Establishment of a PCR Detection Method for the Anti-colistin Gene mcr-3

Abstract

With the widespread use of antibiotics, bacterial resistance has become a growing concern, particularly the emergence of plasmid-mediated colistin resistance genes, which diminishes the efficacy of colistin, a "last-resort" antibiotic. This study designed and established a specific PCR detection method for the mcr-3 gene to support rapid screening and resistance monitoring. A pair of primers was designed targeting a unique region of the mcr-3 gene, located between the middle variable region and the catalytic active domain (1032-1287 bp), yielding a theoretical amplification product of 209 bp. The primers exhibited ideal physicochemical properties, with moderate GC content (55% and 60%) and similar Tm values (approximately 60 ℃). Experimental validation demonstrated high specificity, with only mcr-3-positive templates producing a 209 bp band, while negative controls showed no amplification. Sensitivity tests indicated that 1ng, 100 pg, and 10pg of mcr-3 plasmid DNA could be detected, with band intensity decreasing with lower concentrations. Testing of 16 Salmonella strains (primarily from poultry, with a few from human sources) revealed no mcr-3 gene, suggesting its rarity in the tested samples. This PCR method is simple, rapid, and suitable for screening mcr-3 in livestock, human, and environmental samples, providing critical technical support for antimicrobial resistance monitoring and control strategies.