Haemadipsa japonica is a land leech species endemic to Japan, where it inhabits litter in temperate evergreen and temperate deciduous forests. Haemadipsa japonica feeds on blood of host animals, such as the sika deer (Cervus nippon) and Japanese serow (Capricornis crispus). Recently, the range of H. japonica has expanded to include areas of anthropogenic activity, possibly because of the increase in mammal populations, which has become a problem in several Japanese prefectures. Knowledge of the origin and host animals of the currently expanding H. japonica populations is critical for countermeasure implementations. Therefore, in this study, we addressed the following questions: what are the origins of H. japonica populations currently expanding from native populations in each prefecture or from neighboring prefectures? How did the expansion of H. japonica populations occur in the case of Tochigi Prefecture, Japan? To address the first question, we assessed the genetic structure of 233 H. japonica leeches collected from 24 populations across Japan. We isolated DNA from caudal sucker tissues, and we sequenced the COI region in mitochondrial DNA (mtDNA). To address the second question, we used 12 nuclear microsatellite (nSSR) markers to genotype 150 individuals from one native and five currently expanding populations in Tochigi prefecture. Furthermore, to identify the host animal of H. japonica in Tochigi prefecture, we isolated DNA from undigested blood clots in the digestive system of 16 H. japonica leeches, and sequenced the 16S ribosome RNA (rRNA) gene in mtDNA; we collated them with nucleotide sequence accessions in GenBank.
Results/Conclusions
Haemadipsa japonica exhibited 43 different haplotypes, phylogenetically divided into two distinct lineages (northern/southern Japan). Each lineage comprised several sublineages, suggesting that the current expansion of H. japonica populations in Japan occurs within each region with each sublineage. In Tochigi Prefecture, the nucleotide diversity of mtDNA of the expanding populations was lower than that of the native population, probably because of the founder effect during the current rapid expansion. Additionally, analyses using nSSR genotypes revealed two distinct, narrow, northern and southern H. japonica groups in Tochigi Prefecture. We identified that the main host animal of H. japonica was Cervus nippon in both groups in Tochigi Prefecture. In conclusion, the current expansion of H. japonica populations in Japan might be caused within each narrow region; in Tochigi Prefecture, in particular, this expansion was caused via C. nippon within each northern and southern region.