Mitochondrial DNA sequence divergence between two cryptic species of Lethenteron, with reference to an improved identification technique

Genetic divergences and population structures were examined in the cryptic Lethenteron sp. N and sp. S, based on mitochondrial DNA (mtDNA) cytochrome oxidase subunit I (CO I) region sequences. An improved method of discrimination between L. sp. N and sp. S was found using PCR, with diagnostic primers for each species-specific sequence in the mtDNA CO I region. Identification of 50 individuals of each species by this analysis was consistent with that by allozyme analysis of nuclear DNA. L. sp. N and sp. S, identified on the basis of diagnostic alleles at five allozymic loci, were independently grouped in a neighbour-joining (NJ) tree, with a large sequence difference (mean ± s.d. = 9·10 ± 0·36%) between them. Within each species, the values of sequence divergence among localities were significantly higher in L. sp. S (1·61 ± 0·44%) than in L. sp. N (1·10 ± 0·48%). On the tree and nested clade analyses, several phylogenetic groups comprising geographically close localities were detected in the former, although scarcely detected in the latter, probably resulting from dispersal pattern differences between them.