Winter migratory birds may carry diverse antimicrobial resistance genes into Japan

BACKGROUND: Environmental surveillance of antimicrobial resistance (AMR) in wildlife remains limited, despite increasing recognition that resistance determinants can circulate across human, livestock, and natural ecosystems. Migratory waterbirds move long distances and aggregate at shared stopover and wintering sites, potentially facilitating the acquisition and redistribution of antimicrobial resistance genes (ARGs) across regions. However, nationwide evidence describing the breadth of ARGs carried by winter migratory birds in Japan is scarce. We assessed the diversity and distribution of ARGs in pooled fecal samples from winter migratory birds across Japan. METHODS: We analyzed pooled fecal DNA collected at migratory bird habitats across 12 local governments during the 2021-2022 and 2022-2023 winter seasons (24 pools). Avian host origin was inferred by DNA metabarcoding, and ARGs were profiled by probe-based target enrichment with read-based detection (ARG detected at ≥ 10 reads). RESULTS: Ducks (Anas spp. and Mareca spp.) were the predominant inferred hosts. ARGs were detected in all areas and included genes associated with resistance to multiple antibiotic classes used in livestock production. Across the two seasons, genes associated with resistance to gentamicin, cephalosporins, macrolides, tetracyclines, fosfomycin, clindamycin, penicillins, streptogramins, sulfonamides/trimethoprim, colistin, erythromycin, chloramphenicol, rifampicin, and isoniazid were detected in all 12 areas in at least one season. Genes associated with resistance to agents restricted for use in Japanese livestock production, including colistin, erythromycin, chloramphenicol, and rifampicin, were also detected in all 24 pools. Isoniazid-, gentamicin-, meropenem-, and tigecycline-associated genes were detected in 23/24, 20/24, 11/24, and 9/24 pools, respectively. CONCLUSIONS: These data indicate widespread environmental occurrence of diverse ARGs and support the possibility that migratory birds could contribute to long-distance dispersal of ARGs. Culture-based isolation, phenotypic testing, and quantitative analyses will be needed to identify host bacteria and assess clinical relevance.