V. A. Brykov

Development of Microsatellite Markers for Japanese Scallop (Mizuhopecten yessoensis) and Their Application to a Population Genetic Study

A bstractThe Japanese scallop (Mizuhopectenyessoensis) is one of the main fishery products in Japan, but with the expansion of culture operations of the Japanese scallop, various problems have been encountered including high mortality, poor growth, poor seed production, and so on. Moreover, there is concern that many years of cultivation may have affected the genetic structure of the scallop population. To approach these problems and concerns, we developed microsatellite markers as a molecular tool for population genetic studies. By using 4 microsatellite markers as well as a mitochondrial marker, we investigated the genetic structure of samples from the islands of Hokkaido (14 populations) and Honshu (Tohoku, 3 populations) in Japan, and south Primorye (4 populations) in Russia. All the populations sampled had high genetic diversity (average expected heterozygosity, 0.7011 to 0.7622; haplotype diversity, 0.6090 to 0.8848), and almost all showed a tendency of homozygote excess, which was significant in 2 populations. Hierarchical analysis of molecular variance tests based on the microsatellite and mitochondrial markers indicated that the 3 geographic regions were genetically divergent from one another, with little evidence of divergence within regions. Homogeneity in allele frequency distributions between natural and cultured scallops and allele frequency stability over a period of 2 decades indicated that the culturing operations have probably not had a substantial effect on the genetic structure of the populations.

High male incidence and evolutionary implications of triploid form in northeast Asia Carassius auratus complex

Carassius auratus complex is believed to originate from East Eurasia and consist of diploid and triploid forms. Diploid form reproduces sexually, whereas triploid form possesses mixture modes of unisexual gynogenesis and sexual reproduction, which makes it a unique case to study evolutionary issues among vertebrates. In this study, we identified 337 triploid individuals from 386 specimens of Carassius auratus complex sampled from 4 different sites of Xingkai Lake and Suifen River on the northeast Asia transboundary areas of Russia and China, and found that triploids were ubiquitous, whereas diploids existed only in SII site of Suifen River. Triploid males were detected in all surveyed sites, and an unusually high triploid male incidence (23%) was found in the Chinese reach of Suifen River. Then, nuclear and cytoplasmic markers were used to analyze their genetic diversity and phylogenetic relationship. A total of 61 distinct tf alleles and 35 mtDNA CR haplotypes were revealed. Higher genetic diversity and divergence were confirmed in triploids than in diploids, and identical genetic background between triploid males and females was demonstrated. Moreover, evolutionary implications and roles of triploid males were suggested in population proliferation and diversity creation of the triploid form.

Differences between two subspecies of Dolly Varden, Salvelinus malma, revealed by RFLP-PCR analysis of mitochondrial DNA

Genetic differences between two subspecies of Dolly Varden, northern Salvelinus malma malma and southern Salvelinus malma krascheninnikovi, from rivers of eastern Russia were studied. Mitochondrial DNA was analyzed by restriction fragment length polymorphisms (RFLP) performed on products amplified with polymerase chain reaction. Three adjacent segments (approximately 7670 bp), comprising 47% of the mitochondrial genome were used: two encoding the five complete NADH dehydrogenase subunits and the other the cytochrome b gene and the control region (D-loop). Total composite haplotypes 46 were found among 136 fishes using RFLP analysis with 14 restriction enzymes. The amount of nucleotide divergence between haplotypes of two subspecies of Dolly Varden was estimated to be approximately 4%. The differences in the level of nucleotide diversity, mismatch distribution between haplotypes, and population-genetic structure of two subspecies of Dolly Varden suggest that these two forms have existed separately for a long time.

Population Genetic Structure and Phylogeography of Masu Salmon (Oncorhynchus Masou Masou) Inferred from Mitochondrial and Microsatellite DNA Analyses

The population genetic structure and phylogeography of masu salmon were investigated by using variation in the mitochondrial NADH dehydrogenase subunit 5 gene (ND5) and six polymorphic microsatellite loci among a total of 895 fish representing 18 populations collected from Japan (9), Russia (7), and Korea (2) from 2000 to 2008. An analysis of ND5 nucleotide sequences revealed 22 variable sites in about 560 bp in the 5′ half of the gene, which defined 20 haplotypes, including some associated with geographical regions. Haplotype and nucleotide diversities were greater in the populations in Japan and Korea than in those in Russia, indicating greater genetic diversity in the Japanese and Korean populations than in the Russian populations. All the microsatellite loci examined showed a high level of variation, but the expected heterozygosity indicated a similar level of genetic diversity among the populations of the three regions, contrary to the results for ND5. However, AMOVA and pairwise population F ST estimates for both ND5 and the microsatellite markers indicated a similar pattern of moderate genetic differentiation among populations of the three regions, and large population groups on the coasts of the Sea of Japan, Sea of Okhotsk, and Pacific Ocean in the Far East. From a mismatch distribution analysis and neutrality test, the observed genetic structure appears to have been influenced primarily by bottlenecks during glacial periods and population expansions during interglacial periods in the late Pleistocene.

