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Abstract Background Cis-regulatory mutations often underlie phenotypic evolution. However, because identifying the locations of promoters and enhancers in non-coding regions is challenging, we have fewer examples of identified causative cis-regulatory mutations that underlie naturally occurring phenotypic variations than of causative amino acid-altering mutations. Because cis-regulatory elements have epigenetic marks of specific histone modifications, we can detect cis-regulatory elements by mapping and analyzing them. Here, we investigated histone modifications and chromatin accessibility with cleavage under targets and tagmentation (CUT&Tag) and assay for transposase-accessible chromatin-sequencing (ATAC-seq). Results Using the threespine stickleback (Gasterosteus aculeatus) as a model, we confirmed that the genes for which nearby regions showed active marks, such as H3K4me1, H3K4me3, and high chromatin accessibility, were highly expressed. In contrast, the expression levels of genes for which nearby regions showed repressive marks, such as H3K27me3, were reduced, suggesting that our chromatin analysis protocols overall worked well. Genomic regions with peaks of histone modifications showed higher nucleotide diversity within and between populations. By comparing gene expression in the gills of the marine and stream ecotypes, we identified several insertions and deletions (indels) with transposable element fragments in the candidate cis-regulatory regions. Conclusions Thus, mapping and analyzing histone modifications can help identify cis-regulatory elements and accelerate the identification of causative mutations in the non-coding regions underlying naturally occurring phenotypic variations.

Species occupying dryland river ecosystems often experience "boom‐and‐bust" demographic cycles that coincide with shifts in habitat availability. Knowing whether declines are within natural thresholds versus those caused by acute human disturbance is critical for managing protected species. We investigated temporal shifts in abundance and habitat use of an endangered population of the threespine stickleback Gasterosteus aculeatus in southern California, where a Mediterranean climate leads to ephemeral habitat in one of the regions' least hydrologically modified rivers, the Santa Clara River. We conducted population surveys over a period of below‐average rainfall in the upper watershed in Soledad Canyon, with predefined reaches surveyed multiple times per year to capture different hydrologic conditions. Abundances were stable across years but varied significantly depending on location, with some reaches remaining dry and others drying seasonally to varying degrees. Occupancy models showed that the presence of stable perennial reaches, drying regime, and other site‐specific factors were important predictors of habitat use, and that certain reaches may be key to ensuring source‐sink dynamics as flow dissipates over the dry season. Low occupancy in two sections was driven by different predominant mechanisms, one by diel cycles of evapotranspiration and the other by cattails (Typha spp.), with both having greater effects during the hotter, drier parts of the year. As dryland river ecosystems are vulnerable to the effects of anthropogenic‐induced climate change, this study demonstrates how temporal monitoring can delimit dry‐state benchmarks for improving management interventions (i.e., translocation and habitat restoration) for protected species under conditions that are predicted to worsen in the coming years. [ABSTRACT FROM AUTHOR]

Mate choice plays a fundamental role in speciation, yet we know little about the molecular mechanisms that underpin this crucial decision-making process. Stickleback fish differentially adapted to limnetic and benthic habitats are reproductively isolated and females of each species use different male traits to evaluate prospective partners and reject heterospecific males. Here, we integrate behavioural data from a mate choice experiment with gene expression profiles from the brains of females actively deciding whether to mate. We find substantial gene expression variation between limnetic and benthic females, regardless of behavioural context, suggesting general divergence in constitutive gene expression patterns, corresponding to their genetic differentiation. Intriguingly, female gene co-expression modules covary with male display traits but in opposing directions for sympatric populations of the two species, suggesting male displays elicit a dynamic neurogenomic response that reflects known differences in female preferences. Furthermore, we confirm the role of numerous candidate genes previously implicated in female mate choice in other species, suggesting evolutionary tinkering with these conserved molecular processes to generate divergent mate preferences. Taken together, our study adds important new insights to our understanding of the molecular processes underlying female decision-making critical for generating sexual isolation and speciation. [ABSTRACT FROM AUTHOR]

Background: Cis-regulatory mutations often underlie phenotypic evolution. However, because identifying the locations of promoters and enhancers in non-coding regions is challenging, we have fewer examples of identified causative cis-regulatory mutations that underlie naturally occurring phenotypic variations than of causative amino acid-altering mutations. Because cis-regulatory elements have epigenetic marks of specific histone modifications, we can detect cis-regulatory elements by mapping and analyzing them. Here, we investigated histone modifications and chromatin accessibility with cleavage under targets and tagmentation (CUT&Tag) and assay for transposase-accessible chromatin-sequencing (ATAC-seq). Results: Using the threespine stickleback (Gasterosteus aculeatus) as a model, we confirmed that the genes for which nearby regions showed active marks, such as H3K4me1, H3K4me3, and high chromatin accessibility, were highly expressed. In contrast, the expression levels of genes for which nearby regions showed repressive marks, such as H3K27me3, were reduced, suggesting that our chromatin analysis protocols overall worked well. Genomic regions with peaks of histone modifications showed higher nucleotide diversity within and between populations. By comparing gene expression in the gills of the marine and stream ecotypes, we identified several insertions and deletions (indels) with transposable element fragments in the candidate cis-regulatory regions. Conclusions: Thus, mapping and analyzing histone modifications can help identify cis-regulatory elements and accelerate the identification of causative mutations in the non-coding regions underlying naturally occurring phenotypic variations. [ABSTRACT FROM AUTHOR]

Duplicated genes provide the opportunity for evolutionary novelty and adaptive divergence. In many cases, having more gene copies increases gene expression, which might facilitate adaptation to stressful or novel environments. Conversely, overexpression or misexpression of duplicated genes can be detrimental and subject to negative selection. In this scenario, newly duplicate genes may evade purifying selection if they are epigenetically silenced, at least temporarily, leading them to persist in populations as copy number variations (CNVs). In animals and plants, younger gene duplicates tend to have higher levels of DNA methylation and lower levels of gene expression, suggesting epigenetic regulation could promote the retention of gene duplications via expression repression or silencing. Here, we test the hypothesis that DNA methylation variation coincides with young duplicate genes that are segregating as CNVs in six populations of the three‐spined stickleback that span a salinity gradient from 4 to 30 PSU. Using reduced‐representation bisulfite sequencing, we found DNA methylation and CNV differentiation outliers rarely overlapped. Whereas lineage‐specific genes and young duplicates were found to be highly methylated, just two gene CNVs showed a significant association between promoter methylation level and copy number, suggesting that DNA methylation might not interact with CNVs in our dataset. If most new duplications are regulated for dosage by epigenetic mechanisms, our results do not support a strong contribution from DNA methylation soon after duplication. Instead, our results are consistent with a preference to duplicate genes that are already highly methylated. [ABSTRACT FROM AUTHOR]

