Data analyses

The sequences of the two mitochondrial genes were aligned using the ClustalW plugin on Geneious Prime 2020.1.2 (https://www.geneious.com) and prepared as concatenated sequences. DnaSP 6.12.03 [65] was used for the genetic diversity analysis of mitochondrial DNA, in which the number of haplotypes (H), number of segregating sites (S), haplotype diversity (Hd), nucleotide diversity (π), and average number of nucleotide differences (k) were examined.

Pairwise F_ST values were estimated using Arlequin 3.5 software [66] to investigate genetic differentiation among the populations. Principal coordinate analysis (PCoA) was performed with GenAlEx version 6.51b2 [67] based on pairwise F_ST values.

Analyses of molecular variance (AMOVA) were performed using Arlequin 3.5 [66] with the locus-by-locus option and using 1000 permutations to determine the population structure. Specimens were grouped according to regional groups in South Korea: Group 1 comprised specimens from Gyeonggi-do, Group 2, Gangwon-do; Group 3, Chungcheong-do; Group 4, Gyeongsang-do, and Group 5, Jeolla-do.

To better understand the genealogical relationships, the haplotypes were constructed using the TCS method as implemented in PopART 1.7 [68].

To investigate the demographic history of populations, deviations from selective neutrality were tested by Tajima’s D [69] and Fu’s F_S [70] metrics using Arlequin 3.5 [66]. To confirm whether a population had undergone sudden expansion, a mismatch distribution was determined using DnaSP 6.12.03 [65].