All day-long: Sticklebacks effectively forage on whitefish eggs during all light conditions

The three-spined stickleback Gasterosteus aculeatus invaded Lake Contance in the 1940s and expanded in large numbers from an exclusively shoreline habitat into the pelagic zone in 2012. Stickleback abundance is very high in the pelagic zone in winter near the spawning time of pelagic whitefish Coregonus wartmanni, and it is hypothesized that this is triggered by the opportunity to consume whitefish eggs. Field sampling has qualitatively confirmed predation of whitefish eggs by stickleback, but quantification has proven difficult due to stormy conditions that limit sampling. One fundamental unknown is if freshwater stickleback, known as visual feeders, can successfully find and eat whitefish eggs during twilight and night when whitefish spawn. It is also unknown how long eggs can be identified in stomachs following ingestion, which could limit efforts to quantify egg predation through stomach content analysis. To answer these questions, 144 individuals were given the opportunity to feed on whitefish roe under daylight, twilight, and darkness in controlled conditions. The results showed that stickleback can ingest as many as 100 whitefish eggs under any light conditions, and some individuals even consumed maximum numbers in complete darkness. Furthermore, eggs could be unambiguously identified in the stomach 24 hours after consumption. Whitefish eggs have 28% more energy content than the main diet of sticklebacks (zooplankton) based on bomb-calorimetric measurements, underlining the potential benefits of consuming eggs. Based on experimental results and estimates of stickleback abundance and total egg production, stickleback could potentially consume substantial proportions of the total eggs produced even if relatively few sticklebacks consume eggs. Given the evidence that stickleback can feed on eggs during nighttime spawning and may thereby hamper recruitment, future studies aimed at quantifying actual egg predation and resulting effects on the whitefish population are urgently needed.

Selection, linkage, and population structure interact to shape genetic variation among threespine stickleback genomes

The outcome of selection on genetic variation depends on the geographic organization of individuals and populations as well as the syntenic organization of loci within the genome. Spatially variable selection between marine and freshwater habitats has had a significant and heterogeneous impact on patterns of genetic variation across the genome of threespine stickleback fish. When marine stickleback invade freshwater habitats, more than a quarter of the genome can respond to divergent selection, even in as little as 50 years. This process largely uses standing genetic variation that can be found ubiquitously at low frequency in marine populations, can be millions of years old, and is likely maintained by significant bidirectional gene flow. Here, we combine population genomic data of marine and freshwater stickleback from Cook Inlet, Alaska, with genetic maps of stickleback fish derived from those same populations to examine how linkage to loci under selection affects genetic variation across the stickleback genome. Divergent selection has had opposing effects on linked genetic variation on chromosomes from marine and freshwater stickleback populations: near loci under selection, marine chromosomes are depauperate of variation while these same regions among freshwater genomes are the most genetically diverse. Forward genetic simulations recapitulate this pattern when different selective environments also differ in population structure. Lastly, dense genetic maps demonstrate that the interaction between selection and population structure may impact large stretches of the stickleback genome. These findings advance our understanding of how the structuring of populations across geography influences the outcomes of selection, and how the recombination landscape broadens the genomic reach of selection.

Salinity-induced phenotypic plasticity in threespine stickleback sperm activation

Phenotypic expression may be and often is influenced by an organism's developmental environment, referred to as phenotypic plasticity. The sperm cells of teleosts have been found to be inactive in the seminal plasma and are activated by osmotic shock for most fish species, through release in either hypertonic (for marine fish) or hypotonic (for freshwater fish) water. If this is the case, the regulatory system of sperm mobility should be reversed in salt- and freshwater fish. We tested this hypothesis by first activating sperm of salt- and freshwater populations of threespine stickleback in salt- and freshwater. The sperm from saltwater stickleback could be activated in either salinity, which matches the freshwater colonization history of the species, whereas the sperm from the freshwater population acted as predicted by the osmotic shock theory and was activated in freshwater only. As the freshwater population used here was calculated to be thousands of years old, we went on to test whether the trait(s) were plastic and sperm from freshwater males still could be activated in saltwater after individuals were exposed to saltwater. After raising freshwater stickleback in saltwater, we found the mature males to have active sperm in both saltwater and freshwater. Further, we also found the sperm of wild-caught freshwater stickleback to be active in saltwater after exposing those mature males to saltwater for only 2 days. This illustrates that the ability for stickleback sperm to be activated in a range of water qualities is an environmentally induced plastic trait.

Ancient genomic variation underlies repeated ecological adaptation in young stickleback populations