Extreme temperature events, such as heat waves, can have lasting effects on the behavior, physiology, and reproductive success of organisms. Here, we examine the impact of short-term exposure to a simulated heat wave on condition, parental care, and reproductive success in a population of threespine stickleback (Gasterosteus aculeatus), a small fish with exclusive paternal care, currently experiencing regular heat waves. Males were either exposed to a simulated heat wave (23 °C) for 5 d or held at an ideal temperature (18 °C). Following this 5-d treatment, all males were transferred to 18 °C, where they completed a full parenting cycle. Offspring were raised at 18 °C. We found that while mass and body condition were unaffected in males exposed to a heat wave, cortisol responses were dampened across the nesting cycle compared to control males. In addition, heat wave males had longer latency for eggs to hatch, lower hatching success, and showed lower levels of parental care behavior compared to control males. Offspring of heat wave males had lower body condition, affecting swimming performance. Altogether, our results highlight the long-term impact that even short-term events can have on reproductive success, parental behavior, and subsequent generations, providing insight into population responses to rapid environmental change. [ABSTRACT FROM AUTHOR]

Two forms of gigantism are differentiated in fish, Brobdingnagian and Goliathan gigantism, the former applying to populations whose individuals are all larger than is typical for the taxon, the latter to single individuals within a population. While Brobdingnagian gigantism is largely explained by various ecological and evolutionary rules, Goliathan gigantism is not. A mechanistic hypothesis is proposed which explains Goliathan gigantism in terms of the reduction of oxygen requirements of individual fish via moving to cooler temperatures and/or acquiring larger, more energy‐dense prey, which enable them to get bigger, and, in the process, sometimes generate bimodal size distributions that may qualify as gradual forms between Goliathan and Brobdingnagian gigantism. This mechanism, which relies on the manner in which their gill surface area grows, is more likely to operate in fish that can get big in the first place than in fish that remain small. [ABSTRACT FROM AUTHOR]

Sculpins (coastrange and slimy) and sticklebacks (ninespine and threespine) are widely distributed fishes cohabiting 2 south-central Alaskan lakes (Aleknagik and Iliamna), and all these species are parasitized by cryptic diphyllobothriidean cestodes in the genus Schistocephalus. The goal of this investigation was to test for host-specific parasitic relationships between sculpins and sticklebacks based upon morphological traits (segment counts) and sequence variation across the NADH1 gene. A total of 446 plerocercoids was examined. Large, significant differences in mean segment counts were found between cestodes in sculpin (mean = 112; standard deviation [s.d.] = 15) and stickleback (mean = 86; s.d. = 9) hosts within and between lakes. Nucleotide sequence divergence between parasites from sculpin and stickleback hosts was 20.5%, and Bayesian phylogenetic analysis recovered 2 well-supported clades of cestodes reflecting intermediate host family (i.e. sculpin, Cottidae vs stickleback, Gasterosteidae). Our findings point to the presence of a distinct lineage of cryptic Schistocephalus in sculpins from Aleknagik and Iliamna lakes that warrants further investigation to determine appropriate evolutionary and taxonomic recognition.

Fish species are commonly used as experimental models in the laboratory. DNA is routinely collected from these animals to permit identification of their genotype. The current standard procedure to sample DNA is fin clipping, which involves anaesthetising individuals and removing a portion of the caudal fin. While fin clipping reliably generates good quality DNA samples for downstream applications, there is evidence that it can alter health and welfare, and impact the fish’s behaviour. This in turn can result in greater variation in the data collected. In a recent study we adapted a skin swabbing protocol to collect DNA from small-bodied fish, including sticklebacks and zebrafish, without the use of analgesics, anaesthetics or sharp instruments. A rayon-tipped swab was used to collect mucus from the flank of the fish, which was then used for DNA extraction. We subsequently demonstrated that compared to fin clipping, skin swabbing triggered fewer changes in stress axis activation and behaviour. We also found that gene expression and behaviour data collected from swabbed fish were less variable than similar data collected from fish that had been fin clipped. This potentially allows smaller sample sizes in experimental groups to be used after skin swabbing, thereby reducing animal use. Here we provide a detailed protocol explaining how to collect DNA samples from small laboratory fish using skin swabs.

Development and regeneration are orchestrated by gene regulatory networks that operate in part through transcriptional enhancers. Although many enhancers are pleiotropic and are active in multiple tissues, little is known about whether enhancer pleiotropy is due to 1) site pleiotropy, in which individual transcription factor binding sites (TFBS) are required for activity in multiple tissues, or 2) multiple distinct sites that regulate expression in different tissues. Here, we investigated the pleiotropy of an intronic enhancer of the stickleback Bone morphogenetic protein 6 (Bmp6) gene. This enhancer was previously shown to regulate evolved changes in tooth number and tooth regeneration, and is highly pleiotropic, with robust activity in both fins and teeth throughout embryonic, larval, and adult life, and in the heart and kidney in adult fish. We tested the hypothesis that the pleiotropy of this enhancer is due to site pleiotropy of an evolutionarily conserved predicted Foxc1 TFBS. Transgenic analysis and site-directed mutagenesis experiments both deleting and scrambling this predicted Foxc1 TFBS revealed that the binding site is required for enhancer activity in both teeth and fins throughout embryonic, larval, and adult development, and in the heart and kidney in adult fish. Collectively these data support a model where the pleiotropy of this Bmp6 enhancer is due to site pleiotropy and this putative binding site is required for enhancer activity in multiple anatomical sites from the embryo to the adult.

The mechanisms underlying the divergence of reproductive strategies between closely related species are still poorly understood. Additionally, it is unclear which selective factors drive the evolution of reproductive behavioral variation and how these traits coevolve, particularly during early divergence. To address these questions, we quantified behavioral differences in a recently diverged pair of Nova Scotian three-spined stickleback (Gasterosteus aculeatus) populations, which vary in parental care, with one population displaying paternal care and the other lacking this. We compared both populations, and a full reciprocal F1 hybrid cross, across four major reproductive stages: territoriality, nesting, courtship, and parenting. We identified significant divergence in a suite of heritable behaviors. Importantly, F1 hybrids exhibited a mix of behavioral patterns, some of which suggest sex linkage. This system offers fresh insights into the coevolutionary dynamics of reproductive behaviors during early divergence and offers support for the hypothesis that coevolutionary feedback between sexual selection and parental care can drive rapid evolution of reproductive strategies. [ABSTRACT FROM AUTHOR]

