Genome Analysis and Multiplex PCR Method for the Molecular Detection of Coresistance to Cephalosporins and Fosfomycin in Serovar Heidelberg.

Heidelberg is among the top three serovars associated with human foodborne illness in Canada. Traditional culture and antimicrobial susceptibility testing techniques can be time-consuming to identify Heidelberg resistant to cephalosporins and fosfomycin. Rapid and accurate detection of such antibiotic-resistant Heidelberg isolates is essential to adopt appropriate control measures. In this study, 15 Heidelberg strains isolated from feces of Canadian broiler chickens were characterized by whole genome sequencing. Heidelberg genomes had an average coverage of greater than 80-fold, an average of 4,761 protein-coding genes, and all belonged to multilocus sequence type ST15. Genome sequences were compared with genomes in the National Center for Biotechnology Information Pathogen Detection database ( www.ncbi.nlm.nih.gov/pathogens/ ), including human outbreak isolates. The Canadian broiler isolates clustered with chicken isolates from the United States and an equine clinical isolate from Ontario, Canada. In agreement with their antimicrobial resistance phenotypes, several chromosomally encoded specific antimicrobial resistance genes including and multidrug resistance efflux pump determinants were detected. An AmpC-like β-lactamase gene, , linked with a quaternary ammonium compound resistance gene, on a replicon type 1 plasmid was detected in all 15 broiler Heidelberg isolates. Of the 205,031 published genomes screened in silico, 4,954 (2.4%) contained , 8,143 (4.0%) contained and 919 (0.4%) contained both resistance genes. The combination of both resistance genes ( and ) was detected in 64% of the Heidelberg genomes and in a small proportion of various other serovars. A PCR method was developed to detect Heidelberg in pure culture and chicken feces based on specific primers targeting genes conferring fosfomycin () and third-generation cephalosporin () resistance as well as the -specific gene and the universal 16S rRNA genes. The PCR assay was specific and sensitive for and harboring Heidelberg. However, some other serovars containing these two resistance genes could also be detected by the developed PCR method.