Diminished Susceptibility to Cefoperazone/Sulbactam and Piperacillin/Tazobactam in Enterobacteriaceae Due to Narrow-Spectrum β-Lactamases as Well as Omp Mutation

Abstract

Cefoperazone/sulbactam (CSL) and piperacillin/tazobactam (TZP) are commonly used in clinical practice in China because of their excellent antimicrobial activity. CSL and TZP-nonsusceptible Enterobacteriaceae are typically resistant to extended-spectrum cephalosporins such as ceftriaxone (CRO). However, 11 nonrepetitive Enterobacteriaceae strains, which were resistant to CSL and TZP yet susceptible to CRO, were collected from January to December 2020. Antibiotic susceptibility tests and whole-genome sequencing were conducted to elucidate the mechanism for this rare phenotype. Antibiotic susceptibility tests showed that all isolates were amoxicillin/clavulanic-acid resistant and sensitive to ceftazidime, cefepime, cefepime/tazobactam, cefepime/zidebactam, ceftazidime/avibactam, and ceftolozane/tazobactam. Whole-genome sequencing revealed three of seven Klebsiella pneumoniae strains harbored blaSHV-1 only, and four harbored blaSHV-1 and blaTEM-1B. Two Escherichia coli strains carried blaTEM-1B only, while two Klebsiella oxytoca isolates harbored blaOXY-1-3 and blaOXY-1-1, respectively. No mutation in the β-lactamase gene and promoter sequence was found. Outer membrane protein (Omp) gene detection revealed that numerous missense mutations of OmpK36 and OmpK37 were found in all strains of K. pneumoniae. Numerous missense mutations of OmpK36 and OmpK35 and OmpK37 deficiency were found in one K. oxytoca strain, and no OmpK gene was found in the other. No Omp mutations were found in E. coli isolates. These results indicated that narrow spectrum β-lactamases, TEM-1, SHV-1, and OXY-1, alone or in combination with Omp mutation, contributed to the resistance to CSL and TZP in CRO-susceptible Enterobacteriaceae.Antibiotic susceptibility testsAntibioticsBreakpoint, (μg/ml)Klebsiella pneumoniaeEscherichia couKlebriehd axyocaE1E3E4E7E9E10E11E6E8E2E5CRO≤1≥4≤0.5≤0.5≤0.5≤0.5 1≤0.51≤0.5≤0.511CAZ4 ≥161214444241 1FEP≤2 216 110.2512220.521 1AMC≤8 ≥32≥128≥128≥128≥128≥128≥128≥128≥128≥128≥128≥128CSL≤16 ≥6464646464≥128128≥12864128128≥128TZP≤16 ≥128≥256≥256≥256≥25622562256≥256≥256≥256≥256≥256FPT≤2 ≥1610.50.060.1252120.2510.1250.25FPZ≤2 2160.250.250.060.1250.250.25 10.1250.250.1250.125CZA≤8 216 10.50.250.2510.2510.50.50.50.25CZT≤2 28210.5 1222 1122CROceftriaxone, CAZceftazidime, FEPcefepime, AMC:amoxicillin clavulanic-acid, CSLcefoperazone/sulbactam, TZP:piperadllin/tazobactam, FPT:cefepime tazobactam, FPZ:cefepime/zidebactam, CZA:ceftazidime/avibactam, CZTceftolozane/tazobactam Gene sequencing resultsNumberStrainSTp-Lactamase genePromoter sequence mutationOmp mutationElKpn45blaSHV-1, blaTEM-lBnoneOmpK36, OmpK3 7E3Kpn45blaSHV-1, blaTEM-lBnoneOmpK36. OmpK3 7E4Kpn2854blaSHV-1noneOmpK36, OmpK3 7E7Kpn2358blaSHV-1 - blaTEM-lBnoneOmpK36, OmpK3 7E9Kpn2358blaSHV-1. blaTEM-lBnoneOmpK36. OmpK3 7E10Kpn 189blaSHV-1noneOmpK36. OmpK3 7EllKpn45blaSHV-1noneOmpK36, OmpK3 7E6Eco88blaTEM-lBnonenoneESEco409blaTEM-1BnonenoneE2Kox194blaOXY-1-3noneOmpK36 mutations. OmpK35 and OmpK 37 deficiencyE5Kox 11blaOXY-1-1noneno OmpK (OmpK3 5, OmpK36 and OmpK37) gene found