Abstract
Leeches hold significant medical and pharmaceutical value for antithrombotic treatments, yet their genetic diversity patterns remain poorly understood. We performed population genetic analyses on seven Hirudinaria manillensis populations from southern China using mitochondrial protein-coding genes (MitPCGs). Complete sequences of all 13 MitPCGs were obtained from 74 individuals. Haplotype diversity exhibited a logarithmic relationship with the gene length (R(2) = 0.858, p < 0.001), while nucleotide diversity showed a near-perfect alternating low-high pattern (Z = 2.938, p = 0.003). Concatenated sequence analyses indicated high haplotype diversity (>0.5) and low nucleotide diversity (<0.005) across all populations, suggesting a historical bottleneck followed by rapid expansion and mutation accumulation. The haplotype network, haplotype phylogenetics, and genetic structure analyses revealed moderate genetic differentiation across populations, dividing them into three clades: a basal Yunnan population (YNHH), sub-basal Guangxi populations (GXGG, GXLZ, and GXYL), and distal Guangdong/Hainan populations (GDMM, GDZJ, and HNDA). Analysis of historical population demography revealed five phases from ancient to recent times (P1-5): growth, prolonged stability, rapid decline, rapid growth, and secondary decline. These phases correlate strongly with past climatic events, demonstrating that glacial-interglacial cycles profoundly impacted the leech's effective population size. This study provides a key scientific basis for H. manillensis resource conservation and utilization.