Determining how environmental conditions contribute to divergence among populations and drive speciation is fundamental to resolving mechanisms and understanding outcomes in evolutionary biology. Postglacial freshwater fish species in the Northern Hemisphere are ideal biological systems to explore the effects of environment on diversification in morphology, ecology, and genetics (ecomorph divergences) within lakes. To date, various environmental factors have been implicated in the presence of multiple ecomorphs within particular lakes or regions. However, concerted evidence for generalizable patterns in environmental variables associated with speciation across geographical regions and across species and genera has been lacking. Here, we aimed to identify key biotic and abiotic factors associated with ecological divergence of postglacial freshwater fish species into multiple sympatric ecomorphs, focusing on species in the well‐studied, widespread, and co‐distributed genera Gasterosteus, Salvelinus, and Coregonus (stickleback, charr, and whitefish, respectively). We found that the presence of multiple sympatric ecomorphs tended to be associated with increasing lake surface area, maximum depth, and nutrient availability. In addition, predation, competition, and prey availability were suggested to play a role in divergence into multiple ecomorphs, but the effects of biotic factors require further study. Although we identified several environmental factors correlated with the presence of multiple ecomorphs, there were substantial data gaps across species and regions. An improved understanding of these systems may provide insight into both generalizable environmental factors involved in speciation in other systems, and potential ecological and evolutionary responses of species complexes when these variables are altered by environmental change. [ABSTRACT FROM AUTHOR]

Abstract Duplicated genes provide the opportunity for evolutionary novelty and adaptive divergence. In many cases, having more gene copies increases gene expression, which might facilitate adaptation to stressful or novel environments. Conversely, overexpression or misexpression of duplicated genes can be detrimental and subject to negative selection. In this scenario, newly duplicate genes may evade purifying selection if they are epigenetically silenced, at least temporarily, leading them to persist in populations as copy number variations (CNVs). In animals and plants, younger gene duplicates tend to have higher levels of DNA methylation and lower levels of gene expression, suggesting epigenetic regulation could promote the retention of gene duplications via expression repression or silencing. Here, we test the hypothesis that DNA methylation variation coincides with young duplicate genes that are segregating as CNVs in six populations of the three‐spined stickleback that span a salinity gradient from 4 to 30 PSU. Using reduced‐representation bisulfite sequencing, we found DNA methylation and CNV differentiation outliers rarely overlapped. Whereas lineage‐specific genes and young duplicates were found to be highly methylated, just two gene CNVs showed a significant association between promoter methylation level and copy number, suggesting that DNA methylation might not interact with CNVs in our dataset. If most new duplications are regulated for dosage by epigenetic mechanisms, our results do not support a strong contribution from DNA methylation soon after duplication. Instead, our results are consistent with a preference to duplicate genes that are already highly methylated.

Fish species are commonly used as experimental models in the laboratory. DNA is routinely collected from these animals to permit identification of their genotype. The current standard procedure to sample DNA is fin clipping, which involves anaesthetising individuals and removing a portion of the caudal fin. While fin clipping reliably generates good quality DNA samples for downstream applications, there is evidence that it can alter health and welfare, and impact the fish’s behaviour. This in turn can result in greater variation in the data collected. In a recent study we adapted a skin swabbing protocol to collect DNA from small-bodied fish, including sticklebacks and zebrafish, without the use of analgesics, anaesthetics or sharp instruments. A rayon-tipped swab was used to collect mucus from the flank of the fish, which was then used for DNA extraction. We subsequently demonstrated that compared to fin clipping, skin swabbing triggered fewer changes in stress axis activation and behaviour. We also found that gene expression and behaviour data collected from swabbed fish were less variable than similar data collected from fish that had been fin clipped. This potentially allows smaller sample sizes in experimental groups to be used after skin swabbing, thereby reducing animal use. Here we provide a detailed protocol explaining how to collect DNA samples from small laboratory fish using skin swabs.

Differentiation of foraging traits among predator populations may help explain observed variation in the structure of prey communities. However, few studies have investigated the phenotypic effects of predators on their prey in natural communities. Here, we use a comparative analysis of 78 Greenlandic lakes to examine how foraging trait variation among threespine stickleback populations can help explain variation in zooplankton community composition among lakes. We find that landscape‐scale variation in zooplankton composition was jointly explained by lake properties, such as size and water chemistry, and the presence and absence of both stickleback and arctic char. Additional variation in zooplankton community structure can be explained by stickleback jaw protrusion, a trait with known utility for foraging on zooplankton, but only in lakes where stickleback co‐occur with arctic char. Overall, our results illustrate how trait variation of predators, alongside other ecosystem properties, can influence the composition of prey communities in nature. [ABSTRACT FROM AUTHOR]

The tradition of stocking whitefish (Coregonus spec.) is widespread throughout Europe and dates back more than a century in some lakes. Despite its broad acceptance as a management action, stocking success is less often studied and might change with shifts in ecosystems. Similar to many other large lakes in Central Europe, Lake Constance is stocked with millions of larvae each year at great cost. The last evaluation of this management practice took place in 2003, and since then several potential impacts on whitefish increased in magnitude (e.g. stickleback invasion, increasing water temperature). Forty‐two million dye‐marked larvae were stocked in 2016 and 36 million in 2018. Recaptures made 1.5 and 3.5 years after stocking revealed that around 20% of the 2016 and 2018 whitefish cohorts consisted of stocked whitefish. This amount is significantly lower than was found in previous experiments. To identify potential factors controlling recruitment, a Beverton‐Holt stock‐recruitment model showed that stocking did not significantly affect year‐class recruitment. We conclude that stocking is no longer cost‐effective, and we recommend that stocking should be reduced in the short term. Furthermore, our results suggest that explanatory variables that previously described recruitment variation of the whitefish population, such as April water temperature, may no longer apply. [ABSTRACT FROM AUTHOR]

Genome sequencing enables answering fundamental questions about the genetic basis of adaptation, population structure and epigenetic mechanisms. Yet, we usually need a suitable reference genome for mapping population‐level resequencing data. In some model systems, multiple reference genomes are available, giving the challenging task of determining which reference genome best suits the data. Here, we compared the use of two different reference genomes for the three‐spined stickleback (Gasterosteus aculeatus), one novel genome derived from a European gynogenetic individual and the published reference genome of a North American individual. Specifically, we investigated the impact of using a local reference versus one generated from a distinct lineage on several common population genomics analyses. Through mapping genome resequencing data of 60 sticklebacks from across Europe and North America, we demonstrate that genetic distance among samples and the reference genomes impacts downstream analyses. Using a local reference genome increased mapping efficiency and genotyping accuracy, effectively retaining more and better data. Despite comparable distributions of the metrics generated across the genome using SNP data (i.e. π, Tajima's D and FST), window‐based statistics using different references resulted in different outlier genes and enriched gene functions. A marker‐based analysis of DNA methylation distributions had a comparably high overlap in outlier genes and functions, yet with distinct differences depending on the reference genome. Overall, our results highlight how using a local reference genome decreases reference bias to increase confidence in downstream analyses of the data. Such results have significant implications in all reference‐genome‐based population genomic analyses. [ABSTRACT FROM AUTHOR]

