Genetic characterization and transmission of the multidrug resistance gene cfr in fecal and environmental pathways on a chicken farm in China

The emergence and spread of the multidrug-resistant gene cfr have raised significant public health concerns worldwide. To investigate its prevalence and dissemination dynamics, 18 cfr-positive strains were isolated in 2021 from fecal and environmental samples. Antimicrobial susceptibility testing showed that all strains were 100 % multidrug-resistant. Conjugation experiments demonstrated that a cfr- carrying IncFII(K)-IncR-IncFIB multi-replicon plasmid could transfer to E. coli J53. S1-nuclease digestion and Southern blotting identified cfr on plasmids of varying sizes, while whole-genome sequencing confirmed its presence on multiple plasmid types: IncX4, IncN, IncFII(K)-IncR-IncFIB, IncFIB-IncFII-IncR-IncHI2-IncHI2A multi-replicon plasmids, and two plasmids of unknown types. Genetic environment analysis revealed that cfr is categorized into five distinct structures (Types I-V). Reverse PCR results showed that Types I, II, and IV can form three circular intermediates of varying lengths (cfr-IS26). Network analysis further indicated strong association between cfr, tet(M), and dfrA14 mediated by IS26. Phylogenetic analysis revealed that the four ST1140 E. coli strains and all nine K. pneumoniae strains showed minimal genetic divergence. These findings suggest both clonal and horizontal transmission of cfr within the poultry farm. Continuous monitoring of cfr in animal-related environments is essential to mitigate its potential transfer to humans.