Improved molecular serogroup typing reveals locally circulating Leptospira serogroups in wildlife in Yamaguchi, Japan

Leptospirosis is a globally distributed zoonotic disease caused by pathogenic Leptospira spp., which are classified into numerous serogroups. Accurate serogroup identification is essential for epidemiological understanding and public health interventions in endemic regions. The microscopic agglutination test (MAT) is the conventional serological method for serogroup identification; however, it is labor-intensive and depends on extensive reference strain panels. Molecular serogroup typing (MST) has been proposed as an alternative approach, but its performance for region-specific strains has not been sufficiently evaluated. In this study, we improved an MST method optimized for Japanese Leptospira isolates and applied it to determine the serogroups of pathogenic Leptospira spp. detected in kidneys of wild boars (Sus scrofa) and sika deer (Cervus nippon) collected in Yamaguchi Prefecture, Japan, between 2016 and 2025. Pathogenic Leptospira DNA was detected in 23 of the 190 wild boars (12.1%) and 22 of the 275 sika deer (8.0%) by real-time PCR targeting lipL32. The improved MST identified serogroups Autumnalis, Australis, Canicola, and Hebdomadis in both host species, with Autumnalis predominating. In contrast, MAT using standard reference strains detected antibodies mainly against Hebdomadis and Australis, indicating discrepancies in the predominant serogroups between molecular and serological results. Notably, MAT using a local Autumnalis strain successfully detected Autumnalis antibodies in both hosts. These findings demonstrate that the improved MST complements serological testing.