The primary function of animal nests is to protect developing offspring from hostile and fluctuating environments. Animal builders have been shown to adjust nest construction in response to changes in their environment. However, the extent of this plasticity, and its dependence on an evolutionary history of environmental variability, is not well understood. To test whether an evolutionary history with flowing water impacts male ability to adjust nests in response to flow regime, we collected three-spined sticklebacks (Gasterosteus aculeatus) from three lakes and three rivers, and brought them into reproductive condition in controlled laboratory aquaria. Males were then allowed to nest under both flowing and static conditions. Nest building behaviour, nest structure and nest composition were all recorded. In comparison to males building nests under static conditions, males building in flowing water took longer to construct their nests and invested more in nesting behaviour. Moreover, nests built in flowing water contained less material, were smaller, more compact, neater and more elongated than nests built under static conditions. Whether males came from rivers or lakes had little impact on nesting activities, or male capacity to adjust behaviours in response to flow treatment. Our findings suggest that aquatic animals which have experienced a stable environment over a long period of time retain plasticity in nest-building behaviours that allow them to adjust nests to ambient flow conditions. This ability may prove crucial in coping with the increasingly unpredictable flow regimes found in anthropogenically altered waterways and those resulting from global climate change. This article is part of the theme issue 'The evolutionary ecology of nests: a cross-taxon approach'. [ABSTRACT FROM AUTHOR]

Host–parasite coevolution may lead to patterns of local adaptation in either the host or parasite. For parasites with complex multi‐host life cycles, this coevolution may be more challenging as they must adapt to multiple geographically varying hosts. The tapeworm Schistocephalus solidus exhibits some local adaptation to its second intermediate host, threespine stickleback, to which the parasite is strictly specialized. However, the tapeworm's adaptation to its first intermediate host (any of a number of copepod species) is not documented. We investigated if there was local adaptation and host specify in the tapeworm Schistocephalus solidus to its copepod first intermediate hosts. We exposed copepods from five lakes in Vancouver Island (BC, Canada) to local (i.e. same lake) and foreign tapeworms in a reciprocal exposure experiment. Results indicate that the tapeworm is not locally adapted to the copepods. Instead, we observed moderate‐effect host specificity, infection rates being higher in certain copepod species than in others. Infection rates also varied among cestode populations. These results show that although S. solidus infects multiple copepod genera, they are not equally competent hosts. Differences in S. solidus epidemiology among lakes is likely to be driven more by this partial specialization, than by local adaptation to first intermediate hosts. [ABSTRACT FROM AUTHOR]

Epigenetic mechanisms such as DNA methylation (DNAme) are thought to comprise an invaluable adaptive toolkit in the early stages of local adaptation, especially when genetic diversity is constrained. However, the link between genetic diversity and DNAme has been scarcely examined in natural populations, despite its potential to shed light on the evolutionary forces acting on methylation state. Here, we analyzed reduced-representation bisulfite sequencing and whole-genome pool-seq data from marine and freshwater stickleback populations to examine the relationship between DNAme variation (between- and within-population) and nucleotide diversity in the context of freshwater adaptation. We find that sites that are differentially methylated between populations have higher underlying standing genetic variation, with diversity higher among sites that gained methylation in freshwater than those that lost it. Strikingly, although nucleotide diversity is generally lower in the freshwater population as expected from a population bottleneck, this is not the case for sites that lost methylation, which instead have elevated nucleotide diversity in freshwater compared with marine. Subsequently, we show that nucleotide diversity is higher among sites with ancestrally variable methylation and also positively correlates with the sensitivity to environmentally induced methylation change. The results suggest that as selection on the control of methylation state becomes relaxed, so too does selection against mutations at the sites themselves. Increased epigenetic variance in a population is therefore likely to precede genetic diversification. [ABSTRACT FROM AUTHOR]

Loss of body armour, sometimes including a reduction in or loss of pelvic spines, is an adaptation observed in many isolated freshwater populations. Pelvic reduction in sticklebacks has previously been associated with recurrent, but variant, deletions within pelvic enhancer regions PelA and PelB, which regulate expression of the homeodomain transcription factor gene Pitx1. We investigated variation in nucleotide sequences of pelvic enhancers in sticklebacks collected from two small freshwater lakes in the same watercourse and a nearby marine site in subarctic Norway. Spineless, as well as asymmetrically spined and completely spined sticklebacks are present in the upper lake, while only specimens with complete spines are found at the other lake and the marine site. Observed variation at PelA between the three sites was mainly due to variable numbers of repeats at three fragile TG-repeat loci. The length of PelA, mainly at one of the TG-repeat loci, was consistently shorter among individuals in the upper lake compared with specimens from the two other sites. However, no obvious association was revealed between enhancer variants and pelvic status. No polymorphism was found at PelB. Thus, additional genetic factors and/or environmental cues need to be identified to fully explain the occurrence of pelvic reduction in sticklebacks in this lake. [ABSTRACT FROM AUTHOR]

Abstract The vertebrate sodium–iodide symporter (NIS or SLC5A5) transports iodide into the thyroid follicular cells that synthesize thyroid hormone. The SLC5A protein family includes transporters of vitamins, minerals, and nutrients. Disruption of SLC5A5 function by perchlorate, a pervasive environmental contaminant, leads to human pathologies, especially hypothyroidism. Perchlorate also disrupts the sexual development of model animals, including threespine stickleback (Gasterosteus aculeatus) and zebrafish (Danio rerio), but the mechanism of action is unknown. To test the hypothesis that SLC5A5 paralogs are expressed in tissues necessary for the development of reproductive organs, and therefore are plausible candidates to mediate the effects of perchlorate on sexual development, we first investigated the evolutionary history of Slc5a paralogs to better understand potential functional trajectories of the gene family. We identified two clades of slc5a paralogs with respect to an outgroup of sodium/choline cotransporters (slc5a7); these clades are the NIS clade of sodium/iodide and lactate cotransporters (slc5a5, slc5a6, slc5a8, slc5a8, and slc5a12) and the SGLT clade of sodium/glucose cotransporters (slc5a1, slc5a2, slc5a3, slc5a4, slc5a10, and slc5a11). We also characterized expression patterns of slc5a genes during development. Stickleback embryos and early larvae expressed NIS clade genes in connective tissue, cartilage, teeth, and thyroid. Stickleback males and females expressed slc5a5 and its paralogs in gonads. Single‐cell transcriptomics (scRNA‐seq) on zebrafish sex‐genotyped gonads revealed that NIS clade‐expressing cells included germ cells (slc5a5, slc5a6a, and slc5a6b) and gonadal soma cells (slc5a8l). These results are consistent with the hypothesis that perchlorate exerts its effects on sexual development by interacting with slc5a5 or its paralogs in reproductive tissues. These findings show novel expression domains of slc5 genes in stickleback and zebrafish, which suggest similar functions across vertebrates including humans, and provide candidates to mediate the effects of perchlorate on sexual development.

