-day diversity remains poorly understood. Here, we used whole-genome sequencing data from modern and ancient samples across the Palearctic range to address this knowledge gap. Reconstruction of autosomal phylogeny revealed that Palearctic true lemmings exhibit relatively shallow genetic structure, contrasting with the deep divergence inferred from mitochondrial genomes. Genetic variation largely follows an isolation-by-distance pattern, and no elevated nuclear divergence was detected between the major mitochondrial lineages. Window-based phylogenetic analyses identified several peripheral populations with high concordance factors, including Norway and Amur lemmings. The high degree of phylogenetic concordance along the genome in these populations is likely a consequence of postglacial bottlenecks and isolation, as indicated by reduced heterozygosity and the presence of runs of homozygosity in them. Overall, our results indicate that the modern genomic structure of Palearctic lemmings was shaped primarily by range fragmentation and population isolation following the broad distribution of the genus during the Last Glacial Maximum. Thus, the current genetic structure appears to represent only a fraction of the Late Pleistocene true lemming diversity. This is illustrated by a genetically distinct ancient specimen ([~]40 ka BP) from the Indigirka River basin that does not cluster with any modern lineage. From a taxonomic perspective, these findings do not support strong species-level differentiation among the major Palearctic lineages and highlight the discrepancy between mitochondrial and nuclear patterns of diversity within the genus.

Genomic variation within populations reflects both past and contemporary evolutionary processes. The genetic structure of the European peatland plant species is shaped by complex postglacial recolonization and recent habitat loss. Here, we investigate population genomic patterns in the vulnerable mire plant, round-leaved sundew (Drosera rotundifolia L.). Using ddRAD sequencing of 311 individuals from 38 populations across Europe, we detected significant genetic differentiation (Fst = 0.02-0.44, p < 0.05) and a consistent deficit of heterozygosity (Fis = 0.248) across populations. In contrast, sequencing of 10,449 bp of chloroplast DNA revealed extremely low variation, with only one SNP detected. Clustering analyses (Admixture and DAPC) both identified pronounced genetic structure comprising three major clusters (Western, Northern, and Eastern) that broadly correspond to European biogeographic regions. Genetic differentiation was partially explained by geographic distance (3.3%; p = 0.0001; Mantel test), while climatic variables, particularly temperature and precipitation, accounted for 2.9% of genomic variation (p < 0.0001; redundancy analysis). The observed pattern is consistent with polygenic responses to climatic gradients and is supported by 1,022 SNPs, distributed across 632 loci, that were significantly associated with environmental variables. Demographic reconstruction revealed distinct evolutionary trajectories among clusters, with the Western cluster showing a more recent expansion. Together, our results suggest that the genetic structure of D. rotundifolia is a result of postglacial recolonization, geographic isolation, and climate influence. We highlight the importance of conserving genetic diversity of D. rotundifolia across all three clusters and accounting for potential local adaptation and population vulnerability in future conservation strategies.

Understanding spatial and temporal connectivity among individuals with different migration strategies is essential for migratory ecology and effective conservation, yet it often requires integrating multiple data sources. In Northeast Atlantic European sea bass (Dicentrarchus labrax), electronic tagging has revealed partial migration, with both resident and long-distance migrants showing fidelity to summer feeding and winter spawning areas. However, the role of regional spawning-site philopatry in shaping migration patterns and stock connectivity remains unclear. Here, we combine reconstructed migration trajectories with genome-wide analyses of gene flow and recent relatedness in 708 individuals sampled from 10 French Atlantic locations. We identify a seasonally shifting genetic discontinuity between the Bay of Biscay (BOB) and Northern (NS) stocks, located off western Brittany during winter spawning and displaced northeastward into the central English Channel during summer feeding. Despite seasonal mixing in the English Channel, an association between individual genetic composition and spawning-site selection supports regional spawning-site philopatry. Analyses of long genomic segments shared identical-by-descent reveal substantially greater connectivity within stocks than between stocks, indicating that philopatry constrains effective gene flow despite seasonal mixing. Reanalysis of independent genomic data further shows that sea bass from the northern Atlantic range predominantly belong to the Northern stock. Together, these results show how seasonal movements reshape spatial genetic structure while maintaining demographic subdivision, with direct implications for fisheries management.

