Piperacillin/tazobactam resistance in a clinical isolate of Escherichia coli due to IS26-mediated amplification of blaTEM-1B

Alasdair T M Hubbard,Alice J Fraser,Jenifer Mason,Caroline Corless,Jon Van Aartsen,Alex Howard,Christopher M Parry,Christopher M Parry,Christopher M Parry,Christopher M Parry,Issra Bulgasim,Adam P Roberts,Thomas Edwards,Paul Roberts,Emily R Adams

doi:10.1038/s41467-020-18668-2

Abstract

A phenotype of Escherichia coli and Klebsiella pneumoniae, resistant to piperacillin/tazobactam (TZP) but susceptible to carbapenems and 3rd generation cephalosporins, has emerged. The resistance mechanism associated with this phenotype has been identified as hyperproduction of the β-lactamase TEM. However, the mechanism of hyperproduction due to gene amplification is not well understood. Here, we report a mechanism of gene amplification due to a translocatable unit (TU) excising from an IS26-flanked pseudo-compound transposon, PTn6762, which harbours blaTEM-1B. The TU re-inserts into the chromosome adjacent to IS26 and forms a tandem array of TUs, which increases the copy number of blaTEM-1B, leading to TEM-1B hyperproduction and TZP resistance. Despite a significant increase in blaTEM-1B copy number, the TZP-resistant isolate does not incur a fitness cost compared to the TZP-susceptible ancestor. This mechanism of amplification of blaTEM-1B is an important consideration when using genomic data to predict susceptibility to TZP.

Highlights

A phenotype of Escherichia coli and Klebsiella pneumoniae, resistant to piperacillin/tazobactam (TZP) but susceptible to carbapenems and 3rd generation cephalosporins, has emerged
Putative clonality of these two isolates was confirmed with whole-genome sequencing; both isolates were identified as serotype H30 O86, sequence type 315 and had an average nucleotide identify (ANI) of 100%, with 36 single nucleotide polymorphisms (SNP) difference between the two isolates
Tazobactam is able to inhibit the activity of class A β-lactamases[32], and the presence of blaTEM-1 within the genome of an E. coli isolate should not result in resistance to TZP

Summary

Introduction

A phenotype of Escherichia coli and Klebsiella pneumoniae, resistant to piperacillin/tazobactam (TZP) but susceptible to carbapenems and 3rd generation cephalosporins, has emerged. Mechanisms leading to hyperproduction include mutations in the promoter region of blaTEM, changing it from a weak promoter (P3) to a stronger promoter (P4 or P5)[18] or a single point mutation further upstream resulting in the overlapping, stronger promoter Pa/Pb superseding the weaker P3 promoter[19], increasing the production of TEM Another such mechanism proposed to cause TZP-resistance but 3rd generation cephalosporin and carbapenem susceptibility is the increase in copy number of blaTEM present in either a plasmid or chromosome[17,20]. We identify a pair of clonal E. coli isolates, isolated from a single patient across two separate infection episodes, which display within-patient evolution to TZP resistance In this isolate, amplification of blaTEM-1B occurs when a TU excises from a pseudocompound transposon flanked by directly repeated IS26 present in the chromosome. Replicating the evolutionary event in vitro in the TZP-susceptible isolate leads to the capture of the TU in a plasmid which contains a copy of IS26

Methods

Results

Conclusion