Nobuhiko Taniguchi

Microsatellite multiplex panels for population genetic analysis of red sea bream Pagrus major

In this study, multiplex PCR panels were developed to amplify 20 microsatellite markers for red sea bream, Pagrus major, with the aim of reducing labor and material costs associated with genetic analysis of this species. The usefulness of these panels was validated in 200 fish collected at five sampling locations. The occurrence of null alleles was suggested at five markers, which were dropped from further analysis. The remaining 15 microsatellite loci showed high levels of heterozygosity (HE range 0.34–0.97, HO range 0.32–1) and a wide range in the number of alleles per locus (A; range 5–46). Increasing the number of microsatellite loci from three to ten and 15 improved the performance of the panels for population differentiation (FST) and estimation of Nei’s (DS) genetic distance. The results of this study confirm the usefulness of genotyping a large number of microsatellite DNA markers to expand our knowledge of red sea bream population genetics.

Allozyme variation of littleneck clam Ruditapes philippinarum and genetic mixture analysis of foreign clams in Ariake Sea and Shiranui Sea off Kyushu Island, Japan

Allozyme variation of the littleneck clam Ruditapes philippinarum was evaluated in four samples from Nameishi and Matsuo in the Ariake Sea, Ryugatake and Ushibuka in the Shiranui Sea off Kyushu Island, Japan, and in one sample from Jinzhou, China, in the Bohai Sea. A Ruditapes bruguieri sample imported from the Korean Bay off Nampo, North Korea was also studied. Among the R. philippinarum samples, heterozygosity varied from 0.265 to 0.301 and Fis estimates indicated significant homozygosity excess in 15 of 40 loci analyzed. Deviations from Hardy-Weinberg equilibrium were significant in all samples (P<0.05). Pairwise FST estimates indicate that genetic differences between the Chinese and Japanese samples were very low, but significantly different from zero. Mixture proportions with 95% confidence intervals of Chinese R. philippinarum in Nameishi and Matsuo were estimated at 0.4098 [0.2512, 0.5705] and 0.4899 [0.3262, 0.6540], respectively. However, genetic invasion of stocked Chinese R. philippinarum into wild populations in the Ariake Sea remains uncertain due to the low precision of the estimates caused by the high similarity of allele frequencies between Jinzhou and the Ariake Sea.

Effects of Large-Scale Releases on the Genetic Structure of Red Sea Bream (Pagrus major, Temminck et Schlegel) Populations in Japan.

Large-scale hatchery releases are carried out for many marine fish species worldwide; nevertheless, the long-term effects of this practice on the genetic structure of natural populations remains unclear. The lack of knowledge is especially evident when independent stock enhancement programs are conducted simultaneously on the same species at different geographical locations, as occurs with red sea bream (Pagrus major, Temminck et Schlegel) in Japan. In this study, we examined the putative effects of intensive offspring releases on the genetic structure of red sea bream populations along the Japanese archipelago by genotyping 848 fish at fifteen microsatellite loci. Our results suggests weak but consistent patterns of genetic divergence (F ST = 0.002, p < 0.001). Red sea bream in Japan appeared spatially structured with several patches of distinct allelic composition, which corresponded to areas receiving an important influx of fish of hatchery origin, either released intentionally or from unintentional escapees from aquaculture operations. In addition to impacts upon local populations inhabiting semi-enclosed embayments, large-scale releases (either intentionally or from unintentional escapes) appeared also to have perturbed genetic structure in open areas. Hence, results of the present study suggest that independent large-scale marine stock enhancement programs conducted simultaneously on one species at different geographical locations may compromise native genetic structure and lead to patchy patterns in population genetic structure.

