Coarse- and fine-grained phenotypic divergence among threespine stickleback from alternating lake and stream habitats

Variation in the gillrakers and diet of threespine sticklebacks (Gasterosteus aculeatus) was evident throughout their European distribution. The gillrakers of completely-plated sticklebacks were more numerous, longer, and more closely spaced than those of low-plated sticklebacks. Among populations of either form, sticklebacks from lentic (marine, lake and pond) habitats had more gillrakers than did those from lotic (river and stream) habitats. Genetic control of gillraker number was demonstrated for one lake-dwelling and one stream-dwelling population. Lake-dwelling sticklebacks were found to consume more zooplankton than did stream-dwelling individuals although benthic invertebrates were important prey in both habitats. Variation in gillrakers could be attributed only in part to selection pressures arising from the greater abundance of zooplankton prey in lake than stream habitats. Evidence of stabilizing selection led us to conclude that conflicting selection pressures determined the habitat-characteristic patterns of geographic variation. THREESPINE sticklebacks (Gasterosteus aculeatus) are small teleosts well known for their morphological variability. They are widely distributed in the middle and high latitudes of the Northern Hemisphere where environmental instability is thought to favor the capacity for adaptive variation (Nikolsky and Vasilyev, 1973). Recent studies along the Pacific Coast of North America and in Europe have shown that variation of some morphological characters has a genetic basis, and that local selection pressures may often explain both habitat-specific and regional trends of variation (e.g., Bell and Haglund, 1978; Gross, 1977, 1978; McPhail, 1977;

Hydrodynamic forces and their absence appear to exert differential selection pressure on aquatic biodiversity in lake and stream habitats, creating a tight fit between organismal phenotypes and their environments. Ecophenotypic variants may be the result of genetic differentiation or phenotypic plasticity, where a genotype can produce multiple phenotypes dependent on the environment. Freshwater mussels possess a wide degree of morphological variation that frequently covaries with the environment, making them a good system to understand the mechanisms of ecophenotypic variation across hydrological conditions. We designed a two‐year experiment where individuals from the same Pyganodon grandis maternal brood (half and full siblings) were reared at a controlled site and four natural sites involving one lake and three streams. At the end of the experiment, shell shape was quantified for recaptured (N = 70), wild (N = 206), and zoo‐reared (N = 305) mussels. The maternal individual and 46 recaptured mussels were sequenced for genomic single nucleotide polymorphisms to test for multiple paternity and its effect on offspring morphology. Analysis of covariance found significant differences in shell shape between rearing sites, particularly between stream and lake habitats, but no shape differences were detected across the three stream sites. At two of the four sites, the shell shape of recaptured individuals was not significantly different than that of wild populations. Genomic sequencing and parentage analysis identified 11–27 different fathers among recaptured individuals. Yet no genetic differences were present between stream and lake habitats, and there was no paternal effect on shell shape. Taken together, phenotypic plasticity, over genetic differentiation, is identified as the primary mechanism of ecophenotypic variation. Plasticity is likely ubiquitous across freshwater mussels and may be a key adaptation given their high variance in habitat use. Multiple paternity may also play a role in the evolution of phenotypic plasticity, allowing more males from greater distances opportunities for fertilization, thus increasing genetic connectivity. Lastly, phenotypic plasticity and multiple paternity are convenient properties for freshwater mussel conservation and propagation. Multiple fathers increase the genetic variation of propagated broods, while plasticity may provide resilience to the release of stocked individuals across environmental heterogeneity.

