Differences in Salinity Tolerance and Gene Expression Between Two Populations of Atlantic Cod (Gadus morhua) in Response to Salinity Stress

Populations of marine fish, even from contrasting habitats, generally show low genetic differentiation at neutral genetic markers. Nevertheless, there is increasing evidence for differences in gene expression among populations that may be ascribed to adaptive divergence. Studying variation in salinity tolerance and gene expression among Atlantic cod (Gadus morhua) from two populations distributed across a steep salinity gradient, we observed high mortality (45% North Sea cod and 80% Baltic Sea cod) in a reciprocal common garden setup. Quantitative RT-PCR assays for expression of hsp70 and Na/K-ATPase α genes demonstrated significant differences in gene regulation within and between populations and treatment groups despite low sample sizes. Most interesting are the significant differences observed in expression of the Na/K-ATPase α gene in gill tissue between North Sea and Baltic cod. The findings strongly suggest that Atlantic cod are adapted to local saline conditions, despite relatively low levels of neutral genetic divergence between populations.     

Genetic population structure of turbot (Scophthalmus maximus L.) supports the presence of multiple hybrid zones for marine fishes in the transition zone between the Baltic Sea and the North Sea

Genetic population structure of turbot (Scophthalmus maximus L.) in the Northeast Atlantic was investigated using eight highly variable microsatellite loci. In total 706 individuals from eight locations with temporal replicates were assayed, covering an area from the French Bay of Biscay to the Aaland archipelago in the Baltic Sea. In contrast to previous genetic studies of turbot, we found significant genetic differentiation among samples with a maximum pairwise FST of 0.032. Limited or no genetic differentiation was found among samples within the Atlantic/North Sea area and within the Baltic Sea, suggesting high gene flow among populations in these areas. In contrast, there was a sharp cline in genetic differentiation going from the low saline Baltic Sea to the high saline North Sea. The data were explained best by two divergent populations connected by a hybrid zone; however, a mechanical mixing model could not be ruled out. A significant part of the genetic variance could be ascribed to variation among years within locality. Nevertheless, the population structure was relatively stable over time, suggesting that the observed pattern of genetic differentiation is biologically significant. This study suggests that hybrid zones are a common phenomenon for marine fishes in the transition area between the North Sea and the Baltic Sea and highlights the importance of using interspecific comparisons for inferring population structure in high gene flow species such as most marine fishes.     

Population genomic evidence for adaptive differentiation in the Baltic Sea herring

Detecting and estimating the degree of genetic differentiation among populations of highly mobile marine fish having pelagic larval stages is challenging because their effective population sizes can be large, and thus, little genetic drift and differentiation is expected in neutral genomic sites. However, genomic sites subject to directional selection stemming from variation in local environmental conditions can still show substantial genetic differentiation, yet these signatures can be hard to detect with low‐throughput approaches. Using a pooled RAD‐seq approach, we investigated genomewide patterns of genetic variability and differentiation within and among 20 populations of Atlantic herring in the Baltic Sea (and adjacent Atlantic sites), where previous low‐throughput studies and/or studies based on few populations have found limited evidence for genetic differentiation. Stringent quality control was applied in the filtering of 1 791 254 SNPs, resulting in a final data set of 68 182 polymorphic loci. Clear differentiation was identified between Atlantic and Baltic populations in many genomic sites, while differentiation within the Baltic Sea area was weaker and geographically less structured. However, outlier analyses – whether including all populations or only those within the Baltic Sea – uncovered hundreds of directionally selected loci in which variability was associated with either salinity, temperature or both. Hence, our results support the view that although the degree of genetic differentiation among Baltic Sea herring populations is low, there are many genomic regions showing elevated divergence, apparently as a response to temperature‐ and salinity‐related natural selection. As such, the results add to the increasing evidence of local adaptation in highly mobile marine organisms, and those in the young Baltic Sea in particular.     

