Molecular Epidemiology of Third-Generation-Cephalosporin-Resistant Enterobacteriaceae in Southeast Queensland,...

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. In addition to routine mass spectrometry and antibiotic sensitivity testing, isolates were examined using rapid (∼40-min) real-time PCR assays targeting the most common extended-spectrum β-lactamases (ESBLs; bla_CTX-M-1 and bla_CTX-M-9 groups, plus bla_TEM, bla_SHV, and an internal 16S rRNA gene control). AmpC CMY β-lactamase (bla_CMY) 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%) harboring bla_CTX-M-1 group, 92 (31.9%) harboring bla_CTX-M-9 group, 48 (16.7%) harboring bla_SHV, 133 (46.2%) harboring bla_TEM, and 34 (11.8%) harboring bla_CMY genes. 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 ESBL and carbapenemase genes, including bla_NDM and bla_IMP, 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.