While aztreonam-avibactam is a potent β-lactam-β-lactamase-inhibitor combination, reduced in vitro activity against some Enterobacterales isolates has been reported. In this study, globally collected clinical isolates of Enterobacterales with elevated minimum inhibitory concentrations (MICs) for aztreonam-avibactam were examined for potential resistance mechanisms. Isolates with aztreonam-avibactam MICs ≥8 μg/mL (n = 55: Escherichia coli, n = 38; Enterobacter cloacae, n = 10; Klebsiella pneumoniae, n = 3; others, n = 4) and <8 μg/mL (n = 18) collected for the INFORM global surveillance programme were characterized by short read whole-genome sequencing. Sequences were inspected for the presence of β-lactamase genes, penicillin-binding protein (PBP) mutations, and disruptions in the coding sequences of porin genes. All isolates of E. coli testing with aztreonam-avibactam MIC values ≥8 μg/mL carried a previously documented four-amino-acid insertion in PBP3 at position 333 of YRI(K/N/P). Such mutations were absent in isolates with MICs <2 μg/mL (n = 6). Among other species, carriage of PER- or VEB-type β-lactamases was identified in 10/17 (58.8%) of isolates testing with aztreonam-avibactam MICs ≥8 μg/mL, but no isolates with lower MIC values (n = 11). PBP3 mutations are known to confer resistance to aztreonam in E. coli, providing a rationale for the elevated MIC values for aztreonam-avibactam in these isolates. Elevated MICs in other isolates were associated with the carriage of PER-type β-lactamases, which have been previously shown to be inhibited less effectively by avibactam than other Class A β-lactamases and may contribute to this phenotype. Other resistance mechanisms contributing to poor in vitro activity for aztreonam-avibactam in some of these isolates are not yet elucidated.

We evaluated the activity of ceftolozane/tazobactam (C/T), and comparators against clinical Pseudomonas aeruginosa isolates collected for the global Study for Monitoring Antimicrobial Resistance Trends (SMART) surveillance program in ten countries in the Middle East and Africa to augment scarce standardized surveillance data in this region. Minimum inhibitory concentrations (MICs) were determined using Clinical and Laboratory Standards Institute broth microdilution and interpreted with European Committee on Antimicrobial Susceptibility Testing breakpoints. P. aeruginosa isolates testing with C/T MIC >4 mg/l or imipenem MIC >2 mg/l were screened for β-lactamase genes. C/T was active against 91.4% and 87.0% of P. aeruginosa isolates from the Middle East and Africa, respectively (14-21 and 7-16 percentage points higher than most β-lactam comparators, respectively). Considerable variation in susceptibility was seen across countries, which largely correlated with the observed prevalence of carbapenemases and/or extended-spectrum β-lactamases. Differences across countries were smaller for C/T than for the β-lactam comparators, ranging from 81% C/T-susceptible among isolates from Jordan to 95% for Qatar. Among subsets resistant to meropenem, ceftazidime, or piperacillin/tazobactam, C/T maintained activity against 51-73% of isolates from the Middle East and against 27-54% from Africa (where metallo-β-lactamase and GES carbapenemase rates were higher). Given the desirability of β-lactam use among clinicians, C/T represents an important option in the treatment of infections caused by P. aeruginosa.