Phenotypic variability represents an important factor allowing species to adapt to local environmental conditions, but mechanisms underlying such variation are incompletely understood. This study investigated whether habitat-specific demands on swimming performance or difference in trophic relationships in lakes (pelagic, littoral) and streams (riffle, pool) were significant predictors of phenotypic variation exhibited by brook char (Salvelinus fontinalis), the only fish in the study habitats. Specifically, we hypothesized that pelagic and riffle habitats would impose greater selective pressures associated with swimming, resulting in body morphologies that were dorsoventrally compressed, anterior–posteriorly elongated, and that exhibited a long, narrow caudal peduncle. Geometric morphometrics was applied in a quantitative analysis of body morphology among habitats, whereas stable isotope analysis was used to differentiate between food sources. Analyses revealed that while body morphology differed between lake and stream habitats, there was convergence between the pelagic and riffle habitats, as well as among littoral and riffle and pool environments. The littoral and pool habitats were thought to be more structurally complex, thereby selecting for increased maneuverability but lower sustained swimming and correspondingly deeper bodies with shorter, dorsoventrally expanded caudal peduncles. Carbon source and trophic position did not differ among habitats with a system, suggesting that feeding was not the main influence on morphological plasticity; however, fish in the stream were feeding at a higher trophic position than fish in the lake. These findings suggest that individual species may take advantage of morphological variation to better adapt local surroundings.

The lateral line is a mechanoreceptive sensory system that allows fish to sense objects and motion in their local environment. Variation in lateral line morphology may allow fish in different habitats to differentially sense and respond to salient cues. Threespine sticklebacks (Gasterosteus aculeatus) occupy a diverse range of aquatic habitats; we therefore hypothesized that populations within the G. aculeatus species complex might show variation in the morphology of the lateral line sensory system. We sampled 16 threespine stickleback populations from marine, stream and lake (including benthic and limnetic) habitats and examined the distribution, type and number of neuromasts on different regions of the body. We found that the threespine stickleback has a reduced lateral line canal system, completely lacking canal neuromasts. Although the arrangement of lines of superficial neuromasts on the body was largely the same in all populations, the number of neuromasts within these lines varied across individuals, populations and habitats. In pairwise comparisons between threespine sticklebacks adapted to divergent habitats, we found significant differences in neuromast number. Stream residents had more neuromasts than marine sticklebacks living downstream in the same watershed. In two independent lakes, benthic sticklebacks had more trunk neuromasts than sympatric limnetic sticklebacks, providing evidence for parallel evolution of the lateral line system. Our data provide the first demonstration that the lateral line sensory system can vary significantly between individuals and among populations within a single species, and suggest that this sensory system may experience different selection regimes in alternative habitats.

The repeated evolution of similar phenotypes in independent populations (i.e. parallel or convergent evolution) provides an opportunity to identify genetic and ecological factors that influence the process of adaptation. Threespine stickleback fish (Gasterosteus aculeatus) are an excellent model for such studies, as they have repeatedly adapted to divergent habitats across the Northern hemisphere. Here, we use genomic, ecological and morphological data from 16 independent pairs of stickleback populations adapted to divergent lake and stream habitats. We combine a population genomic approach to identify regions of the genome that are likely under selection in these divergent habitats with an association mapping approach to identify regions of the genome that underlie variation in ecological factors and morphological traits. Over 37% of genomic windows are repeatedly differentiated across lake–stream pairs. Similarly, many genomic windows are associated with variation in abiotic factors, diet items and morphological phenotypes. Both the highly differentiated windows and candidate trait windows are non-randomly distributed across the genome and show some overlap. However, the overlap is not significant on a genome-wide scale. Together, our data suggest that adaptation to divergent food resources and predation regimes are drivers of differentiation in lake–stream stickleback, but that additional ecological factors are also important.This article is part of the theme issue ‘Convergent evolution in the genomics era: new insights and directions’.

Parallel evolution of similar traits by independent populations in similar environments is considered strong evidence for adaptation by natural selection. Often, however, replicate populations in similar environments do not all evolve in the same way, thus deviating from any single, predominant outcome of evolution. This variation might arise from non-adaptive, population-specific effects of genetic drift, gene flow or limited genetic variation. Alternatively, these deviations from parallel evolution might also reflect predictable adaptation to cryptic environmental heterogeneity within discrete habitat categories. Here, we show that deviations from parallel evolution are the consequence of environmental variation within habitats combined with variation in gene flow. Threespine stickleback (Gasterosteus aculeatus) in adjoining lake and stream habitats (a lake-stream 'pair') diverge phenotypically, yet the direction and magnitude of this divergence is not always fully parallel among 16 replicate pairs. We found that the multivariate direction of lake-stream morphological divergence was less parallel between pairs whose environmental differences were less parallel. Thus, environmental heterogeneity among lake-stream pairs contributes to deviations from parallel evolution. Additionally, likely genomic targets of selection were more parallel between environmentally more similar pairs. In contrast, variation in the magnitude of lake-stream divergence (independent of direction) was better explained by differences in lake-stream gene flow; pairs with greater lake-stream gene flow were less morphologically diverged. Thus, both adaptive and non-adaptive processes work concurrently to generate a continuum of parallel evolution across lake-stream stickleback population pairs.