Populations are subjected to diverse environmental conditions that affect fitness and induce evolutionary or plastic responses, resulting in phenotypic divergence. Some authors contend that such divergence is concentrated along a single major axis of trait covariance even if that axis does not lead populations directly toward a fitness optimum. Other authors argue that divergence can occur readily along many phenotype axes at the same time. We use populations of threespine stickleback (Gasterosteus aculeatus) from 14 lakes with contrasting ecological conditions to find some resolution along the continuum between these two extremes. Unlike many previous studies, we included several functional suites of traits (defensive, swimming, trophic) potentially subject to different sources of selection. We find that populations exhibit dimensionality of divergence that is high enough to preclude a history of constraint along a single axis—both for divergence in multivariate mean trait values and for the structure of trait covariances. Dimensionality varied among trait suites and were strongly influenced by the inclusion of specific traits, and integration of trait suites varied between populations. We leverage this variation into new insights about the process of divergence and suggest that similar analyses could increase understanding of other adaptive radiations. [ABSTRACT FROM AUTHOR]

Submersion in the anaesthetic MS-222 is a well-established and effective method used during the euthanasia of fish, but the consequences of treatment with this anaesthetic for mitochondrial respiration are yet to be established. This is important to evaluate, as an increasing amount of research is conducting high-resolution respirometry to measure respiration across multiple species of fish, including looking at thermal sensitivity and mitochondrial responses to the warmer temperatures faced with climate change. Analysis often occurs after euthanasia with MS-222 without knowledge of how MS-222 itself affects any measured parameters of mitochondrial respiration, leaving potential for a misinterpretation of results. Here, high-resolution respirometry was conducted to explore how MS-222 affects oxidative phosphorylation in the brain and skeletal muscle of the three-spined stickleback, Gasterosteus aculeatus, which is a model species in evolutionary ecology. In the brain, differences in respiration were observed between three-spined sticklebacks euthanised with MS-222 and those where no anaesthetic was implemented. No differences between treatments were observed in the skeletal muscle, although variation between individuals was high and oxygen flux was lower than in the brain. Overall, this study highlights the need for a consistent method of euthanasia when conducting high-resolution respirometry in fish, as MS-222 may alter measures of oxidative phosphorylation. [ABSTRACT FROM AUTHOR]

The context and cause of adaptive radiations have been widely described and explored but why rapid evolutionary diversification does not occur in related evolutionary lineages has yet to be understood. The standard answer is that evolutionary diversification is provoked by ecological opportunity and that some lineages do not encounter the opportunity. Three‐spined sticklebacks on the Scottish island of North Uist show enormous diversification, which seems to be associated with the diversity of aquatic habitats. Sticklebacks on the neighboring island of South Uist have not been reported to show the same level of evolutionary diversity, despite levels of environmental variation that we might expect to be similar to North Uist. In this study, we compared patterns of morphological and environmental diversity on North and South Uist. Ancestral anadromous sticklebacks from both islands exhibited similar morphology including size and bony "armor." Resident sticklebacks showed significant variation in armor traits in relation to pH of water. However, North Uist sticklebacks exhibited greater diversity of morphological traits than South Uist and this was associated with greater diversity in pH of the waters of lochs on North Uist. Highly acidic and highly alkaline freshwater habitats are missing, or uncommon, on South Uist. Thus, pH appears to act as a causal factor driving the evolutionary diversification of stickleback in local adaptation in North and South Uist. This is consistent with diversification being more associated with ecological constraint than ecological opportunity. [ABSTRACT FROM AUTHOR]

Species competing for resources also commonly share predators. While competition often drives divergence between species, the effects of shared predation are less understood. Theoretically, competing prey species could either diverge or evolve in the same direction under shared predation depending on the strength and symmetry of their interactions. We took an empirical approach to this question, comparing antipredator and trophic phenotypes between sympatric and allopatric populations of threespine stickleback and prickly sculpin fish that all live in the presence of a trout predator. We found divergence in antipredator traits between the species: in sympatry, antipredator adaptations were relatively increased in stickleback but decreased in sculpin. Shifts in feeding morphology, diet and habitat use were also divergent but driven primarily by stickleback evolution. Our results suggest that asymmetric ecological character displacement indirectly made stickleback more and sculpin less vulnerable to shared predation, driving divergence of antipredator traits between sympatric species. [ABSTRACT FROM AUTHOR]

Development and regeneration are orchestrated by gene regulatory networks that operate in part through transcriptional enhancers. Although many enhancers are pleiotropic and are active in multiple tissues, little is known about whether enhancer pleiotropy is due to 1) site pleiotropy, in which individual transcription factor binding sites (TFBS) are required for activity in multiple tissues, or 2) multiple distinct sites that regulate expression in different tissues. Here, we investigated the pleiotropy of an intronic enhancer of the stickleback Bone morphogenetic protein 6 (Bmp6) gene. This enhancer was previously shown to regulate evolved changes in tooth number and tooth regeneration, and is highly pleiotropic, with robust activity in both fins and teeth throughout embryonic, larval, and adult life, and in the heart and kidney in adult fish. We tested the hypothesis that the pleiotropy of this enhancer is due to site pleiotropy of an evolutionarily conserved predicted Foxc1 TFBS. Transgenic analysis and site-directed mutagenesis experiments both deleting and scrambling this predicted Foxc1 TFBS revealed that the binding site is required for enhancer activity in both teeth and fins throughout embryonic, larval, and adult development, and in the heart and kidney in adult fish. Collectively these data support a model where the pleiotropy of this Bmp6 enhancer is due to site pleiotropy and this putative binding site is required for enhancer activity in multiple anatomical sites from the embryo to the adult. [Display omitted] • Enhancer in intron 4 of stickleback Bone morphogenetic protein 6 is pleiotropic. • Enhancer is active in multiple sites including teeth, fins, heart, and kidney. • Enhancer contains predicted Foxc1 transcription factor binding site (TFBS). • This TFBS is required for enhancer activity in all domains. • Results suggest further genetic similarities between tooth and hair regeneration. [ABSTRACT FROM AUTHOR]

