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TEM-1, mediated by plasmid and transposon, is the most commonly encountered β-lactamase in Gram-negative bacteria. Four different promoters upstream of  bla  TEM -related genes have been identified: the weak  P3  promoter, and the strong promoters  Pa/Pb ,  P4 , and  P5.  In this study, we investigated the genetic basis of a clinical strain of  Escherichia coli  (RJ904), which was found to be resistant to BLBLIs (β-lactam/β-lactamase inhibitors), including amoxicillin-clavulanate, ticarcillin-clavulanate (TCC), and piperacillin-tazobactam (TZP) but sensitive to third-generation cephalosporins. The conjugation test and S1-nuclease pulsed-field gel electrophoresis (S1-PFGE) demonstrated that transfer of this resistance was mediated by a ca. 100 kb plasmid. The transformant with TZP resistance was screened out with the shortgun cloning. Sequence analysis revealed that the recombinant plasmid contained a  bla  TEM-1b  gene with the strong promoter  Pa/Pb . Different plasmids were cloned based on the clone vector pACYC184 with the insertion of the  bla  TEM-1b  gene with promoters  Pa/Pb  or  P3 . Susceptibility to TZP was determined by the  E -test, agar dilution, and broth microdilution. The level of  bla  TEM-1b -specific transcription was determined by quantitative real-time PCR. Substitution of  Pa/Pb  for  P3  resulted in a 128-fold decline of the MIC value of TZP, from &gt;1024 mg/L to 8 mg/L, and a significantly lower  bla  TEM-1b  expression level. Hyperproduction of TEM-1 β-lactamase mediated by the promoter  Pa/Pb  was responsible for high resistance to TZP in  E. coli . Our data show possible risks of resistance development in association with the clinical use of TZP. The  bla  TEM  promoter modifications should be considered for whole genome whole-genome sequencing-inferred bacterial antimicrobial susceptibility testing.

Piperacillin-Tazobactam (TZP) Resistance in Escherichia coli Due to Hyperproduction of TEM-1 β-Lactamase Mediated by the Promoter Pa/Pb