Antimicrobial Susceptibility Testing and Plasmid Conjugation Assay

As recommended by Clinical & Laboratory Standards Institute (CLSI) M100-S30,² CLSI VET01-A4 (see text footnote 2), and the European Committee on Antimicrobial Susceptibility Testing (EUCAST),³ minimum inhibitory concentrations (MICs) of colistin were evaluated by broth microdilution, and 11 antibiotics (meropenem, amikacin, florfenicol, sulfamethoxazole-trimethoprim, piperacillin-tazobactam, tetracycline, ciprofloxacin, gentamicin, tigecycline, cefotaxime, and ceftiofur) were determined with the agar dilution method. E. coli ATCC 25922 was used as a quality control strain. The transferability of mcr-1 was determined by plasmid conjugation assay; streptomycin-resistant E. coli C600 serves as the recipient and MCRPEC as the donor strains. Their transconjugants were selected on MacConkey agar with colistin (2 mg/L) and streptomycin (500 mg/L), and confirmed by ERIC-PCR fingerprinting (Amin et al., 2020) for E. coli C600 and PCR analysis for the mcr-1 gene.