Sex chromosomes vary greatly in their age and levels of differentiation across the tree of life. This variation is largely due to the rates of sex chromosome turnover in different lineages; however, we still lack an explanation for why sex chromosomes are so conserved in some lineages (e.g. mammals, birds) but so labile in others (e.g. teleosts, amphibians). To identify general mechanisms driving transitions in sex determination systems or forces which favour their conservation, we first require empirical data on sex chromosome systems from multiple lineages. Stickleback fishes are a valuable model lineage for the study of sex chromosome evolution due to variation in sex chromosome systems between closely‐related species. Here, we identify the sex chromosome and a strong candidate for the master sex determination gene in the brook stickleback, Culaea inconstans. Using whole‐genome sequencing of wild‐caught samples and a lab cross, we identify AmhY, a male specific duplication of the gene Amh, as the candidate master sex determination gene. AmhY resides on Chromosome 20 in C. inconstans and is likely a recent duplication, as both AmhY and the sex‐linked region of Chromosome 20 show little sequence divergence. Importantly, this duplicate AmhY represents the second independent duplication and recruitment of Amh as the sex determination gene in stickleback and the eighth example known across teleosts. We discuss this convergence in the context of sex chromosome turnovers and the role that the Amh/AmhrII pathway, which is crucial for sex determination, may play in the evolution of sex chromosomes in teleosts. [ABSTRACT FROM AUTHOR]

DNA samples are often used to identify fish before they are utilised in other experiments. Our recent research has shown that skin swabbing can be used to collect DNA for genotyping, and that swabbing causes less harm to fish than fin clipping, another common technique. In this study we investigated potential refinements to the skin swabbing protocol by pre-treating fish with the analgesic lidocaine. We could not detect any differences in cortisol release, behaviour or expression of stress axis marker genes in skin swabbed sticklebacks or zebrafish regardless of lidocaine application. In contrast, fin clipping caused changes in cortisol release, gene expression and behaviour when analgesia was not used. These changes were rescued by pre-treatment with lidocaine confirming that analgesia was effective. The results demonstrates that skin swabbing is a refined technique for DNA collection that does not require analgesia.

Abstract Background Sexual signals produced by males play a central role in sexual selection, but the relationship between these traits and the quality of the bearer are often ambiguous. Secondary sexual traits may represent genetic quality of the bearer, resulting in positive relationships with physiological state, or may be costly to produce, showing trade-off with physiological state. A number of studies have explored the relationships between secondary sexual traits and other functional traits, but few have studied their fitness consequences. We studied the link between diverse physiological traits and both morphological and behavioural sexual traits and examined how their interplay influences offspring viability in the three-spined stickleback. Results Male sticklebacks showing nest building and courtship behaviour were smaller than those not investing in reproductive activities. There was no evidence that the expression of red nuptial colouration and the quality of courtship behaviour of males are positively related to their metabolic rates, swim ability, oxidative damage and mtDNA copy number. However, individuals showing larger red nuptial colour areas had higher levels of oxidative DNA damage in their sperm. Male courtship behaviour and aggressiveness, but not red colour area, were good predictors of offspring hatching and survival. Conclusions Our results suggest that, in our study population at the southern edge of the species’ distribution, sexual colouration of male sticklebacks was not a good indicator of their body state, but both courtship quality and aggressiveness during the courtship are reliable cues of their gamete quality, influencing the viability of their offspring. Thus, females that choose mates based on their courtship behaviour will have high fitness. In the study population, which represents a fast pace-of-life with high reproductive rate and short lifespan, sexual ornaments of males may not honestly signal their physiological and physical state because they invest at maximum in a single reproductive season despite high costs.

Establishing how collective behaviour emerges is central to our understanding of animal societies. Previous research has highlighted how universal interaction rules shape collective behaviour, and that individual differences can drive group functioning. Groups themselves may also differ considerably in their collective behaviour, but little is known about the consistency of such group variation, especially across different ecological contexts that may alter individuals' behavioural responses. Here, we test if randomly composed groups of sticklebacks differ consistently from one another in both their structure and movement dynamics across an open environment, an environment with food, and an environment with food and shelter. Based on high-resolution tracking data of the free-swimming shoals, we found large context-associated changes in the average behaviour of the groups. But despite these changes and limited social familiarity among group members, substantial and predictable behavioural differences between the groups persisted both within and across the different contexts (group-level repeatability): some groups moved consistently faster, more cohesively, showed stronger alignment and/or clearer leadership than other groups. These results suggest that among-group heterogeneity could be a widespread feature in animal societies. Future work that considers group-level variation in collective behaviour may help understand the selective pressures that shape how animal collectives form and function.

Ecological character displacement between the sexes, and sexual selection, integrate into a convergent set of factors that produce sexual variation. Ecologically modulated, sexually mediated variation within and between sexes may be a major contributor to the amount of total variation that selection can act on in species. Threespine stickleback (Gasterosteus aculeatus) display rapid adaptive responses and sexual variation in many phenotypic traits. We examined phenotypic variation in the skull, pectoral and pelvic girdles of threespine stickleback from two freshwater and two coastal marine sites on the Sunshine Coast of British Columbia, Canada, using an approach that avoids a priori assumptions about bimodal patterns of variation. We quantified shape and size of the cranial, pectoral and pelvic regions of sticklebacks in marine and freshwater habitats using 3D geometric morphometrics and an index of sexually mediated variation. We show that the expression of phenotypic variation is structured in part by the effects of both habitat marine vs freshwater and the effects of individual sites within each habitat. Relative size exerts variable influence, and patterns of phenotypic variation associated with sex vary among body regions. This fine‐grained quantification of sexually mediated variation in the context of habitat difference and different anatomical structures indicates a complex relationship between genetically inferred sex and environmental factors, demonstrating that the interplay between shared genetic background and sexually mediated, ecologically based selective pressures structures the phenotypic expression of complex traits. [ABSTRACT FROM AUTHOR]

Abstract Intraspecific niche differentiation can contribute to population persistence in changing environments. Following declines in large predatory fish, eutrophication, and climate change, there has been a major increase in the abundance of threespine stickleback (Gasterosteus aculeatus) in the Baltic Sea. Two morphotype groups with different levels of body armor—completely plated and incompletely plated—are common in coastal Baltic Sea habitats. The morphotypes are similar in shape, size, and other morphological characteristics and live as one apparently intermixed population. Variation in resource use between the groups could indicate a degree of niche segregation that could aid population persistence in the face of further environmental change. To assess whether morphotypes exhibit niche segregation associated with resource and/or habitat exploitation and predator avoidance, we conducted a field survey of stickleback morphotypes, and biotic and abiotic ecosystem structure, in two habitat types within shallow coastal bays in the Baltic Sea: deeper central waters and shallow near‐shore waters. In the deeper waters, the proportion of completely plated stickleback was greater in habitats with greater biomass of two piscivorous fish: perch (Perca fluviatilis) and pike (Esox lucius). In the shallow waters, the proportion of completely plated stickleback was greater in habitats with greater coverage of habitat‐forming vegetation. Our results suggest niche segregation between morphotypes, which may contribute to the continued success of stickleback in coastal Baltic Sea habitats.