Inference of Potential Genetic Risks Associated with Large-Scale Releases of Red Sea Bream in Kanagawa Prefecture, Japan Based on Nuclear and Mitochondrial DNA Analysis

Since 1978, millions of hatchery-reared red sea bream (Pagrus major) juveniles have been released in Sagami Bay and Tokyo Bay in Kanagawa Prefecture, Japan. The stock enhancement program has contributed to total catch; however, no information regarding the genetic interactions with wild counterparts is available. Here, we combined 15 microsatellite loci and mitochondrial D-loop sequencing to characterize the genetic resources of red sea bream in Sagami Bay and Tokyo Bay and to elucidate the potential harmful genetic effects associated with fish releases. Both types of markers evidenced higher levels of genetic diversity in wild samples (SB and TB) compared with offspring before stocking (H07 and H08) as well as a hatchery-released sample recaptured in Sagami Bay (HR). Microsatellite FST estimates and Bayesian clustering analysis found significant genetic differences among samples (FSTu2009=u20090.013–0.054), except for the two wild samples (FSTu2009=u20090.002) and HR vs. H07 (FSTu2009=u20090.007). On the other hand, mitochondrial-based ФST suggested haplotypic similarity between SB, H07, and HR. The low effective number of females contributing to the offspring over multiple generations may be responsible for the lack of haplotypic differentiation. Moreover, the putative hatchery origin to three fish (8xa0%) without deformity in the inter-nostril epidermis was inferred for the first time. Our results showed the usefulness of combining nuclear and mitochondrial markers to elucidate genetic interactions between hatchery-released and wild red sea bream and warned about potential harmful genetic effects should interbreeding takes place.

Estimation of the mode of inheritance of thermal tolerance in the guppy Poecilia reticulata

High water temperature influences the survival, growth, and maturation of fish. Genetically characterizing thermal tolerance is one of the most important subjects in fish culture. To identify the genetic characterization of thermal tolerance, this characteristic was compared among strains, and among parents and their offspring, in the guppy Poecilia reticulata. In the strain comparison, significant differences in survival rate were observed among the five strains examined, and between females and males. Females exhibited greater tolerance than males in four of five strains examined. In the comparison between parents and their offspring, stronger influence of female parent than of male parent was observed. Offspring obtained from surviving females exhibited greater tolerance than those from dead females. This tendency was typically observed in male offspring. The survival rate in male offspring obtained from dead female parents was lower than that of those from surviving females. The high-temperature tolerance of male parents did not influence this characteristic in offspring as strongly as that of female parents. These results suggest that the major gene or genes, which has a dominant resistant allele and a recessive sensitive allele, are probably passed on by sex-linked inheritance, located on the X chromosome.

Population structure and conservation genetics of anadromous white-spotted char (Salvelinus leucomaenis) on Hokkaido Island: Detection of isolation-by-distance

To increase the quantity of anadromous and freshwater fish resources and conserve natural populations effectively, it is important to identify conservation units and population structures, as well as to ensure sufficient genetic variability within populations. Here, we used 12 microsatellite loci to evaluate the population structure, effective population size, and bottlenecks in seven anadromous white-spotted char (Salvelinus leucomaenis leucomaenis) populations inhabiting the rivers of Hokkaido Island, Japan. Low migration rates were detected among populations, with significant genetic differentiation being observed, suggesting high homing rates. In addition, isolation-by-distance was observed among the evaluated populations, indicating that the populations in this region are at equilibrium between migration and drift. We identified a genetic bottleneck footprint in all seven of the analyzed white-spotted char populations by using the M ratio test. In contrast, heterozygote excess tests showed that all seven populations showed no signatures of population decline. This discrepancy may have been caused by differences in statistical power among tests. Alternatively, this discrepancy may be consistent with a strong founder effect during the late Pleistocene, followed by a subsequent low migration rate among populations. In conclusion, future conservation genetic management strategies should ensure that anadromous white-spotted char populations successfully exhibit homing behavior in the rivers of Hokkaido Island, Japan.

Isolation and characterization of 13 microsatellite markers for the viviparous surfperch Ditrema temmincki (Embiotocidae) and cross-species amplification

Thirteen microsatellite loci were isolated from a size‐selected genomic library of the surfperch (Ditrema temmincki Bleeker). All loci displayed a high degree of length polymorphism, as observed in the total number of alleles per locus (two to 23) and a high degree of estimated heterozygosity, ranging from 0.080 to 0.893. The primers developed for D. temmincki were also tested for their ability to amplify homologous sequences in D. viride and Neoditrema ransonetii. Distinct differences were observed among three species of surfperches, in both genetic variability and the frequency distribution of the alleles.