ABSTRACTAdaptation in the wild often involves standing genetic variation (SGV), which allows rapid responses to selection on ecological timescales. However, we still know little about how the evolutionary histories and genomic distributions of SGV influence local adaptation in natural populations. Here, we address this knowledge gap using the threespine stickleback fish (Gasterosteus aculeatus) as a model. We extend the popular restriction site-associated DNA sequencing (RAD-seq) method to produce phased haplotypes approaching 700 base pairs (bp) in length at each of over 50,000 loci across the stickleback genome. Parallel adaptation in two geographically isolated freshwater pond populations consistently involved fixation of haplotypes that are identical-by-descent. In these same genomic regions, sequence divergence between marine and freshwater stickleback, as measured by dXY, reaches ten-fold higher than background levels and structures genomic variation into distinct marine and freshwater haplogroups. By combining this dataset with a de novo genome assembly of a related species, the ninespine stickleback (Pungitius pungitius), we find that this habitat-associated divergent variation averages six million years old, nearly twice the genome-wide average. The genomic variation that is involved in recent and rapid local adaptation in stickleback has actually been evolving throughout the 15-million-year history since the two species lineages split. This long history of genomic divergence has maintained large genomic regions of ancient ancestry that include multiple chromosomal inversions and extensive linked variation. These discoveries of ancient genetic variation spread broadly across the genome in stickleback demonstrate how selection on ecological timescales is a result of genome evolution over geological timescales, and vice versa.IMPACT STATEMENTAdaptation to changing environments requires a source of genetic variation. When environments change quickly, species often rely on variation that is already present – so-called standing genetic variation – because new adaptive mutations are just too rare. The threespine stickleback, a small fish species living throughout the Northern Hemisphere, is well-known for its ability to rapidly adapt to new environments. Populations living in coastal oceans are heavily armored with bony plates and spines that protect them from predators. These marine populations have repeatedly invaded and adapted to freshwater environments, losing much of their armor and changing in shape, size, color, and behavior.Adaptation to freshwater environments can occur in mere decades and probably involves lots of standing genetic variation. Indeed, one of the clearest examples we have of adaptation from standing genetic variation comes from a gene, eda, that controls the shifts in armor plating. This discovery involved two surprises that continue to shape our understanding of the genetics of adaptation. First, freshwater stickleback from across the Northern Hemisphere share the same version, or allele, of this gene. Second, the ‘marine’ and ‘freshwater’ alleles arose millions of years ago, even though the freshwater populations studied arose much more recently. While it has been hypothesized that other genes in the stickleback genome may share these patterns, large-scale surveys of genomic variation have been unable to test this prediction directly.Here, we use new sequencing technologies to survey DNA sequence variation across the stickleback genome for patterns like those at the eda gene. We find that every region of the genome associated with marine-freshwater genetic differences shares this pattern to some degree. Moreover, many of these regions are as old or older than eda, stretching back over 10 million years in the past and perhaps even predating the species we now call the threespine stickleback. We conclude that natural selection has maintained this variation over geological timescales and that the same alleles we observe in freshwater stickleback today are the same as those under selection in ancient, now-extinct freshwater habitats. Our findings highlight the need to understand evolution on macroevolutionary timescales to understand and predict adaptation happening in the present day.

The genomics of ecological vicariance in threespine stickleback fish

Populations occurring in similar habitats and displaying similar phenotypes are increasingly used to explore parallel evolution at the molecular level. This generally ignores the possibility that parallel evolution can be mimicked by the fragmentation of an ancestral population followed by genetic exchange with ecologically different populations. Here we demonstrate such an ecological vicariance scenario in multiple stream populations of threespine stickleback fish divergent from a single adjacent lake population. On the basis of demographic and population genomic analyses, we infer the initial spread of a stream-adapted ancestor followed by the emergence of a lake-adapted population, that selective sweeps have occurred mainly in the lake population, that adaptive lake–stream divergence is maintained in the face of gene flow from the lake into the streams, and that this divergence involves major inversion polymorphisms also important to marine-freshwater stickleback divergence. Overall, our study highlights the need for a robust understanding of the demographic and selective history in evolutionary investigations.

Cline coupling and uncoupling in a stickleback hybrid zone

Strong ecological selection on a genetic locus can maintain allele frequency differences between populations in different environments, even in the face of hybridization. When alleles at divergent loci come into tight linkage disequilibria, selection acts on them as a unit and can significantly reduce gene flow. For populations interbreeding across a hybrid zone, linkage disequilibria between loci can force clines to share the same slopes and centers. However, strong ecological selection can push clines away from the others, reducing linkage disequilibria and weakening the barrier to gene flow. We looked for this ‘cline uncoupling’ effect in a hybrid zone between stream resident and anadromous sticklebacks at two genes known to be under divergent natural selection (Eda and ATP1a1) and five morphological traits that repeatedly evolve in freshwater stickleback. We used 10 anonymous SNPs to characterize the shape of the zone. We found that the clines at Eda, ATP1a1, and four morphological traits were concordant and coincident, suggesting that direct selection on each is outweighed by the indirect selection generated by linkage disequilibria. Interestingly, the cline for pectoral fin length was much steeper and displaced 200m downstream, and two anonymous SNPs also had steep clines.

Two developmentally temporal quantitative trait loci underlie convergent evolution of increased branchial bone length in sticklebacks

In convergent evolution, similar phenotypes evolve repeatedly in independent populations, often reflecting adaptation to similar environments. Understanding whether convergent evolution proceeds via similar or different genetic and developmental mechanisms offers insight towards the repeatability and predictability of evolution. Oceanic populations of threespine stickleback fish, Gasterosteus aculeatus , have repeatedly colonized countless freshwater lakes and streams, where new diets lead to morphological adaptations related to feeding. Here, we show that heritable increases in branchial bone length have convergently evolved in two independently derived freshwater stickleback populations. In both populations, an increased bone growth rate in juveniles underlies the convergent adult phenotype, and one population also has a longer cartilage template. Using F 2 crosses from these two freshwater populations, we show that two quantitative trait loci (QTL) control branchial bone length at distinct points in development. In both populations, a QTL on chromosome 21 controls bone length throughout juvenile development, and a QTL on chromosome 4 controls bone length only in adults. In addition to these similar developmental profiles, these QTL show similar chromosomal locations in both populations. Our results suggest that sticklebacks have convergently evolved longer branchial bones using similar genetic and developmental programmes in two independently derived populations.