Development of a Method for the Fast Detection of Extended-Spectrum β-Lactamase- and Plasmid-Mediated AmpC β-Lactamase-Producing Escherichia coli and Klebsiella pneumoniae from Dogs and Cats in the USA

Antibiotic resistance, such as resistance to beta-lactams and the development of resistance mechanisms, is associated with multifactorial phenomena and not only with the use of third-generation cephalosporins. Many methods have been recommended for the detection of ESBL and pAmpC beta-lactamase production but they are very subjective and the appropriate facilities are not available in most laboratories, especially not in clinics. Therefore, for fast clinical antimicrobial selection, we need to rapidly detect ESBL- and pAmpC beta-lactamase-producing bacteria using a simple method with samples containing large amounts of bacteria. For the detection of ESBL- and pAmpC phenotypes and genes, the disk diffusion test, DDST and multiplex PCR were conducted. Of the 109 samples, 99 (90.8%) samples were grown in MacConkey broth containing cephalothin, and 71 samples were grown on MacConkey agar containing ceftiofur. Of the 71 samples grown on MacConkey agar containing ceftiofur, 58 Escherichia coli and 19 Klebsiella pneumoniae isolates, in particular, harbored beta-lactamase genes. Of the 38 samples that did not grow in MacConkey broth containing cephalothin or on MacConkey agar containing ceftiofur, 32 isolates were identified as E. coli, and 10 isolates were identified as K. pneumoniae; beta-lactamase genes were not detected in these E. coli and K. pneumoniae isolates. Of the 78 ESBL- and pAmpC beta-lactamase-producing E. coli and K. pneumoniae, 55 (70.5%) isolates carried one or more ESBL genes and 56 (71.8%) isolates carried one or more pAmpC beta-lactamase genes. Our method is a fast, and low-cost tool for the screening of frequently encountered ESBL- and pAmpC beta-lactamase-producing bacteria and it would assist in diagnosis and improve therapeutic treatment in animal hospitals.

Automated summary

A simple and fast method for detecting ESBL- and pAmpC beta-lactamase-producing bacteria in samples with high bacterial load is important for clinical antimicrobial selection. The disk diffusion test, DDST, and multiplex PCR were used for phenotypic and genotypic detection of ESBL and pAmpC. The majority of isolates carried beta-lactamase genes, especially in samples grown on MacConkey agar containing cephalothin or ceftiofur.