Isolation and characterization of novel microsatellite loci in the wrinkled frog.

The wrinkled frog, Rana rugosa, is a species complex that inhabits plains and mountains near freshwater bodies throughout East Asia, encompassing China, Korea, Japan, and the Russian Primorye region. Although extensive efforts are required to estimate cryptic diversity in the R. rugosa complex, no specifically designed microsatellite loci are available. Here, novel and polymorphic microsatellites were isolated based on the construction of a microsatellite-enriched library and characterized using R. rugosa specimens collected on the Korean Peninsula. A total of 72 primer sets were designed from approximately 400 positive clones, and 22 were validated as being reliably amplified and polymorphic. Overall, high genetic variability was observed (mean number of alleles per locus = 22.23; mean observed and expected heterozygosities = 0.770 and 0.816, respectively) from a total of 60 individuals sampled from two geographically isolated localities. In the two sites analyzed, an extremely low level of relatedness was inferred from the estimation of pairwise relatedness, and no evidence of a genetic bottleneck was detected. The two sites showed a high level of genetic differentiation, suggesting a clear signature of isolation following colonization. With high statistical power in parentage and sibship exclusion, these microsatellite loci will be suitable for the identification of cryptic diversity and population structure as well as the recognition of individuals in social interaction and captive breeding practice.

Development of polymorphic microsatellite loci in Odontobutis obscura using Illumina paired-end sequencing and the test of cross-species amplification.

Odontobutis obscura is a bottom-dwelling freshwater fish native to East Asia. Its range encompasses southwest China, western Japan, and Geoje Island in South Korea. Despite its widespread range in China and Japan, only a small and spatially isolated population is found in South Korea. We developed a total of 23 novel and polymorphic microsatellite loci of O. obscura using Illumina paired-end shotgun sequencing and characterized them using 80 Japanese and Korean samples. An extensive genetic polymorphism was detected at these 23 loci, with the observed number of alleles at a locus ranging from 2 to 15 and expected and observed heterozygosities ranging from 0 to 0.656 and 0 to 0.547, respectively. Korean O. obscura exhibited a much lower level of genetic variability than the Japanese population did, probably as a result of long-term isolation combined with historical bottlenecks. The Japanese and Korean populations showed a high level of genetic differentiation with FST = 0.700 and RST = 0.913. Many of our primer sets were successfully transferable to congeneric O. interrupta and O. platycephala, which exhibited even greater polymorphism than Korean O. obscura. In conclusion, our study showed that these 23 microsatellite markers are useful for understanding the conservation biology and population genetic structure of O. obscura and other congeneric species.

Microsatellite genetic differentiation among populations of the Trinidadian guppy.

Insight into the processes of evolutionary change can be obtained by studying the distribution of genetic diversity among populations. Such diversity can be shaped by historical colonization events, population connectivity and adaptation to local selection pressures. Here we examine genetic differentiation of Trinidadian guppies, Poecilia reticulata, by genotyping 373 individuals from 15 populations located in three drainages (northern coast, Caroni and Oropouche) with 7 microsatellite loci. Our data provide little evidence to support previous claims of two major genetic lineages of guppies in northern Trinidad but instead suggest a more complex pattern of gene flow among populations from different drainages. First, some of the populations in the Caroni drainage show genetic signatures similar to those in the Oropouche drainage. Second, the populations in the northern coast are all highly differentiated from those in either the Caroni or Oropouche drainages. Despite differing selection regimes owing to predation pressure, populations from upstream and downstream locales typically cluster together, albeit upstream populations consistently have less genetic variability than the corresponding downstream population. There is, however, no overall pattern of isolation by distance. We also find evidence that an artificially transplanted population from the Caroni drainage is successfully invading into other populations within the Oropouche system. Our analysis details the genetic and phylogeographic structure of Trinidadian guppies in the northern range and provides insight into evolutionary processes at different timescales that have shaped genetic heterogeneity in this fish.