The ability for an organism to extract energy from its environment is central to its survival, reproduction and fitness. In addition to traits within the host that directly influence digestion, such as intestinal physiology and morphology, an animal's gut bacterial constituents can also strongly influence its digestive capabilities. In fact, bacterial communities that live in and on an organism can strongly influence the host's condition and fitness. Although fish are the most diverse vertebrate group, little is known about their microbial constituents or how they impact their physiology, ecology and evolution, and likewise, how fish affect the bacteria in their environment. To explore factors that shape fish bacterial communities, I investigated the variation in gut bacteria across fish species and populations. Through a meta-analysis of gut bacteria from diverse fish hosts, patterns of variation in response to alterations in diet, trophic level, and habitat were apparent from the 18 fish species included in the analysis. After focusing on a broad array of fish taxa, I concentrated on the Trinidadian guppy, Poecilia reticulata, which serves as a model system for evolutionary biology. I investigated the composition of guppy gut microbial communities as well as other elements of their digestive physiology that may affect their ability to extract energy from their environment. Through field surveys and a dietary manipulation, I found that populations of guppies differ in digestive traits, including digestive enzymatic profiles and gut morphology. The bacterial communities in guppies appeared to be largely divided by stream of origin, but because bacteria found in guppy guts were distinct from the bacteria in their environment, they appear to be driven, in part, by guppy population background. Finally, I expanded my investigation to examine how fish influence the community structure of bacteria in their environment. Using the threespine stickleback (Gasterosteus aculeatus) from lake and stream habitats in Switzerland, I explored how their presence and population background affect the structure of the bacterial communities in their experimental mesocosms. I also explored whether the effect of population background on environmental bacterial community structure is subject to plasticity due to the diet upon which they were reared. I found that the presence of fish strongly shapes the microbes that are present in their environment as well as their abundance. Additionally, the interaction of rearing diet and fish population background can also influence bacteria present in the mesocosms. The results from this dissertation show how fish ecology and environmental factors interact to shape the communities of gut bacteria in fish, and how fish can also influence the bacteria in their environment. This work is a key step to establish how bacterial communities are structured in fish and in their aquatic environments. It also lays the groundwork for future studies on the functional contributions of bacteria to fish ecology, which should lead to a better understanding of how fish interact with their environments.

Threespine sticklebacks (Gasterosteus aculeatus) from different habitats have been observed to differ in shoaling behavior, both in the wild and in laboratory studies. In the present study, we surveyed the shoaling behavior of sticklebacks from a variety of marine, lake, and stream habitats throughout the Pacific Northwest. We tested the shoaling tendencies of 113 wild-caught sticklebacks from 13 populations using a laboratory assay that was based on other published shoaling assays in sticklebacks. Using traditional behavioral measures for this assay, such as time spent shoaling and mean position in the tank, we were unable to find population differences in shoaling behavior. However, simple plotting techniques revealed differences in spatial distributions during the assay. When we collapsed individual trials into population-level data sets and applied information theoretic measurements, we found significant behavioral differences between populations. For example, entropy estimates confirm that populations display differences in the extent of clustering at various tank positions. Using log-likelihood analysis, we show that these population-level observations reflect consistent differences in individual behavioral patterns that can be difficult to discriminate using standard measures. The analytical techniques we describe may help improve the detection of potential behavioral differences between fish groups in future studies.

