High in vitro activity of aztreonam/avibactam against NDM-producing Enterobacterales with emerging resistance in Escherichia coli in the United States Mid-Atlantic region

ABSTRACT New Delhi metallo-β-lactamase (NDM)-producing Enterobacterales represent a critical antimicrobial resistance threat with limited therapeutic options. Aztreonam/avibactam, which combines the stability of aztreonam against metallo-β-lactamases with the inhibition of serine β-lactamases by avibactam, is a promising therapeutic candidate. However, emerging resistance associated with ftsI (penicillin-binding protein 3 PBP3) insertions and expression of AmpC β-lactamases has been described. We evaluated 95 NDM-producing Enterobacterales isolates submitted to the Maryland Department of Health Laboratories Administration through the AR Lab Network between 2020 and 2025. Susceptibility testing for aztreonam, ceftazidime/avibactam, and aztreonam/avibactam was performed by broth microdilution. Whole-genome sequencing was used to identify sequence types, resistance genes, and ftsI alterations. The isolates included Escherichia coli ( n = 40), Klebsiella pneumoniae ( n = 32), Enterobacter cloacae complex ( n = 8), and other Enterobacterales ( n = 15). All isolates carried NDM; 12 harbored OXA-48-like, and 3 harbored KPC carbapenemases. All isolates were resistant to ceftazidime/avibactam, and 83% were non-susceptible to aztreonam alone. In contrast, 86% (82/95) were susceptible to aztreonam/avibactam, 7% intermediate, and 6% resistant. Non-susceptibility was concentrated in E. coli (30%, 12/40) and related to four–amino acid ftsI insertions (YRIN, KYRI), particularly when combined with CMY-type AmpC β-lactamases. Non-susceptible E. coli was genetically diverse and distributed across multiple jurisdictions with no evidence of clonal spread. Aztreonam/avibactam demonstrated high in vitro activity against NDM-producing Enterobacterales, but resistance is emerging, predominantly in E. coli with PBP3 alterations and AmpC co-production. These findings underscore the importance of susceptibility testing and genomic surveillance to preserve the clinical utility of aztreonam/avibactam as its clinical use expands. IMPORTANCE The approval of aztreonam/avibactam provides a critical therapeutic option for treating infections caused by metallo-β-lactamase-producing Enterobacterales, including NDM-positive strains that are resistant to nearly all other β-lactams. However, emerging resistance due to PBP3 ( ftsI ) insertions and AmpC co-production threatens its efficacy. This study demonstrates that while aztreonam/avibactam remains highly active against most NDM-producing Enterobacterales, resistance is already emerging in diverse Escherichia coli lineages carrying specific ftsI insertions. By correlating phenotypic susceptibility with genomic determinants, our findings underscore the need for laboratories to implement standardized testing for aztreonam/avibactam and for continued genomic surveillance to detect evolving resistance. This work provides critical regional data supporting stewardship and diagnostic preparedness as clinical use of aztreonam/avibactam expands.