Riho Gross

Matrilinear phylogeography of Atlantic salmon ( Salmo salar L.) in Europe and postglacial colonization of the Baltic Sea area

Sixty‐four samples from 46 salmon populations totalling 2369 specimens were used for polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) analysis of the mitochondrial ND1 region. The final analyses included 3095 specimens from 60 populations in Northern Europe. A subsample was analysed by RFLP of ND3/4/5/6. Representative RFLP haplotypes from different parts of the distribution area were sequenced and the phylogeny of European haplotypes and their relations to the North American lineage was described. The four common European haplotypes derive from the ancestral ND1‐BBBA (rooting the European clade to the North American) by one‐step substitutions: AAAA < AABA < BBBA > BBBB. The Swedish west‐coast populations differ from the geographically close southern Baltic, indicating absence of inward and limited outward gene flow through the Danish straits during the last 8000 years. Within the Baltic Sea, only three ND1 haplotypes were detected and there was no variation for ND3/4/5/6. In the whole southern Baltic and in lakes Vänern, Ladoga and Onega the haplotype AABA dominated. Proposed postglacial colonization routes to the Baltic Sea are discussed in relation to the haplotype distribution pattern.

Genetic variability and structure of common carp (Cyprinus carpio) populations throughout the distribution range inferred from allozyme, microsatellite and mitochondrial DNA markers

Domesticated/captive stocks and wild/feral populations of common carp from Europe, Central Asia and East/South-East Asia were examined for allozyme (23 populations), microsatellite (11 populations) and mitochondrial DNA (21 populations) variation. Allozyme variability (1.06–1.81 alleles per locus, expected heterozygosity 0.006–0.136 at 16 loci) was much lower than microsatellite variability (2.5–14.0 alleles per locus, expected heterozygosity 0.426–0.887 at four loci). Differences in variability between domesticated/captive stocks and wild-caught ones were more pronounced at microsatellite loci than at allozyme loci, suggesting that microsatellites are better suited to detect population bottlenecks and loss of variation due to inbreeding. All but one European population were fixed for a single composite mtDNA haplotype, which also dominated in Central Asia but was completely missing in East/South-East Asia, indicating a single origin of European carp in Central Asia. All three classes of genetic markers clustered populations into two highly divergent groups: Europe/Central Asia and East/South-East Asia. Hierarchical partition of genetic diversity showed that for microsatellite loci most of variation was due to the within-population component while the highest proportion of mtDNA variation and substantial proportion of allozyme variation was accounted for by differences between geographical regions. Genetic data support the subspecies status of C. c. carpio assigned to the European carp and C. c. haematopterus assigned to the East/South-East Asian carp but do not justify a separate subspecies status (C. c. aralensis) for the Central Asian carp. As demonstrated for a wild/feral carp population from R. Danube, Germany, the genetic markers used in our study may be effectively applied to detect mixing and introgression of intra-species units in the presence of sufficient genetic differentiation.

Mitochondrial and nuclear DNA phylogeography of European grayling (Thymallus thymallus): evidence for secondary contact zones in central Europe

Mitochondrial and microsatellite DNA markers were applied to infer the phylogeography, intraspecific diversity and dynamics of the distributional history of European grayling (Thymallus thymallus) with focus on its central and northern European distribution range. Phylogenetic and nested clade analyses revealed at least four major mtDNA lineages, which evolved in geographical isolation during the Pleistocene. These lineages should be recognized as the basic evolutionary significant units (ESUs) for grayling in central and northern Europe. In addition, and in contrast to previous work on grayling, the results of Bayesian analysis of individual admixture coefficients, two‐dimensional scaling analysis and spatial analysis of molecular variance provided evidence for a high level of admixture among major lineages in contact zones between drainages (e.g. the low mountain range of Germany), most likely resulting from glacial perturbations and ancient river connections between drainages during the Pleistocene glaciations. Even within river systems, a high level of differentiation among populations was revealed as indicated by the microsatellite data. Grayling sampled from 29 sites displayed high levels of differentiation (overall FST = 0.367), a high number of private alleles and high bootstrap support for the genetic distance‐based population clusters across 12 loci. We specifically discuss our results in context of phylogeograpic studies on other European freshwater fish species with habitat preferences similar to those of grayling. Our study shows that both large‐scale phylogeographical and detailed genetic analyses on a fine scale are mandatory for developing appropriate conservation guidelines of endangered species.

