Long-term changes in the genetic diversity of the endangered freshwater fish Microphysogobio koreensis (Gobioninae) in Korea

BACKGROUND: Microphysogobio koreensis is an endangered freshwater fish endemic to the Nakdong and Seomjin River systems in Korea. Because its distribution is restricted and population decline has been reported, genetic monitoring is needed to evaluate temporal changes in diversity and to define conservation units. In this study, we developed novel microsatellite markers and assessed long-term changes in genetic diversity, effective population size, and population structure in M. koreensis. RESULTS: We analyzed two populations from the Nakdong and Seomjin Rivers sampled in 2012 and 2021 using seven newly developed microsatellite loci, together with mitochondrial cytochrome b sequences. Across the four population samples, observed heterozygosity ranged from 0.783 to 0.832 and expected heterozygosity ranged from 0.822 to 0.827, indicating that overall heterozygosity was largely maintained over the 9-year interval. However, the number of alleles declined in both river systems in 2021 relative to 2012. Significant bottleneck signals were detected in 12ND, 12SJ, and 21SJ, and effective population size estimates ranged from 19 to 43, except for 12SJ where estimation was not possible. Pairwise genetic differentiation, Bayesian clustering, discriminant analysis of principal components, and haplotype network analysis consistently showed clear genetic separation between the Nakdong and Seomjin River populations. Year-specific STRUCTURE and DAPC analyses further confirmed that this differentiation was consistently maintained within both sampling years. Mitochondrial data also revealed strong differentiation between the two river systems. CONCLUSIONS: Although overall heterozygosity remained relatively stable over time, the loss of alleles and very small effective population sizes indicate ongoing genetic vulnerability in M. koreensis. The Nakdong and Seomjin River populations should be treated as separate management units, and both recent populations require urgent conservation attention. Continued long-term genetic monitoring will be important for guiding conservation management and population reinforcement strategies.