Environmental correlates of population differentiation in Atlantic herring

The marine environment is characterized by few physical barriers, and pelagic fishes commonly show high migratory potential and low, albeit in some cases statistically significant, levels of genetic divergence in neutral genetic marker analyses. However, it is not clear whether low levels of differentiation reflect spatially separated populations experiencing gene flow or shallow population histories coupled with limited random genetic drift in large, demographically isolated populations undergoing independent evolutionary processes. Using information for nine microsatellite loci in a total of 1951 fish, we analyzed genetic differentiation among Atlantic herring from eleven spawning locations distributed along a longitudinal gradient from the North Sea to the Western Baltic. Overall genetic differentiation was low (theta = 0.008) but statistically significant. The area is characterized by a dramatic shift in hydrography from the highly saline and temperature stable North Sea to the brackish Baltic Sea, where temperatures show high annual variation. We used two different methods, a novel computational geometric approach and partial Mantel correlation analysis coupled with detailed environmental information from spawning locations to show that patterns of reproductive isolation covaried with salinity differences among spawning locations, independent of their geographical distance. We show that reproductive isolation can be maintained in marine fish populations exhibiting substantial mixing during larval and adult life stages. Analyses incorporating genetic, spatial, and environmental parameters indicated that isolating mechanisms are associated with the specific salinity conditions on spawning locations.     

Population structure of flounder (Platichthys flesus) in the Baltic Sea: differences among demersal and pelagic spawners

We found significant population structure and isolation by distance among samples of flounder (Platichthys flesus) in the Baltic, Kattegat and Skagerrak seas using microsatellite genetic markers. This pattern was almost entirely due to a difference between flounder that have demersal spawning in the northern Baltic, as compared to pelagic spawners in the southern Baltic and on the west coast of Sweden. Among demersal spawners we found neither genetic differentiation nor any isolation by distance among sampling sites. We speculate that demersal flounder are descendants of a population that colonized the Baltic previous to pelagic spawners. The demersal flounder may thus have had longer time to adapt to the low salinity in the Baltic, and accordingly display egg characteristics that make it possible to reproduce at the low salinity levels in the northern Baltic. Among pelagic spawners significant isolation by distance was detected. Pelagic spawners have previously been shown to display clinal variation in egg size, which allows them to float also at the moderate salinity levels up to the region north of the island Bornholm. Management units for harvesting should ideally be based on true biological populations, and for the commercially important flounder up to 15 different management stocks in the Baltic have been suggested. We could not find a population genetic foundation for such a high number of management units, and our data suggest three management units: the northern Baltic (demersal populations), southern Baltic with the Oresund straits and the most northwestern sampling sites (Skagerrak, Kattegat and North Sea).     

Local adaptation and oceanographic connectivity patterns explain genetic differentiation of a marine diatom across the North Sea–Baltic Sea salinity gradient

Drivers of population genetic structure are still poorly understood in marine micro‐organisms. We exploited the North Sea–Baltic Sea transition for investigating the seascape genetics of a marine diatom, Skeletonema marinoi. Eight polymorphic microsatellite loci were analysed in 354 individuals from ten locations to analyse population structure of the species along a 1500‐km‐long salinity gradient ranging from 3 to 30 psu. To test for salinity adaptation, salinity reaction norms were determined for sets of strains originating from three different salinity regimes of the gradient. Modelled oceanographic connectivity was compared to directional relative migration by correlation analyses to examine oceanographic drivers. Population genetic analyses showed distinct genetic divergence of a low‐salinity Baltic Sea population and a high‐salinity North Sea population, coinciding with the most evident physical dispersal barrier in the area, the Danish Straits. Baltic Sea populations displayed reduced genetic diversity compared to North Sea populations. Growth optima of low salinity isolates were significantly lower than those of strains from higher native salinities, indicating local salinity adaptation. Although the North Sea–Baltic Sea transition was identified as a barrier to gene flow, migration between Baltic Sea and North Sea populations occurred. However, the presence of differentiated neutral markers on each side of the transition zone suggests that migrants are maladapted. It is concluded that local salinity adaptation, supported by oceanographic connectivity patterns creating an asymmetric migration pattern between the Baltic Sea and the North Sea, determines genetic differentiation patterns in the transition zone.     