Extensive immigration from compensatory hatchery releases into wild Atlantic salmon population in the Baltic sea: spatio-temporal analysis over 18 years

Genetic homogenization has been recognized as a serious threat in an increasing number of species, including many salmonid fishes. We assessed the rate and impact of immigration from the main hatchery stocks of Atlantic salmon in the Gulf of Bothnia into one of the largest wild salmon populations in the Baltic Sea, the River Vindelälven, within a temporal framework of 18 years (from 1985–2003). We provide genetic evidence based on mtDNA and microsatellite markers, using mixed-stock analysis, that a large proportion (66%) of fin-damaged spawners (n=181) caught in the Ume/Vindelälven during 1997–2003 originated from the hatcheries in the Rivers Ångermanälven, Luleälven and Ljusnan. The maximum-likelihood estimate of immigration rate from these hatcheries into the wild Vindelälven population was 0.068 (95% CI 0.021–0.128) over the studied time period (1985–2003) and reached up to a quarter (m=0.249, 95% CI 0.106–0.419) of the total population during 1993–2000. This resulted in significant (P<0.01) genetic homogenization trend between the wild Vindelälven population and hatchery stocks of the Ångermanälven and Luleälven. Our results demonstrate extensive straying from geographically distant hatchery releases into wild salmon population and emphasize the genetic risks associated with current large-scale stocking practices in the Baltic Sea.

Restriction fragment length polymorphism at the growth hormone 1 gene in Atlantic salmon (Salmo salar L.) and its association with weight among the offspring of a hatchery stock

Abstract Variation was studied within an 1825 bp fragment of the Atlantic salmon growth hormone 1 ( GH1 ) gene, encompassing the sequence from the first to the fourth exon. Two novel polymorphisms were detected in a hatchery strain of landlocked salmon population from the Lake Vanern, Sweden (spawns in the River Gullspang) after PCR amplification and subsequent digestion of the GH1 fragment with Taq I restriction enzyme: restriction site polymorphism within a 562 bp fragment and length polymorphism of a 187 bp fragment. Altogether six different banding patterns were observed that arose from random association of three haplotypes. In order to study possible association of the detected variation with the weight of one-year-old fish, progeny of the hatchery strain was graded into three size groups. Significant heterogeneity of the GH1 haplotype and genotype frequencies among the size groups was detected ( P P >0.05).

PCR–RFLP analysis of the mitochondrial ND-3/4 and ND-5/6 gene polymorphisms in the European and East Asian subspecies of common carp (Cyprinus carpio L.)

Abstract Polymorphism within the mitochondrial NADH-3,4 dehydrogenase ( ND-3/4 ) and NADH-5,6 dehydrogenase ( ND-5/6 ) gene regions was studied by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) analysis among common carp populations belonging to the European (two farmed strains and three wild populations) and East Asian (Amur wild carp, Vietnamese wild carp and Japanese Koi carp) subspecies, Cyprinus carpio carpio and C. c. haematopterus , respectively. Polymorphism was detected using eight and six restriction enzymes, respectively, and a total of seven composite haplotypes were identified. Both distance-based and maximum likelihood phylogenetic inference methods clustered the haplotypes into four distinct groups—the European (two haplotypes), Amur (two haplotypes), Vietnamese (two haplotypes) and Koi (one haplotype), and their distributions strictly follow the geographic origin of populations. The populations are clustered into two highly divergent groups (average net nucleotide divergence, 2.4%), the European and the East Asian populations, suggesting an ancient separation. Six enzymes ( Hin fI, Alu I, Hpa II and Taq I at ND-3/4 ; Eco 47I and Bsu RI at ND-5/6 ) yielded diagnostic restriction sites for discriminating between the European and East Asian maternal lineages that can be applied to monitoring of genetic purity of the European farmed strains.