Phylogeography of northern Dolly Varden Salvelinus malma (Salmoniformes: Salmonidae) from Asia and North America: An analysis based on the mitochondrial DNA genealogy

Spawning in habitats affected by Pleistocene glacial advances over most of its natural range, northern Dolly Varden Salvelinus malma malma typifies Arctic fauna distributed in northeastern Asia and northwestern North America. We reconstructed a genealogy of mtDNA haplotypes from 27 Alaskan and Asian populations to study the influence of historical events on the phylogeography and contemporary population genetic structure. Analysis of molecular variance partitioned most of the mtDNA variability to the intrapopulation component (72.5%) with much reduced differences between populations (21.1%) and regions (6.4%). Similar patterns of variation apparent from hierarchical diversity and nested clade phylogeographical analysis (NCPA) of mtDNA haplotypes identify weak spatial differentiation and low levels of divergence. These findings suggest (1) that demographic history has been influenced by historical range expansions and recent isolation by distance, (2) that present populations from Asia and North America were colonized from one main Beringian Refugium, and (3) that this taxon’s ancestral population probably experienced a bottleneck in the Beringian Refugium during the late Pleistocene (Wisconsin) glacial period. The genealogical and NCPA analyses, and mismatch distribution of S. m. malma mtDNA haplotypes do not confirm the assumptions about presence of the two refugia on the territories of the Beringian Land, in which allopatric S. m. malma ancestral populations evolved, and independent origin of the Sea of Okhotsk populations.

Mitochondrial DNA variation in northwestern Bering Sea walleye pollock, Theragra chalcogramma (Pallas)

Genetic variability of the highly valuable gadid species, walleye pollock, Theragra chalcogramma (Pallas 1811), from five spatially separated northwestern Bering Sea areas (Ozernoi Bay, Olutorskyi Bay, Koryak shelf, Navarin region, and Anadyr Gulf) was investigated. Haplotype diversity within the samples ranged from 0.8788±0.0393 to 0.9436±0.0162. Nucleotide diversity within the samples ranged from 0.0108 to 0.0127. Nucleotide diversity among the regional collections ranged from 0% to 0.18%. Walleye pollock from the Anadyr Gulf appeared genetically separate from the other four samples in a clustering of genetic distances. Total heterogeneity among all five samples was significant, while there was no heterogeneity among the four samples excluding that from the Anadyr Gulf. Pair-wise comparisons using tests for heterogeneity and Fst supported the dendrogram of genetic distances in that only the collection from the Anadyr Gulf was significantly different from the others. The observed pattern of genetic differentiation among walleye pollock from the northwestern Bering Sea presumably emerged as a result of a population of the species subdividing in the northernmost part of its geographic range, though further analysis is needed to verify this supposition.

Phylogenetic relationships of silver crusian carp Carassius auratus gibelio, C. auratus cuvieri, crucian carp Carassius carassius, and common carp Cyprinus carpio as inferred from mitochondrial DNA variation

PCR-RFLP analysis of the ND3/ND4L/ND4 and 12S/16S rRNA regions and nucleotide sequence variation of the cytochrome b gene were used to study the mtDNA divergence in species of the family Cyprinidae, to examine the phylogenetic relationships of the species, and to identify their taxonomic status. The results indicated that an ancestral form diverged into silver crucian carp and crucian carp after its separation from the common carp lineage. The divergence of continental Carassius auratus gibelio and Japanese C. auratus cuvieri occurred more recently. Two well distinguishable mtDNA phylogroups, suggesting divergent evolution, were observed in continental C. auratus gibelio populations. The divergence was possibly related to the formation of two silver crucian carp groups with different types of reproduction, triploid gynogenetic and diploid gonochoric. At the same time, the results supported the high probability of current genetic exchange between the forms. In view of these findings and high morphological similarity of the two forms, they were not considered to be separate species.

[Analysis of mtDNA and Nuclear Markers Points to Homoploid Hybrid Origin of the New Species of Far Eastern Redfins of the Genus Tribolodon (Pisces, Cyprinidae)].

The nucleotide sequences of two mitochondrial genes (cytochrome c oxidase subunit I, COI, and cytochrome b, cyt b) and four nuclear genes (growth hormone gene GH1, ribosomal protein S7 gene RP1, recombination activating gene RAG1, and rhodopsin gene RH) from the Far Eastern redfins of the genus Tribolodon were examined to clarify the status of the southern form of T. hakonensis. Nucleotide sequence analysis of the mitochondrial genes showed differences of 2.6% between individuals of T. hakonensis inhabiting the north and south of the range. Analysis of the nuclear genes showed that Tribolodon sp. (southern form of T. hakonensis) has a mosaic of nuclear genes received from the Pacific redfin T. brandtii and big-scaled redfin T. hakonensis. It is suggested that the new species could have formed as a result of homoploid hybridization between the true T. hakonensis (by original description) and T. brandtii, which probably made it possible for this species to occupy a new ecological niche.

Divergence of Mitochondrial DNA of Two Dolly Varden Trout (Salvelinus malma; Salmonidae: Pisces) Subspecies

Chars of the genus Salvelinus are characterized by a large number of intraspecies forms and ecotypes, as well as by a low degree of species divergence, because this group is relatively young. This hinders evolutionary and taxonomic studies of chars. Most researchers currently accept the Dolly Varden trout, Salvelinus malma (Walbaum) as an independent species and distinguish three subspecies or intraspecies forms morphologically and karyologically differing from one another: S. malma malma, S. malma krascheninnikovi , and S. malma lordi [1]. Although the morphology of the Dolly Varden trout has been studied in detail [2–4], the taxonomic status of its geographic forms is still a subject of discussion.

Microsatellite variability in chum salmon of Primorye

In this study, 12 samples of chum salmon from the southern and central parts of Primorye were studied with ten microsatellite loci. All studied localities of chum salmon of Primorye formed three main genetically different groups: (1) the Narva–Barabashevka–Ryazanovka cluster of southern Primorye, (2) Kievka River, and (3) Avvakumovka River. The revealed genetic heterogeneity of chum salmon showed clear population structure in accordance with the geographical location of the samples. The study suggests that, for the purposes of artificial reproduction of chum salmon, it is desirable to perform egg planting with regard to the described population structure of chum salmon of Primorye.