Evolution of TEM-type extended-spectrum β-lactamases in Escherichia coli by cephalosporins

J Glob Antimicrob Resist. 2019 Dec:19:32-39. doi: 10.1016/j.jgar.2019.03.010. Epub 2019 Apr 30.

Abstract

Objectives: This study was conducted to examine the molecular mechanisms responsible for the evolution of TEM-type extended-spectrum β-lactamases (ESBLs) following selective pressure from four third-generation cephalosporins, namely ceftazidime, cefotaxime, ceftriaxone and ceftibuten. In addition, selective enrichment for ESBL detection in environmental samples was investigated.

Methods: Using experimental evolution, resistant variants were isolated and mutations in TEM-1 were examined by DNA sequencing. Resistance levels and the development of cross-resistance were determined for ESBL-producing isolates by Etest and disk diffusion assay. Selective plating with or without prior growth in selective broth was used to examine the approach of selective enrichment for ESBL detection.

Results: The third-generation cephalosporins ceftazidime, cefotaxime and ceftriaxone selected for ESBLs, whereas ceftibuten did not. All ESBL variants additionally remained susceptible to ceftibuten. DNA sequencing of the TEM-1 coding sequence of mutants revealed mutations not previously isolated through selection. This indicates that the potential for ESBL evolution is much broader than can be inferred from sequence analysis of clinical samples alone. The results also indicate that selective enrichment for enhanced detection of ESBL-producers may give unreliable results owing to the selection of spontaneous mutations in narrow-spectrum β-lactamases resulting in TEM-type ESBL-producers.

Conclusion: These results help explain the molecular changes responsible for evolution of TEM-type ESBLs and meanwhile question the appropriate use of selective enrichment for detection of ESBLs in environmental samples.

Keywords: Antimicrobial resistance; Escherichia coli; Evolution; Extended-spectrum β-lactamase; TEM-1; Third-generation cephalosporin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / pharmacology*
  • Cefotaxime / pharmacology
  • Ceftazidime / pharmacology
  • Ceftriaxone / pharmacology
  • Cephalosporins / pharmacology*
  • Drug Resistance, Multiple, Bacterial / genetics
  • Escherichia coli / drug effects*
  • Escherichia coli / enzymology*
  • Escherichia coli / genetics
  • Evolution, Molecular
  • Humans
  • Microbial Sensitivity Tests
  • Mutation
  • Sequence Analysis, DNA
  • beta-Lactamases / genetics
  • beta-Lactamases / metabolism*

Substances

  • Anti-Bacterial Agents
  • Cephalosporins
  • Ceftriaxone
  • Ceftazidime
  • beta-Lactamases
  • beta-lactamase TEM-1
  • Cefotaxime