Abstract Ecological character displacement between the sexes, and sexual selection, integrate into a convergent set of factors that produce sexual variation. Ecologically modulated, sexually mediated variation within and between sexes may be a major contributor to the amount of total variation that selection can act on in species. Threespine stickleback (Gasterosteus aculeatus) display rapid adaptive responses and sexual variation in many phenotypic traits. We examined phenotypic variation in the skull, pectoral and pelvic girdles of threespine stickleback from two freshwater and two coastal marine sites on the Sunshine Coast of British Columbia, Canada, using an approach that avoids a priori assumptions about bimodal patterns of variation. We quantified shape and size of the cranial, pectoral and pelvic regions of sticklebacks in marine and freshwater habitats using 3D geometric morphometrics and an index of sexually mediated variation. We show that the expression of phenotypic variation is structured in part by the effects of both habitat marine vs freshwater and the effects of individual sites within each habitat. Relative size exerts variable influence, and patterns of phenotypic variation associated with sex vary among body regions. This fine‐grained quantification of sexually mediated variation in the context of habitat difference and different anatomical structures indicates a complex relationship between genetically inferred sex and environmental factors, demonstrating that the interplay between shared genetic background and sexually mediated, ecologically based selective pressures structures the phenotypic expression of complex traits.

Interspecific hybridization has occurred relatively frequently during the evolution of vertebrates. This process usually abolishes reproductive isolation between the parental species. Moreover, it results in the exchange of genetic material and can lead to hybridogenic speciation. Hybridization between species has predominately been observed at the interspecific level, whereas intergeneric hybridization is rarer. Here, using whole-genome sequencing analysis, we describe clear and reliable signals of intergeneric introgression between the three-spined stickleback (Gasterosteus aculeatus) and its distant mostly freshwater relative the nine-spined stickleback (Pungitius pungitius) that inhabit northwestern Russia. Through comparative analysis, we demonstrate that such introgression phenomena apparently take place in the moderate-salinity White Sea basin, although it is not detected in Japanese sea stickleback populations. Bioinformatical analysis of the sites influenced by introgression showed that they are located near transposable elements, whereas those in protein-coding sequences are mostly found in membraneassociated and alternative splicing-related genes. [ABSTRACT FROM AUTHOR]

Research on among individual variation in behavior has increased rapidly in recent years. It is intuitively appealing that variation in the behavior of individuals has ecological consequences, for example, on trophic niche. Bold individuals, with a tendency to be explorative and risk‐tolerant, can be less likely to alter their foraging behavior based on context and, therefore, forage more consistently. Stable isotopes are a useful tool to retrospectively estimate ecological niche and have been found to correlate with foraging behavior in the wild and can be used to extend studies to further examine the ecological or evolutionary relevance of animal personality measured in the laboratory. We examined if common behavioral traits related to ecological niche in the wild using a rapid behavioral assay and δ13C and δ15N stable isotopes from fin and muscle, reflecting ecological niche for the previous weeks and months. We found that bolder stickleback (and females) were enriched in δ13C compared with shyer stickleback (and males) but the covariation of gender and stable isotope values complicate inferences. Nevertheless, the current results highlight the long‐term ecological importance of among individual variation in behavior and are among the first to support a correlation between laboratory measures of behavior and ecological niche in the wild and to document gender differences in stable isotope values of fish. [ABSTRACT FROM AUTHOR]

The vertebrate sodium–iodide symporter (NIS or SLC5A5) transports iodide into the thyroid follicular cells that synthesize thyroid hormone. The SLC5A protein family includes transporters of vitamins, minerals, and nutrients. Disruption of SLC5A5 function by perchlorate, a pervasive environmental contaminant, leads to human pathologies, especially hypothyroidism. Perchlorate also disrupts the sexual development of model animals, including threespine stickleback (Gasterosteus aculeatus) and zebrafish (Danio rerio), but the mechanism of action is unknown. To test the hypothesis that SLC5A5 paralogs are expressed in tissues necessary for the development of reproductive organs, and therefore are plausible candidates to mediate the effects of perchlorate on sexual development, we first investigated the evolutionary history of Slc5a paralogs to better understand potential functional trajectories of the gene family. We identified two clades of slc5a paralogs with respect to an outgroup of sodium/choline cotransporters (slc5a7); these clades are the NIS clade of sodium/iodide and lactate cotransporters (slc5a5, slc5a6, slc5a8, slc5a8, and slc5a12) and the SGLT clade of sodium/glucose cotransporters (slc5a1, slc5a2, slc5a3, slc5a4, slc5a10, and slc5a11). We also characterized expression patterns of slc5a genes during development. Stickleback embryos and early larvae expressed NIS clade genes in connective tissue, cartilage, teeth, and thyroid. Stickleback males and females expressed slc5a5 and its paralogs in gonads. Single‐cell transcriptomics (scRNA‐seq) on zebrafish sex‐genotyped gonads revealed that NIS clade‐expressing cells included germ cells (slc5a5, slc5a6a, and slc5a6b) and gonadal soma cells (slc5a8l). These results are consistent with the hypothesis that perchlorate exerts its effects on sexual development by interacting with slc5a5 or its paralogs in reproductive tissues. These findings show novel expression domains of slc5 genes in stickleback and zebrafish, which suggest similar functions across vertebrates including humans, and provide candidates to mediate the effects of perchlorate on sexual development. [ABSTRACT FROM AUTHOR]

Here, we present and characterize the spontaneous X-linked recessive mutation casper, which causes oculocutaneous albinism in threespine sticklebacks (Gasterosteus aculeatus). In humans, Hermansky-Pudlak syndrome results in pigmentation defects due to disrupted formation of the melanin-containing lysosomal-related organelle (LRO), the melanosome. casper mutants display not only reduced pigmentation of melanosomes in melanophores, but also reductions in the iridescent silver color from iridophores, while the yellow pigmentation from xanthophores appears unaffected. We mapped casper using high-throughput sequencing of genomic DNA from bulked casper mutants to a region of the stickleback X chromosome (chromosome 19) near the stickleback ortholog of Hermansky-Pudlak syndrome 5 (Hps5). casper mutants have an insertion of a single nucleotide in the sixth exon of Hps5, predicted to generate an early frameshift. Genome editing using CRISPR/Cas9 induced lesions in Hps5 and phenocopied the casper mutation. Injecting single or paired Hps5 guide RNAs revealed higher incidences of genomic deletions from paired guide RNAs compared to single gRNAs. Stickleback Hps5 provides a genetic system where a hemizygous locus in XY males and a diploid locus in XX females can be used to generate an easily scored visible phenotype, facilitating quantitative studies of different genome editing approaches. Lastly, we show the ability to better visualize patterns of fluorescent transgenic reporters in Hps5 mutant fish. Thus, Hps5 mutations present an opportunity to study pigmented LROs in the emerging stickleback model system, as well as a tool to aid in assaying genome editing and visualizing enhancer activity in transgenic fish.

