Detection of AmpC β-lactamases in gram-negative bacteria.

AmpC β-lactamase genes are clinically important because they often confer resistance to most β-lactams other than 4th-generation cephalosporins and carbapenems. However, traditional and existing detection methods are expensive, labor-intensive and range-limited. We established an efficient multiplex PCR method to simultaneously identify six families of C β-lactamase genes, ACC, EBC, CIT, DHA, MOX and FOX, and evaluated the sensitivity and specificity of this assay. The multiplex method could accurately identify ACC, EBC, CIT, DHA, MOX and FOX variants among a total of 175 C β-lactamase genes. The minimum concentration of genomic DNA that could be detected was 1.0×10 copies/μL. We subsequently used this method to analyze 2 spp. with carrying CMY-2 and DHA-1, and 167 isolates in blinded PCR testing. Positive isolates produced bright bands that corresponded with their genotype. Results were in concordance with those of the traditional method but showed increased sensitivity and accuracy. This indicates that the newly developed multiplex PCR system could be used as a diagnostic tool to accurately distinguish the six families of C β-lactamase genes with high efficiency, wide range, easy operation and good discrimination.