Parallel (or convergent) evolution provides strong evidence for a deterministic role of natural selection: similar phenotypes evolve when independent populations colonize similar environments. In reality, however, independent populations in similar environments always show some differences: some nonparallel evolution is present. It is therefore important to explicitly quantify the parallel and nonparallel aspects of trait variation, and to investigate the ecological and genetic explanations for each. We performed such an analysis for threespine stickleback (Gasterosteus aculeatus) populations inhabiting lake and stream habitats in six independent watersheds. Morphological traits differed in the degree to which lake-stream divergence was parallel across watersheds. Some aspects of this variation were correlated with ecological variables related to diet, presumably reflecting the strength and specifics of divergent selection. Furthermore, a genetic scan revealed some markers that diverged between lakes and streams in many of the watersheds and some that diverged in only a few watersheds. Moreover, some of the lake-stream divergence in genetic markers was associated within some of the lake-stream divergence in morphological traits. Our results suggest that parallel evolution, and deviations from it, are primarily the result of natural selection, which corresponds in only some respects to the dichotomous habitat classifications frequently used in such studies.

Speciation can be initiated by adaptive divergence between populations in ecologically different habitats, but how sexually based reproductive barriers contribute to this process is less well understood. We here test for sexual isolation between ecotypes of threespine stickleback fish residing in adjacent lake and stream habitats in the Lake Constance basin, Central Europe. Mating trials in outdoor mesocosms allowing for natural reproductive behavior reveal that mating occurs preferentially between partners of the same than of the opposed ecotype. Compared to random mating, this sexual barrier reduces gene flow between the ecotypes by some 36%. This relatively modest strength of sexual isolation is surprising because comparing the males between the two ecotypes shows striking differentiation in traits generally considered relevant to reproductive behavior (body size, breeding coloration, nest size). Analyzing size differences among the individuals in the mating trials further indicates that assortative mating is not related to ecotype differences in body size. Overall, we demonstrate that sexually based reproductive isolation promotes divergence in lake-stream stickleback along with other known reproductive barriers, but also caution against inferring strong sexual isolation from the observation of strong population divergence in sexually relevant traits.,Dryad.stickleback.mate.choiceComplete data set used for all analyses in the study. The rows are the replicate experimental trials (N = 58). The first column specifies the date of termination of the trials. The subsequent columns describe the experimental females, then the lake males, then the stream males (each trial involved a single female and a male of each ecotype). The last column indicates the male that mated with the female. Missing data are coded as 'NA'.,

Major histocompatibility complex (MHC) genes encode proteins that play a central role in vertebrates' adaptive immunity to parasites. MHC loci are among the most polymorphic in vertebrates' genomes, inspiring many studies to identify evolutionary processes driving MHC polymorphism within populations and divergence between populations. Leading hypotheses include balancing selection favouring rare alleles within populations, and spatially divergent selection. These hypotheses do not always produce diagnosably distinct predictions, causing many studies of MHC to yield inconsistent or ambiguous results. We suggest a novel strategy to distinguish balancing vs. divergent selection on MHC, taking advantage of natural admixture between parapatric populations. With divergent selection, individuals with immigrant alleles will be more infected and less fit because they are susceptible to novel parasites in their new habitat. With balancing selection, individuals with locally rare immigrant alleles will be more fit (less infected). We tested these contrasting predictions using three-spine stickleback from three replicate pairs of parapatric lake and stream habitats. We found numerous positive and negative associations between particular MHC IIβ alleles and particular parasite taxa. A few allele-parasite comparisons supported balancing selection, and others supported divergent selection between habitats. But, there was no overall tendency for fish with immigrant MHC alleles to be more or less heavily infected. Instead, locally rare MHC alleles (not necessarily immigrants) were associated with heavier infections. Our results illustrate the complex relationship between MHC IIβ allelic variation and spatially varying multispecies parasite communities: different hypotheses may be concurrently true for different allele-parasite combinations.

About 15,000years earlier, the Northern half of Europe and North America was buried under a few kilometres of ice. Since then, many organisms have colonized and rapidly adapted to the new, vacant habitats. Some, like the threespine stickleback fish, have done so more successfully than others: from the sea, stickleback have adapted to a multitude of lake and stream habitats with a vast array of complex phenotypes and life histories. Previous studies showed that most of these "ecotypes" differ in multiple divergently selected genes throughout the genome. But how are well-adapted ecotypes of one habitat protected from maladaptive gene flow from ecotypes of another, adjacent habitat? According to a From the Cover meta-analysis in this issue of Molecular Ecology (Samuk etal., 2017), low recombination rate regions in the genome offer such protection. While inversions have often been highlighted as an efficient way to maintain linkage disequilibrium among sets of adaptive variants in the face of gene flow, Samuk etal. (2017) show that variation in recombination rate across the genome may perform a similar role in threespine stickleback. With this study, theoretical predictions for the importance of low recombination regions in adaptation are for the first time tested with a highly replicated population genomic data set. The findings from this study have implications for the adaptability of species, speciation and the evolution of genome architecture.