Regional environmental pressure influences population differentiation in turbot (Scophthalmus maximus)

Unravelling the factors shaping the genetic structure of mobile marine species is challenging due to the high potential for gene flow. However, genetic inference can be greatly enhanced by increasing the genomic, geographical or environmental resolution of population genetic studies. Here, we investigated the population structure of turbot (Scophthalmus maximus) by screening 17 random and gene‐linked markers in 999 individuals at 290 geographical locations throughout the northeast Atlantic Ocean. A seascape genetics approach with the inclusion of high‐resolution oceanographical data was used to quantify the association of genetic variation with spatial, temporal and environmental parameters. Neutral loci identified three subgroups: an Atlantic group, a Baltic Sea group and one on the Irish Shelf. The inclusion of loci putatively under selection suggested an additional break in the North Sea, subdividing southern from northern Atlantic individuals. Environmental and spatial seascape variables correlated marginally with neutral genetic variation, but explained significant proportions (respectively, 8.7% and 10.3%) of adaptive genetic variation. Environmental variables associated with outlier allele frequencies included salinity, temperature, bottom shear stress, dissolved oxygen concentration and depth of the pycnocline. Furthermore, levels of explained adaptive genetic variation differed markedly between basins (3% vs. 12% in the North and Baltic Sea, respectively). We suggest that stable environmental selection pressure contributes to relatively strong local adaptation in the Baltic Sea. Our seascape genetic approach using a large number of sampling locations and associated oceanographical data proved useful for the identification of population units as the basis of management decisions.     

Adaptive and Neutral Genetic Variation and Colonization History of Atlantic Salmon, Salmo salar

Synopsis I combined neutral microsatellite markers with the major histocompatibility complex (MHC) class IIB to study genetic differentiation and colonization history in Atlantic salmon, Salmo salar, in the Baltic Sea and in the north-eastern Atlantic. Baltic salmon populations have lower levels of microsatellite genetic variation, in terms of heterozygosity and allelic richness than Atlantic populations, confirming earlier findings with other genetic markers, suggesting that the Baltic Sea populations have been exposed to genetic bottlenecks, most likely at a founding event. On the other hand, the level of MHC variation was similar in the Baltic and in the north-eastern Atlantic, indicating that positive balancing selection has increased the level of MHC-variation. Both microsatellite and MHC class IIB genetic variation give strong support to the hypothesis that the Baltic salmon are of a biphyletic origin, the southern population in this study is strongly differentiated from both the northern Baltic salmon populations and from the north-eastern Atlantic populations. Salmon may have colonized the northern Baltic Sea either from the south, via the so called “N?rke strait” or from the north, via a proposed historical connection between the White Sea and the northern Baltic. At microsatellites, no significant isolation-by distance was found at either colonization route. At the MHC, populations were significantly isolated by distance when assuming that colonization occurred via the “N?rke strait”.     

Intraspecific variation in expression of candidate genes for osmoregulation, heme biosynthesis and stress resistance suggests local adaptation in European flounder (Platichthys flesus)

Despite the recent discovery of significant genetic structuring in a large number of marine organisms, the evolutionary significance of these often minute genetic differences are still poorly understood. To elucidate the adaptive relevance of low genetic differentiation among marine fish populations, we studied expression differences of osmoregulatory and stress genes in genetically weakly differentiated populations of the European flounder (Platichthys flesus), distributed across a natural salinity gradient. Flounders were maintained in a long-term reciprocal transplantation experiment mimicking natural salinities in the North Sea and the Baltic Sea. Applying real-time quantitative PCR and microarray analysis we studied expression of four candidate genes (hsp70, angiotensinogen, Na/K-ATPase-alpha and 5-aminolevulinic acid synthase (ALAS)) in gill, kidney and liver tissues. Genes involved in osmoregulative processes (Na/K-ATPases-alpha and angiotensinogen) showed highly plastic but similar expression in the two populations dependent on environmental salinity. However, we observed a unique sixfold up-regulation of hsp70 in kidney tissue of flounder from the North Sea following long-term acclimation to Baltic salinities. Similarly, significant differences between North Sea and Baltic flounders in expression of ALAS in response to different salinities were found in gill and liver tissue. These findings strongly suggest that gene expression in flounders is shaped by adaptation to local environmental conditions. This identification of adaptive differences in high gene flow marine organisms adds a new dimension to our current understanding of evolutionary processes in the sea and is of paramount importance for identification, protection and sustainable management of marine biodiversity.     