Species complex demographic history, including population size changes, isolation and gene flow, shapes patterns of genetic variation and deleterious load. In small and declining populations, understanding these processes is critical for predicting inbreeding depression and extinction risk. The Rices whale (Balaenoptera ricei) is a newly described baleen whale species resident to the heavily industrialized Gulf of Mexico, with a current abundance estimate of 51 (95 % CI: 20-130) individuals, making it one of the most endangered baleen whales globally. Using whole-genome sequences from 25 individuals, we reconstructed the evolutionary and demographic history of Rices whale and assessed its genomic health. Our analyses reinforce its distinctiveness from Brydes whales, and suggest that Rices whale has persisted as a small and isolated population in the Gulf of Mexico for tens of thousands of years. Despite its long-term small effective population size, genomes show modest impacts of inbreeding, including few long runs of homozygosity. We detected a distinct pulse of introgression [~]350 years ago from a Brydes whale-like lineage that resulted in windows of elevated heterozygosity in Rices whale, though it did not alter the burden of deleterious variation. Forward simulations indicate that a recent population collapse to [~]100 breeding individuals places the species at high risk of future inbreeding and genomic erosion unless population growth occurs. These findings highlight that while gene flow can increase genetic diversity, demographic recovery is essential to mitigate long-term genomic risks, underscoring the importance of management actions that promote sustained population growth.

Thermal history provenancing can guide the choice of parental broodstock for selective breeding of corals from distinct reefs and has been proposed as an intervention for enhancing climate resilience. However, the genetic and molecular mechanisms underlying resultant offspring responses to heat stress, particularly during early life stages, remain poorly understood. Here, we generated Acropora tersa larvae and recruits by crossing parental colonies from the historically warmer Martin Reef and cooler Davies Reef and assessed the effects of within- and between-reef crosses on genetic diversity and transcriptional responses to heat stress. Genome-wide single nucleotide polymorphism analyses showed that broodstock from Martin and Davies Reefs were weakly differentiated (FST = 0.008) and exhibited comparable heterozygosity, as did all larval offspring groups. Transcriptomic analyses of recruits exposed to heat stress (32 {degrees}C for 36 days) revealed that both within- and between-reef offspring groups activated conserved stress-response pathways, with seven genotype-independent heat-responsive genes detected across all offspring groups. Differential expression and enrichment analyses showed induction of defence, protein homeostasis, intracellular transport, and metabolic processes alongside repression of growth- and signalling-related functions, consistent with the Type A General Coral Stress Response. Taken together, these findings suggest that the benefits of thermal history provenancing-informed selective breeding may be limited in low-differentiation systems and that targeted pre-screening of broodstock may help capture functional genetic variation relevant to restoration applications.

Understanding the drivers of genomic health and their consequences for population viability is often overlooked but potentially important to effective conservation amidst the biodiversity crisis of the Anthropocene. Leatherback turtle (Dermochelys coriacea) populations have declined globally due to anthropogenic factors, with some populations losing over 90% of their abundance over the past 30-50 years. While conservation efforts have been successful in stabilizing some populations, others continue to decline, and the reasons for these differential trajectories remain unclear. To assess how recent demographic factors, such as population size and decline, influence population genomic health, we combined population monitoring information with medium depth whole-genome and reduced representation resequencing data from globally representative populations. We found that small-stable populations have lower genomic diversity and higher inbreeding than large declining populations, reflecting prolonged small population sizes and limited gene flow. Yet, small-stable populations also show evidence of deleterious allele purging, suggesting genetic resilience. This, combined with lack of detectable genomic erosion over the study period, provides hope for potential recovery of healthy leatherback populations provided that anthropogenic threats are effectively mitigated. However, potential time lags and possible recent increases in inbreeding among close relatives in recently declined populations warrant continued monitoring and assessment. Genomic and abundance-based metrics were less aligned following rapid population declines, emphasizing the different timescales of the evolutionary and demographic processes they reflect, respectively, and the strength in their complementary, integrative use for extinction risk assessments. This also supports that it is not too late to turn the tide for recently declined leatherback populations and that continued investment in conservation efforts and threat reductions are warranted. Collectively, our results highlight how recent and historical demography shapes current genomic health and recovery potential in leatherback turtles, aids understanding of current risks and informs future conservation and management strategies.

