Unusual Escherichia coli PBP 3 Insertion Sequence Identified from a Collection of Carbapenem-Resistant Enterobacteriaceae Tested In Vitro with a Combination of Ceftazidime-, Ceftaroline-, or Aztreonam-Avibactam

Abstract

Carbapenemase-producing Enterobacteriaceae isolates (n ϭ 110) from health care centers in central Indiana (from 2010 to 2013) were tested for susceptibility to combinations of avibactam (4 g/ml) with ceftazidime, ceftaroline, or aztreonam. MIC 50 /MIC 90 values were 1/2 g/ml (ceftazidime-avibactam), 0.5/2 g/ml (ceftaroline-avibactam), and 0.25/0.5 g/ml (aztreonam-avibactam.) A ␤-lactam MIC of 8 g/ml was reported for the three combinations against one Escherichia coli isolate with an unusual TIPY insertion following Tyr344 in penicillin-binding protein 3 (PBP 3) as the result of gene duplication.KEYWORDS carbapenem-resistant, CRE, avibactam, ceftazidime, ceftaroline, aztreonam, carbapenemase, resistance, PBP 3 C arbapenem-resistant Enterobacteriaceae (CRE) have been designated one of the most urgent antibiotic resistance threats by multiple global health organizations (1, 2). Carbapenemase-producing CRE are most worrisome, due primarily to plasmidencoded KPC, SME, and OXA families with serines at their active sites and the NDM and VIM metallo-␤-lactamase (MBL) families (3). The recently approved ␤-lactamase inhibitor combination ceftazidime-avibactam provides the potential for treating many CRE infections caused by pathogens with serine carbapenemases that are inhibited by avibactam (4). Although avibactam does not inhibit MBLs, its combination with aztreonam protects the monobactam against hydrolysis by serine enzymes, while aztreonam remains functionally active due to its stability to MBLs (5). In addition, a ceftarolineavibactam combination potentially could allow for the treatment of mixed infections due to methicillin-resistant Staphylococcus aureus and CRE (6). In this study, the three avibactam combinations were tested against 110 recent multidrug-resistant CRE isolates with well-characterized ␤-lactamase profiles (7) to determine whether there was a reservoir of contemporary isolates resistant to any of the combinations.Avibactam was furnished by Allergan; all other antibacterial agents were obtained from the U.S. Pharmacopeial Convention (USP, Rockville, MD). Consecutive carbapenemaseproducing CRE isolates from 172 discrete patients (from 2010 to 2013) were provided by a central clinical microbiology laboratory that serviced 14 Indiana health care centers and two large urban hospitals (5, 7). After a Vitek 2 system identified carbapenemresistant isolates, those isolates with positive results in the modified Hodge test (MHT) (8) were tested using the Carba NP test (9). Two SME-producing Serratia marcescens isolates from a second laboratory (2011) were included. Based on bacterial species, susceptibility profiles, and ␤-lactamase composition, 110 representative isolates were

In Vitro Activity of Ceftazidime-Avibactam against Clinical Isolates of Enterobacteriaceae and Pseudomonas aeruginosa Collected in Latin American Countries: Results from the INFORM Global Surveillance Program, 2012 to 2015

“…The mechanism(s) of reduced susceptibility remain to be determined for these isolates but may reflect the presence of an avibactam-insensitive ␤-lactamase that was not detected using the current molecular algorithm (20,21) or a combination of mechanisms, such as increased KPC production with porin deficiency and altered efflux (22)(23)(24). Sequence insertions in penicillinbinding protein 3 have also been reported to result in reduced susceptibility to avibactam-cephalosporin combinations, although ceftazidime-avibactam remained active (MIC Յ8 g/ml) against the isolates reported to date (25,26). Upregulation of efflux transport systems or porin loss alone were not implicated in reduced susceptibility to ceftazidime-avibactam in a previous direct test of those mechanisms (27).…”

In Vitro Activity of Ceftazidime-Avibactam and Aztreonam-Avibactam against OXA-48-Carrying Enterobacteriaceae Isolated as Part of the International Network for Optimal Resistance Monitoring (INFORM) Global Surveillance Program from 2012 to 2015

“…The insertion of YRIN or YRIK after position 333, near the ␤-lactam binding pocket of E. coli PBP3, was reported to decrease susceptibility to ceftazidime by 4-to 8-fold and to aztreonam by 32-fold and resulted in MIC values for aztreonam-avibactam and ceftazidime-avibactam of 4 to 16 g/ml and 2 to 8 g/ml, respectively (65). The insertion of a different four-residue sequence (TIPY inserted after position 344) into the PBP3 of E. coli was proposed to account for MIC values of 8 g/ml observed for ceftaroline-avibactam, ceftazidime-avibactam, and aztreonam-avibactam in a different study (66). No major disruptions in the sequence of pbp3 were identified in the K. pneumoniae, E. cloacae, and S. marcescens isolates sequenced as part of this study, which were assumed to harbor different resistance mechanisms.…”

Stepwise evolution and convergent recombination underlie the global dissemination of carbapenemase-producingEscherichia coli

“…2a). To our knowledge these modifications are the only mutations contributing to β-lactams reported in natural E. coli isolates 19,25 . Nevertheless, the A413V mutation found in association with three types of insertions, has recently been selected during serial passages of E. coli in the presence of aztreonam and shown to lead to a four-fold decreased susceptibility to this antibiotic 38 .…”

“…These insertions result from duplications starting two and three codons upstream the YRIN duplication respectively (Fig. 3a) 25 . To determine whether the association between the acquisition of a carbapenemase gene and a mutated PBP3 characterized by a four AA insertion is specific to ST410 isolates or if is also observed in other E. coli lineages, we analysed for these two features E. coli and Shigella genomes from the NCBI database.…”