An environmental gradient dominates ecological and genetic differentiation of marine invertebrates between the North and Baltic Sea

Environmental gradients have emerged as important barriers to structuring populations and species distributions. We set out to test whether the strong salinity gradient from the marine North Sea to the brackish Baltic Sea in northern Europe represents an ecological and genetic break, and to identify life history traits that correlate with the strength of this break. We accumulated mitochondrial cytochrome oxidase subunit 1 sequence data, and data on the distribution, salinity tolerance, and life history for 28 species belonging to the Cnidaria, Crustacea, Echinodermata, Mollusca, Polychaeta, and Gastrotricha. We included seven non‐native species covering a broad range of times since introduction, in order to gain insight into the pace of adaptation and differentiation. We calculated measures of genetic diversity and differentiation across the environmental gradient, coalescent times, and migration rates between North and Baltic Sea populations, and analyzed correlations between genetic and life history data. The majority of investigated species is either genetically differentiated and/or adapted to the lower salinity conditions of the Baltic Sea. Species exhibiting population structure have a range of patterns of genetic diversity in comparison with the North Sea, from lower in the Baltic Sea to higher in the Baltic Sea, or equally diverse in North and Baltic Sea. Two of the non‐native species showed signs of genetic differentiation, their times since introduction to the Baltic Sea being about 80 and >700 years, respectively. Our results indicate that the transition from North Sea to Baltic Sea represents a genetic and ecological break: The diversity of genetic patterns points toward independent trajectories in the Baltic compared with the North Sea, and ecological differences with regard to salinity tolerance are common. The North Sea–Baltic Sea region provides a unique setting to study evolutionary adaptation during colonization processes at different stages by jointly considering native and non‐native species.     

Population genetic structure of mussels from the Baltic Sea

In a macrogeographic survey, the population genetic structure of mussels from various regions of the Baltic Sea, a large semi-enclosed brackish-water basin, was examined with reference toMytilus edulis andM. galloprovincialis samples from the North Sea, Irish coast and southern Portugal. Electrophoretically detectable variation was analysed at 6 polymorphic enzyme loci (Ap, Est-D, Lap-2, Odh, Pgi andPgm). Evidence was provided of a remarkably large amount of biochemical genetic differentiation among ecologically and morphologically divergent mussel populations in the Baltic. Patterns of allele frequencies in low-salinity populations from the area of the Baltic Proper were demonstrated to be widely homogeneous but contrast strongly with those of the western Baltic, the latter resembling populations from marine habitats of the North Sea. Associated with a pronounced salinity gradient, the spatial heterogeneity in gene-pool structure is indicated by steep clines of allele frequency changes in the area of the eastern Danish isles. The adaptive significance of the observed allozymic variation is suggested. From genetic distance estimates, the subdivision of population structure is discussed in relation to the significant amount of differentiation detected withinMytilus populations to date and to the evolutionary time required for the divergence of Baltic mussel populations. The allozymic data provide evidence for the genetic distinctiveness of mussels from the low-salinity areas of the Baltic. Their position at the specific or subspecific level of classification requires further consideration.     

Patterns of Post-Glacial Genetic Differentiation in Marginal Populations of a Marine Microalga

This study investigates the genetic structure of an eukaryotic microorganism, the toxic dinoflagellate Alexandrium ostenfeldii, from the Baltic Sea, a geologically young and ecologically marginal brackish water estuary which is predicted to support evolution of distinct, genetically impoverished lineages of marine macroorganisms. Analyses of the internal transcribed spacer (ITS) sequences and Amplified Fragment Length Polymorphism (AFLP) of 84 A. ostenfeldii isolates from five different Baltic locations and multiple external sites revealed that Baltic A. ostenfeldii is phylogenetically differentiated from other lineages of the species and micro-geographically fragmented within the Baltic Sea. Significant genetic differentiation (F _ST) between northern and southern locations was correlated to geographical distance. However, instead of discrete genetic units or continuous genetic differentiation, the analysis of population structure suggests a complex and partially hierarchic pattern of genetic differentiation. The observed pattern suggests that initial colonization was followed by local differentiation and varying degrees of dispersal, most likely depending on local habitat conditions and prevailing current systems separating the Baltic Sea populations. Local subpopulations generally exhibited low levels of overall gene diversity. Association analysis suggests predominately asexual reproduction most likely accompanied by frequency shifts of clonal lineages during planktonic growth. Our results indicate that the general pattern of genetic differentiation and reduced genetic diversity of Baltic populations found in large organisms also applies to microscopic eukaryotic organisms.     

