AUTHOR=Ma Ke , Zong Zhiyong 

TITLE=Resistance to aztreonam-avibactam due to CTX-M-15 in the presence of penicillin-binding protein 3 with extra amino acids in Escherichia coli

JOURNAL=Frontiers in Microbiology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2022.1047109

DOI=10.3389/fmicb.2022.1047109

ISSN=1664-302X

ABSTRACT=<p>Aztreonam-avibactam is a promising combination to treat carbapenem-resistant Enterobacterales including coverage for metallo-β-lactamases. <italic>Escherichia coli</italic> strains resistant to aztreonam-avibactam have emerged but resistance mechanisms remain to be elucidated. We performed a study to investigate the mechanism for aztreonam-avibactam in a carbapenem-resistant <italic>Escherichia coli</italic> clinical strain. This strain was resistant to aztreonam-avibactam (aztreonam MIC, 16 mg/L in the presence of 4 mg/L avibactam). Whole genome sequencing revealed that the strain carried metallo-β-lactamase gene <italic>bla</italic><sub>NDM-4</sub> and the extended-spectrum β-lactamase (ESBL) gene <italic>bla</italic><sub>CTX-M-15</sub> and had a YRIK four amino acid insertion in penicillin-binding protein 3 (PBP3). <italic>bla</italic><sub>CTX-M-15</sub> was cloned into pET-28a(+), followed by the transformation, with the gene, of <italic>E. coli</italic> strain 035125∆pCMY42 possessing the YRIK insertion in PBP3 and strain BL21 with the wildtype PBP3. <italic>bla</italic><sub>CTX-M-14</sub>, another common ESBL gene, and <italic>bla</italic><sub>CTX-M-199</sub>, a hybrid of <italic>bla</italic><sub>CTX-M-14</sub> and <italic>bla</italic><sub>CTX-M-15</sub> were also individually cloned into both <italic>E. coli</italic> strains for comparison. Aztreonam-avibactam resistance was only observed in the <italic>E. coli</italic> strains with the YRIK insertion in PBP3 that produced CTX-M-15 or its hybrid enzyme CTX-M-199. Checkerboard titration assays were performed to determine the synergistic effects between aztreonam-avibactam and ceftazidime or meropenem. Doubling avibactam concentration <italic>in vitro</italic> reversed aztreonam-avibactam resistance, while the combination of aztreonam-avibactam and ceftazidime or meropenem did not. In conclusion, CTX-M enzymes with activity against aztreonam, (e.g., CTX-M-15 and CTX-M-199), can confer resistance in the combination of PBP3 with YRIK insertions in metallo-β-lactamase-producing carbapenem-resistant <italic>E. coli</italic>. Doubling the concentration of avibactam may overcome such resistance.</p>