Discovery and application of insertion-deletion (INDEL) polymorphisms for QTL mapping of early life-history traits in Atlantic salmon

BackgroundFor decades, linkage mapping has been one of the most powerful and widely used approaches for elucidating the genetic architecture of phenotypic traits of medical, agricultural and evolutionary importance. However, successful mapping of Mendelian and quantitative phenotypic traits depends critically on the availability of fast and preferably high-throughput genotyping platforms. Several array-based single nucleotide polymorphism (SNP) genotyping platforms have been developed for genetic model organisms during recent years but most of these methods become prohibitively expensive for screening large numbers of individuals. Therefore, inexpensive, simple and flexible genotyping solutions that enable rapid screening of intermediate numbers of loci (~75-300) in hundreds to thousands of individuals are still needed for QTL mapping applications in a broad range of organisms.ResultsHere we describe the discovery of and application of insertion-deletion (INDEL) polymorphisms for cost-efficient medium throughput genotyping that enables analysis of >75 loci in a single automated sequencer electrophoresis column with standard laboratory equipment. Genotyping of INDELs requires low start-up costs, includes few standard sample handling steps and is applicable to a broad range of species for which expressed sequence tag (EST) collections are available. As a proof of principle, we generated a partial INDEL linkage map in Atlantic salmon (Salmo salar) and rapidly identified a number of quantitative trait loci (QTLs) affecting early life-history traits that are expected to have important fitness consequences in the natural environment.ConclusionsThe INDEL genotyping enabled fast coarse-mapping of chromosomal regions containing QTL, thus providing an efficient means for characterization of genetic architecture in multiple crosses and large pedigrees. This enables not only the discovery of larger number of QTLs with relatively smaller phenotypic effect but also provides a cost-effective means for evaluation of the frequency of segregating QTLs in outbred populations which is important for further understanding how genetic variation underlying phenotypic traits is maintained in the wild.

Idengification of the origin of an Atlantic salmon (Salmo salar L.) population in a recently recolonized river in the Baltic Sea

The founder event in a recently recolonized salmon population in the Baltic Sea (Gulf of Finland) was investigated. To idengify the origin of the founders, four wild populations and two hatchery stocks were analysed using six microsatellite loci. The results of assignment tests and factorial correspondence analysis suggest that the initial recolonizers of the river Selja originated from the geographically nearest (7 km) wild population (river Kunda) but as the result of stocking activities, interbreeding between recolonizers and hatchery individuals has occurred in subsequent years. Although the hatchery releases are outnumbering the wild salmon recruitment in the Baltic Sea at present, our results suggest that the native populations may still have an important role in colonization processes of the former salmon rivers.

Application of heteroduplex analysis for detecting variation within the growth hormone 2 gene in Salmo trutta L. (brown trout)

A new method to detect variation at a single copy nuclear gene in brown trout, Salmo trutta L., is provided. The technique entails (i) selective gene amplification by the polymerase chain reaction (PCR), (ii) digestion of amplification products by restriction endonucleases to obtain fragments of suitable size, (iii) hybridization with heterologous DNA followed by denaturation and reannealing to obtain heteroduplex molecules, and (iv) screening for variation in polyacrylamide gels. Variation was studied within a growth hormone 2 gene 1489 bp segment and polymorphism was detected in two HinfI-digested fragments. Formation of different heteroduplex patterns in experimental mixtures of digested amplification products from brown trout and Atlantic salmon, Salmo salar L., allowed us to determine the genotype of the brown trout. Polymorphism was observed in four out of six studied populations.

Analysis of gene associated tandem repeat markers in Atlantic salmon (Salmo salar L.) populations: implications for restoration and conservation in the Baltic Sea

Patterns of genetic diversity and differentiation among five wild and four hatchery populations of Atlantic salmon in the Baltic Sea were assessed based on eight assumedly neutral microsatellite loci and six gene-associated markers, including four expressed sequence tag (EST) linked and two major histocompatibility complex (MHC) linked tandem repeat markers (micro- and mini-satellites). The coalescent simulations based on the method of Beaumont and Nichols (1996, Proc. R. Soc. Lond. Ser. B – Biol. Sci., 263, 1619–1626) indicated that two loci (MHCIIα and Ssa171, with the lowest and highest overall FST estimates, respectively) exhibited significant departures (P<0.05) from the neutral expectations. Another coalescent-based test for selective neutrality (Vitalis et al. 2001, Genetics, 158, 1811–1823) further supported the outlier status of the Ssa171 microsatellite locus but not of the MHCIIα linked minisatellite. In addition, actin related protein linked microsatellite locus was identified with this test as an outlier in six pairwise population comparisons. All genetic diversity estimates revealed more genetic variation in hatchery stocks than in the small wild salmon populations from the Gulf of Finland. However, the wild populations possessed alleles at gene-associated markers (e.g. MHCI and IGF) not found in the hatchery stocks, which together with moderate genetic differentiation and distinctive environmental conditions justifies the special conservation measures for the last remaining native salmon populations in the Gulf of Finland.