Diversity of Aphanizomenon flos-aquae (cyanobacterium) populations along a Baltic Sea salinity gradient

Colony-forming cyanobacteria of the genus Aphanizomenon form massive blooms in the brackish water of the Baltic Sea during the warmest summer months. There have been recent suggestions claiming that the Baltic Sea Aphanizomenon species may be different from Aphanizomenon flos-aquae found in lakes. In this study, we examined variability in the morphology and 16S-23S rRNA internal transcribed spacer (ITS) sequences of A. flos-aquae populations along a salinity gradient from a string of lakes to a fjord-like extension of the Baltic Sea to the open Baltic Sea. Morphological differences among the populations were negligible. We found that the Baltic Sea was dominated (25 out of 27 sequences) by one ITS1-S (shorter band of ITS 1 [ITS1]) genotype, which also was found in the lakes. The lake populations of A. flos-aquae tended to be genetically more diverse than the Baltic Sea populations. Since the lake ITS1-S genotypes of A. flos-aquae are continuously introduced to the Baltic Sea via inflowing waters, it seems that only one ITS1 genotype is able to persist in the Baltic Sea populations. The results suggest that one of the ITS1-S genotypes found in the lakes is better adapted to the conditions of the Baltic Sea and that natural selection removes most of the lake genotypes from the Baltic Sea A. flos-aquae populations.     

The Baltic Sea as a model system for studying postglacial colonization and ecological differentiation,exemplified by the red alga Ceramium tenuicorne

The Baltic Sea provides a unique model system for studying genetic effects of postglacial colonization and ecological differentiation, because all marine organisms must have immigrated after the opening of the Danish Straits 8000 years ago and responded to the development of the steep Skagerrak-Baltic salinity gradient. The red alga Ceramium tenuicorne shows conspicuous variation in growth and reproduction along this gradient. Herein we obtained reproductive data coupled with two types of molecular markers, one organellar (cox2-3 spacer sequences of mitochondrial DNA; mtDNA) and one mainly nuclear (random amplified polymorphic DNAs; RAPDs). Nine main populations were sampled in a nested spatial hierarchy including three salinity regions (Oslofjorden, Kattegat, and the Baltic Sea), and nine additional populations were sampled for the mtDNA analysis. Asexuality was frequent at low (Baltic) and medium (Kattegat) salinities but virtually absent at the highest salinity (Oslofjorden). Five mtDNA haplotypes were observed, of which two highly divergent ones were common. One was restricted to and fixed in Oslofjorden, and the other, which was closely related to the three rare haplotypes, was found from southernmost Norway via Kattegat into the Baltic. The RAPD data revealed, on the other hand, a continuous cline corresponding to the salinity gradient, with 27.4% divergence among salinity regions and most of the variation stored at the smallest spatial scale analysed (64.2%; within 1 m2 subpopulations). The combined data suggest colonization from a diverse Atlantic glacial gene pool followed by (1) lineage sorting of ancestral mtDNA polymorphisms and (2) strong differential selection among nuclear genotypes along the salinity gradient, including selection for nonrecombinant multiplication of those best fit to the marginal low-salinity habitats.     

Polymorphism in developmental mode and its effect on population genetic structure of a spionid polychaete, Pygospio elegans