Strong population contractions can leave a persistent genomic legacy that can influence populations long after their demographic recovery. While bottlenecks facilitate the removal of strongly deleterious mutations, the effectiveness of purging may be limited in historically small populations. The Kirtlands warbler (Setophaga kirtlandii) is a rare North American songbird with an ancestrally small population. After narrowly evading extinction, they are one of few species that have been delisted from federal protections in the USA. Despite their recovery, a previous study showed evidence for recent inbreeding and a high burden of deleterious mutations that may have not been purged despite strong bottlenecks. Historical DNA offers a unique opportunity to understand the consequences of recent demographic declines on genetic diversity. Here, we use DNA from over 100-year-old museum specimens to estimate changes in genetic load in the Kirtlands warblers pre- and post-bottleneck. We validate our results with forward-in-time genetic simulations and explore how sample size and missing data can affect estimates. Both empirical data and simulations suggest a reduced ability to purge deleterious mutations in this historically small population. Our simulations also highlight that limited sampling design and data quality can constrain the ability to detect changes.

AbstractsThe Japanese giant salamander (Andrias japonicus), an apex predator and a Special Natural Monument in Japan, is threatened by hybridization with introduced Chinese giant salamanders (Andrias davidianus). This hybridization has caused genetic introgression and expansion of hybrid populations, posing a serious conservation risk. Because morphological identification of hybrids is occasionally unreliable and current genetic methods rely on limited markers, a genome-wide approach is required. However, the extremely large genome ([~]50 Gb) of giant salamanders has hindered whole-genome analyses. In this study, we conducted transcriptome-based analyses of Japanese giant salamanders, Chinese giant salamanders, and their hybrids, generating RNA-seq data from 34 individuals. A total of over 419,000 SNP candidates were identified, from which 4,457 high-confidence SNPs in highly expressed genes were selected for analysis. Population structure analyses for Nabari colony revealed that hybrid individuals form two major groups, corresponding to different degrees of genetic contribution from Japanese and Chinese lineages. Most hybrids were inferred to be F2 or backcross individuals, while F1 hybrids were rare. Mitochondrial analysis indicated that all hybrids possessed Japanese-type mitochondrial genome, suggesting male-mediated introgression from Chinese salamanders. Differential expression analysis revealed enhanced stress-response pathways in hybrids and stronger antiviral responses in Japanese individuals. Using the axolotl genome as a reference, we constructed a virtual chromosomal map, identifying large haplotype blocks and supporting recent hybridization with limited recombination. This study provides a genome-wide framework for understanding hybridization dynamics and supports future conservation and evolutionary studies.

Experimental introductions of anadromous stickleback into freshwater habitats lacking this species allow analysis of the process of adaptation to freshwater forward-in-time. We examined the population genomic dynamics during early stages of adaptation in three replicate lakes that were experimentally founded, each using [~]3000 anadromous ancestors. We replicated earlier results that rare individuals carrying large haploblocks of freshwater-adaptive alleles (jackpot carriers) provide most of the allelic variation for adaptation of anadromous Threespine Stickleback to freshwater within only a few generations in each lake. There were population bottlenecks two to three generations after founding in each lake, after which jackpot carriers dramatically increased in frequency and came to dominate the populations. Individuals lacking large adaptive haploblocks experienced low fitness in their new freshwater environments, consistent with our previous report based on a single lake population. Despite similarities of the demographic responses to selection, the alleles that were most common among jackpot carriers were different in each population, suggesting that each lake population likely adapted to conditions in freshwater environments through different genes. These results provide direct evidence for the genomic mechanisms underlying the rapid adaptation of anadromous sticklebacks to freshwater environments, a process that can occur within just a few generations.