The light environment influences an animal's ability to forage, evade predators, and find mates, and consequently is known to drive local adaptation of visual systems. However, the light environment may also vary over fine spatial scales at which genetic adaptation is difficult. For instance, in aquatic systems, the available wavelengths of light change over a few metres depth. Do animals plastically adjust their visual system to such small-scale environmental light variation? Here, we show that in three-spine stickleback (Gasterosteus aculeatus), opsin gene expression (an important determinant of colour vision) changes over a 2-m vertical gradient in nest depth. By experimentally altering the light environment using light filters to cover enclosures in a lake, we found that opsin expression can be adjusted on a short time frame (weeks) in response to the local light environment. This is to our knowledge the smallest spatial scale on which visual adjustments through opsin expression have been recorded in a natural setting along a continuously changing light environment.

Interspecific hybridization has occurred relatively frequently during the evolution of vertebrates. This process usually abolishes reproductive isolation between the parental species. Moreover, it results in the exchange of genetic material and can lead to hybridogenic speciation. Hybridization between species has predominately been observed at the interspecific level, whereas intergeneric hybridization is rarer. Here, using whole-genome sequencing analysis, we describe clear and reliable signals of intergeneric introgression between the three-spined stickleback (Gasterosteus aculeatus) and its distant mostly freshwater relative the nine-spined stickleback (Pungitius pungitius) that inhabit northwestern Russia. Through comparative analysis, we demonstrate that such introgression phenomena apparently take place in the moderate-salinity White Sea basin, although it is not detected in Japanese sea stickleback populations. Bioinformatical analysis of the sites influenced by introgression showed that they are located near transposable elements, whereas those in protein-coding sequences are mostly found in membrane-associated and alternative splicing-related genes.

Abstract Research on among individual variation in behavior has increased rapidly in recent years. It is intuitively appealing that variation in the behavior of individuals has ecological consequences, for example, on trophic niche. Bold individuals, with a tendency to be explorative and risk‐tolerant, can be less likely to alter their foraging behavior based on context and, therefore, forage more consistently. Stable isotopes are a useful tool to retrospectively estimate ecological niche and have been found to correlate with foraging behavior in the wild and can be used to extend studies to further examine the ecological or evolutionary relevance of animal personality measured in the laboratory. We examined if common behavioral traits related to ecological niche in the wild using a rapid behavioral assay and δ13C and δ15N stable isotopes from fin and muscle, reflecting ecological niche for the previous weeks and months. We found that bolder stickleback (and females) were enriched in δ13C compared with shyer stickleback (and males) but the covariation of gender and stable isotope values complicate inferences. Nevertheless, the current results highlight the long‐term ecological importance of among individual variation in behavior and are among the first to support a correlation between laboratory measures of behavior and ecological niche in the wild and to document gender differences in stable isotope values of fish.

Loss of body armour, sometimes including a reduction in or loss of pelvic spines, is an adaptation observed in many isolated freshwater populations. Pelvic reduction in sticklebacks has previously been associated with recurrent, but variant, deletions within pelvic enhancer regions PelA and PelB, which regulate expression of the homeodomain transcription factor gene Pitx1. We investigated variation in nucleotide sequences of pelvic enhancers in sticklebacks collected from two small freshwater lakes in the same watercourse and a nearby marine site in subarctic Norway. Spineless, as well as asymmetrically spined and completely spined sticklebacks are present in the upper lake, while only specimens with complete spines are found at the other lake and the marine site. Observed variation at PelA between the three sites was mainly due to variable numbers of repeats at three fragile TG-repeat loci. The length of PelA, mainly at one of the TG-repeat loci, was consistently shorter among individuals in the upper lake compared with specimens from the two other sites. However, no obvious association was revealed between enhancer variants and pelvic status. No polymorphism was found at PelB. Thus, additional genetic factors and/or environmental cues need to be identified to fully explain the occurrence of pelvic reduction in sticklebacks in this lake.

Understanding of genetic mechanisms underlying variation in sexual dichromatism remains limited, especially for carotenoid‐based colors. We addressed this knowledge gap in a gene expression study with threespine stickleback. We compared male and female throat tissues across five populations, including two in which female red coloration has evolved convergently. We found that the expression of individual genes, gene ontologies, and coexpression networks associated with red female color within a population differed between California and British Columbia populations, suggesting differences in underlying mechanisms. Comparing females from each of these populations to females from populations dominated by dull females, we again found extensive expression differences. For each population, genes and networks associated with female red color showed the same patterns for males only inconsistently. The functional roles of genes showing correlated expression with female color are unclear within populations, whereas genes highlighted through inter‐population comparisons include some previously suggested to function in carotenoid pathways. Among these, the most consistent patterns involved TTC39B (Tetratricopeptide Repeat Domain 39B), which is within a known red coloration QTL in stickleback and implicated in red coloration in other taxa. [ABSTRACT FROM AUTHOR]

Human-induced changes in climate and habitats push populations to adapt to novel environments, including new sensory conditions, such as reduced visibility. We studied how colonizing newly formed glacial lakes with turbidity-induced low-visibility affects anti-predator behaviour in Icelandic threespine sticklebacks. We tested nearly 400 fish from 15 populations and four habitat types varying in visibility and colonization history in their reaction to two predator cues (mechano-visual versus olfactory) in high versus low-visibility light treatments. Fish reacted differently to the cues and were affected by lighting environment, confirming that cue modality and light levels are important for predator detection and evasion. Fish from spring-fed lakes, especially from the highlands (likely more diverged from marine fish than lowland fish), reacted fastest to mechano-visual cues and were generally most active. Highland glacial fish showed strong responses to olfactory cues and, counter to predictions from the flexible stem hypothesis, the greatest plasticity in response to light levels. This study, leveraging natural, repeated invasions of novel sensory habitats, (i) illustrates rapid changes in anti-predator behaviour that follow due to adaptation, early life experience, or both, and (ii) suggests an additional role for behavioural plasticity enabling population persistence in the face of frequent changes in environmental conditions. [ABSTRACT FROM AUTHOR]

Abstract Understanding of genetic mechanisms underlying variation in sexual dichromatism remains limited, especially for carotenoid‐based colors. We addressed this knowledge gap in a gene expression study with threespine stickleback. We compared male and female throat tissues across five populations, including two in which female red coloration has evolved convergently. We found that the expression of individual genes, gene ontologies, and coexpression networks associated with red female color within a population differed between California and British Columbia populations, suggesting differences in underlying mechanisms. Comparing females from each of these populations to females from populations dominated by dull females, we again found extensive expression differences. For each population, genes and networks associated with female red color showed the same patterns for males only inconsistently. The functional roles of genes showing correlated expression with female color are unclear within populations, whereas genes highlighted through inter‐population comparisons include some previously suggested to function in carotenoid pathways. Among these, the most consistent patterns involved TTC39B (Tetratricopeptide Repeat Domain 39B), which is within a known red coloration QTL in stickleback and implicated in red coloration in other taxa.