Population genetic structure of sedentary marine species is expected to be shaped mainly by the dispersal ability of their larvae. Long-lived planktonic larvae can connect populations through migration and gene flow, whereas species with nondispersive benthic or direct-developing larvae are expected to have genetically differentiated populations. Poecilogonous species producing different larval types are ideal when studying the effect of developmental mode on population genetic structure and connectivity. In the spionid polychaete Pygospio elegans, different larval types have been observed between, and sometimes also within, populations. We used microsatellite markers to study population structure of European P. elegans from the Baltic Sea (BS) and North Sea (NS). We found that populations with planktonic larvae had higher genetic diversity than did populations with benthic larvae. However, this pattern may not be related to developmental mode, since in P. elegans, developmental mode may be associated with geography. Benthic larvae were more commonly seen in the brackish BS and planktonic larvae were predominant in the NS, although both larval types also are found from both areas. Significant isolation-by-distance (IBD) was found overall and within regions. Most of the pair-wise F(ST) comparisons among populations were significant, although some geographically close populations with planktonic larvae were found to be genetically similar. However, these results, together with the pattern of IBD, autocorrelation within populations, as well as high estimated local recruitment, suggest that dispersal is limited in populations with planktonic larvae as well as in those with benthic larvae. The decrease in salinity between the NS and BS causes a barrier to gene flow in many marine species. In P. elegans, low, but significant, differentiation was detected between the NS and BS (3.34% in AMOVA), but no clear transition zone was observed, indicating that larvae are not hampered by the change in salinity.     

Local adaptation to salinity in the three‐spined stickleback?

Different lines of evidence suggest that the occurrence and extent of local adaptation in high gene flow marine environments – even in mobile and long‐lived vertebrates with complex life cycles – may be more widespread than earlier thought. We conducted a common garden experiment to test for local adaptation to salinity in Baltic Sea sticklebacks (Gasterosteus aculeatus). Fish from three different native salinity regimes (high, mid and low) were subjected to three salinity treatments (high, mid and low) in a full‐factorial experimental design. Irrespective of their origin, fish subjected to low (and mid) salinity treatments exhibited higher juvenile survival, grew to largest sizes and were in better condition than fish subjected to the high salinity treatment. However, a significant interaction between native and treatment salinities – resulting mainly from the poor performance of fish native to low salinity in the high salinity treatment – provided clear cut evidence for adaptation to local variation in salinity. Additional support for this inference was provided by the fact that the results concur with an earlier demonstration of significant differentiation in a number of genes with osmoregulatory functions across the same populations and that the population‐specific responses to salinity treatments exceeded that to be expected by random genetic drift.     

Impact of 21st century climate change on the Baltic Sea fish community and fisheries

The Baltic Sea is a large brackish semienclosed sea whose species-poor fish community supports important commercial and recreational fisheries. Both the fish species and the fisheries are strongly affected by climate variations. These climatic effects and the underlying mechanisms are briefly reviewed. We then use recent regional – scale climate – ocean modelling results to consider how climate change during this century will affect the fish community of the Baltic and fisheries management. Expected climate changes in northern Europe will likely affect both the temperature and salinity of the Baltic, causing it to become warmer and fresher. As an estuarine ecosystem with large horizontal and vertical salinity gradients, biodiversity will be particularly sensitive to changes in salinity which can be expected as a consequence of altered precipitation patterns. Marine-tolerant species will be disadvantaged and their distributions will partially contract from the Baltic Sea; habitats of freshwater species will likely expand. Although some new species can be expected to immigrate because of an expected increase in sea temperature, only a few of these species will be able to successfully colonize the Baltic because of its low salinity. Fishing fleets which presently target marine species (e.g. cod, herring, sprat, plaice, sole) in the Baltic will likely have to relocate to more marine areas or switch to other species which tolerate decreasing salinities. Fishery management thresholds that trigger reductions in fishing quotas or fishery closures to conserve local populations (e.g. cod, salmon) will have to be reassessed as the ecological basis on which existing thresholds have been established changes, and new thresholds will have to be developed for immigrant species. The Baltic situation illustrates some of the uncertainties and complexities associated with forecasting how fish populations, communities and industries dependent on an estuarine ecosystem might respond to future climate change.     

Effects of temperature and salinity on embryonic development of turbot (Scophthalmus maximus L.) from the North Sea, and comparisons with Baltic populations

Optimum temperature and salinity conditions for viable hatch were studied for turbot (Scophthalmus maximus L.) from the North Sea. Temperatures ranging from 6 to 22°C and salinities from 5 to 35‰ were used. Optimum conditions were observed to be between 12 and 18°C at salinities between 20 and 35‰. This contrasted with corresponding data for turbot from the southern Baltic proper, according to which survival sharply decreased in temperatures below 14°C and was high in salinities of 10 to 15‰. Thus, it is concluded that Baltic and Atlantic turbot should be considered as different races.