An Unusual 'Gift' from Humans: Third-Generation Cephalosporin-Resistant Enterobacterales in migratory birds along the East Asian-Australasian Flyway.

Migratory birds play a pivotal role in the global dissemination of antimicrobial resistance genes (ARGs), with shorebirds relying on coastal wetlands during their long-distance migrations, environments often contaminated and conducive to ARG transmission. However, systematic investigations into antimicrobial resistance (AMR) in shorebirds remain scarce. During spring and autumn of 2023, we collected 893 throat and cloacal swabs from 480 shorebirds, representing 28 species, at Chongming Dongtan, a critical stopover along the East Asian-Australasian Flyway. Our analysis identified six strains, including four extra-intestinal pathogenic E. coli (ExPEC) and two K. pneumoniae, that exhibited resistance to third-generation cephalosporins, with three ExPEC strains exhibiting significant virulence in Galleria mellonella infection assays. We identified two conjugative plasmids: E042113F_p1, carrying the blaCMY-2 gene in E. coli, and M50_p2, carrying the blaKPC-2 gene in a hypervirulent K. pneumoniae with a virulence plasmid harboring the aerobactin system. Bioinformatic and experimental analyses confirmed that these plasmids could transfer without any fitness cost, remaining stable for at least 30 passages. Surprisingly, genomic tracing revealed that among the plasmids similar to E042113F_p1 (blaCMY-2), the earliest was identified in a Chinese swallow in 2015, with subsequent detections in wild birds from Mongolia (2017), Russia (2018), and Australia (2019). Notably, these E04-CMY-like/M50-KPC-like plasmids predominantly originated from human sources, underscoring the pivotal role of human activity in the cross-species transmission of AMR. This human-mediated transmission of resistance elements into wildlife posed a substantial risk for amplifying and disseminating AMR through long-range migratory bird movements, highlighting the urgent need for international collaboration under a One Health framework. Integrated surveillance, environmental management, and stringent antibiotic stewardship are critical to mitigating the risks posed by migratory birds in amplifying and spreading AMR across ecosystems.