Molecular epidemiology of third-generation cephalosporin-resistant Enterobacteriaceae in Southeast Queensland, Australia.

Third-generation cephalosporin-resistant (3GC-R) Enterobacteriaceae represent a major threat to human health. Here, we captured 288 3GC-R Enterobacteriaceae clinical isolates from 264 patients presenting at a regional Australian hospital over a 14-month period. Alongside routine mass spectrometry speciation and antibiotic sensitivity testing, isolates were examined using rapid (∼40 min) real-time PCR assays targeting the most common extended spectrum β-lactamases (ESBLs; blaCTX-M-1 and blaCTX-M-9 groups, plus blaTEM, blaSHV, and an internal 16S ribosomal DNA control). AmpC CMY β-lactamase (blaCMY) prevalence was also examined. Escherichia coli (80.2%) and Klebsiella pneumoniae (17.0%) were dominant, with Klebsiella oxytoca, Klebsiella aerogenes and Enterobacter cloacae infrequently identified. Ceftriaxone and cefoxitin resistance were identified in 97.0% and 24.5% of E. coli and K. pneumoniae isolates, respectively. Consistent with global findings in Enterobacteriaceae, most (98.3%) isolates harbored at least one β-lactamase gene, with 144 (50%) encoding blaCTX-M-1 group, 92 (31.9%) blaCTX-M-9 group, 48 (16.7%) blaSHV, 133 (46.2%) blaTEM, and 34 (11.8%) blaCMY A subset of isolates (n=98) were subjected to whole-genome sequencing (WGS) to identify the presence of cryptic resistance determinants, and to verify genotyping accuracy. WGS of β-lactamase negative or carbapenem-resistant isolates identified uncommon ESBLs and carbapenemases, including blaNDM and blaIMP, and confirmed all PCR-positive genotypes. We demonstrate that our PCR assays enable the rapid and cost-effective identification of ESBLs in the hospital setting, which has important infection control and therapeutic implications.