River levels in Central Amazonia fluctuate up to 14 m annually, with the flooding period ranging from 50 to 270 days between the rising and falling phases. Vast areas of forest along the rivers contain plant species that are well adapted to annual flooding. We studied the effect of flooding level on tree species richness, diversity, density, and composition in lake, river, and stream habitats in Jaú National Park, Brazil. 3051 trees >10 cm diameter (at 1.3 m diameter at breast height, dbh) were measured and identified in 25 10 m × 40 m randomly selected plots in each habitat. Ordination methods and analysis of variance results showed that forested areas near lakes had significantly lower species richness of trees than riverine and streamside habitats. Plot species richness and diversity were strongly negatively correlated with the water level and duration of flooding. The drier (stream) habitat had more total species (54 species of trees) and more unique species of trees (6 tree species) than the riverine (52 tree species; 3 unique species) and lake (33 tree species; 3 unique species) habitats. Species composition overlap among habitats was surprisingly high (42.6-60.6% overlap), almost one-third of the species were found in all three habitat types, and few species were unique to each habitat. We conclude that: (1) duration of flooding has a strong impact on species richness, diversity and plant distribution patterns; (2) most species are adapted to a wide range of habitats and flood durations; and (3) while flood duration may decrease local diversity, it also creates and maintains high landscape-scale diversity by increasing landscape heterogeneity.

Fitness costs of signalling are essential in order for reliable sexual signalling to prevail when the interests of the sexes conflict. This means that signalling can be subjected to a life history trade-off between present and future signalling effort. Here, I show that three-spined stickleback males (Gasterosteus aculeatus), who have a single breeding season during which they breed repeatedly, change their red nuptial coloration over the season depending on their body size at the start of breeding. Large males that completed several breeding cycles increased their red coloration over the season, whereas small males, who completed only a few cycles, did not. The increase in coloration was accompanied by an increase in parental success when males were energy constrained, but not when they had access to an unlimited food supply. Red coloration was thus an honest signal of male parental ability despite changes in signal expression when both signalling and parental care were costly and the investments in them changed simultaneously over the reproductive lifetime. However, the honesty of the signal varied over a lifetime. At the penultimate cycle, bright males cannibalized some of their eggs, probably to increase survival to the last cycle, whereas males cared for their offspring independent of coloration at the ultimate cycle.

1. Despite the great interest in characterising fish reproductive habitat, the relationship between the selection of a given spawning site and individual fitness has not been experimentally tested. 2. In this study, we used an in situ experimental approach to determine (i) the relative contribution of substrate characteristics as well as hydrological and physicochemical variables to small-scale redd site selection by brook charr, Salvelinus fontinalis (Mitchill) and (ii) if hatching and emergence success, used as a surrogate of fitness, are improved in selected compared to non-selected sites in both lake and stream habitats. 3. Our results show that upwelling groundwater flow was always significantly higher in selected than in non-selected sites in both lake and stream habitats. We found no significant difference in the mean geometric substrate diameter and no consistent differences in substrate composition between selected and non-selected sites. Oxygen concentration was higher (significantly so in three of four comparisons) and conductivity tended to be lower in selected than in non-selected sites, while temperature showed no significant or consistent variations. We found a significant positive relationship between the selection of a given spawning habitat and hatching and emergence success in these systems. 4. These results show that the main cue that brook charr use to select their spawning sites is upwelling groundwater in lake and stream habitats, and that active selection of these sites increases individual fitness. This suggests that natural selection acted on the same cues in lentic and lotic environments; this could have been highly adaptive in a species that used both habitats as colonisation routes after the last glaciation.