Ceftazidime-avibactam (CZA) and ceftolozane-tazobactam (C/T) are important agents for treating multidrug-resistant P. aeruginosa infections. In this study, we evaluated the molecular characteristics of 300 globally collected clinical P. aeruginosa isolates non-susceptible (NS) to CZA, C/T, or both agents. Isolates were CZA-NS and C/T-NS ( n = 57), CZA-susceptible (S) and C/T-NS ( n = 145), or CZA-NS and C/T-S ( n = 98) selected from the Antimicrobial Testing Leadership and Surveillance (ATLAS) surveillance program from 2020 to 2021. Characterization was by whole-genome sequencing. Analysis was performed to identify β-lactamase genes and mutations that impact efflux regulation, AmpC regulation, and target binding (PBP3). Of the 57 CZA-NS+C/T-NS isolates, 64.9% carried a metallo-β-lactamase (MBL), and a cumulative 84.2% carried any non-intrinsic β-lactamase [i.e., not Pseudomonas- derived cephalosporinase (PDC) or OXA-50-like]. Of the 145 CZA-S+C/T-NS isolates, 26.2% carried an extended-spectrum β-lactamase (ESBL) and no carbapenemase, 17.9% carried a serine-carbapenemase, and 42.1% were negative for non-intrinsic β-lactamases. Of 98 CZA-NS+C/T-S isolates, 34.7% carried mutations previously described as causing an upregulation of the MexAB-OprM efflux pump, while only 9.2% carried a non-intrinsic β-lactamase, and no resistance mechanism was identified in 29.6% of these isolates. MBLs were present in most isolates NS to both agents. More than half of the CZA-S+C/T-NS isolates carried serine β-lactamases. The most frequently identified resistance mechanism identified in CZA-NS+C/T-S isolates was a marker indicating the upregulation of MexAB-OprM. No mechanism was identified that is thought to support resistance to these agents in numerous isolates. This may be due in part to the fact that whole genome sequencing (WGS) cannot directly measure gene expression of chromosomal resistance mechanisms.

Objectives: We assessed the in vitro activity of aztreonam–nacubactam and cefepime–nacubactam against a 2021–2023 global collection of clinical isolates of Gram-negative bacilli (12 613 Enterobacterales, 3748 Pseudomonas aeruginosa and 601 Acinetobacter baumannii). Nacubactam is a novel diazabicyclooctane β-lactamase inhibitor that also possesses intrinsic antimicrobial (PBP2 inhibition) and β-lactam enhancer (for β-lactams that bind PBP3 and other non-PBP2 PBPs) activities against Enterobacterales. Aztreonam–nacubactam and cefepime–nacubactam are in clinical development for the treatment of complicated urinary tract infections, acute uncomplicated pyelonephritis and various infections caused by carbapenem-resistant Enterobacterales (CRE). Methods: MICs were determined for aztreonam–nacubactam (1:1 ratio), cefepime–nacubactam (1:1 ratio) and 13 comparators using the CLSI broth microdilution method. Results: For all Enterobacterales isolates, the MIC90s of aztreonam–nacubactam and cefepime–nacubactam were 1 and 0.5 mg/L, respectively; 99.8% of all Enterobacterales isolates and 99.8% of CRE isolates (n = 917) were inhibited by aztreonam–nacubactam at 4 mg/L and by cefepime–nacubactam at 8 mg/L. Both aztreonam-resistant and cefepime-resistant isolate MICs demonstrated a modal decrease of 64-fold by the addition of nacubactam at a 1:1 ratio. Aztreonam–nacubactam and cefepime–nacubactam were active against the majority of Enterobacterales isolates that were resistant to aztreonam–avibactam, ceftazidime–avibactam, imipenem–relebactam, meropenem–vaborbactam and cefiderocol. The MIC90s for aztreonam–nacubactam and cefepime–nacubactam were 16 and 8 mg/L for P. aeruginosa and >64 and 64 mg/L for A. baumannii. Discussion: The results of this study support the continued development of aztreonam–nacubactam and cefepime–nacubactam as potential therapies for infections caused by CRE where resistance or potential toxicity to marketed antimicrobial agents excludes their use.

The NG-test CARBA 5 and Xpert Carba-R IVD kits were tested against 269 clinical Enterobacterales isolates selected for their previously determined carbapenemase content from the ATLAS Global Surveillance program, primarily from 2020 to 2022. Antimicrobial susceptibility testing by broth microdilution was performed for aztreonam-avibactam, cefiderocol, and othe agents using EUCAST 2025 breakpoints for interpretation. The CARBA 5 and Carba-R IVD kits accurately identified all NDM (93), KPC (36), and OXA-48-like (43) carbapenemases. The CARBA 5 detected all IMP (42) and 45/47 VIM, and the Carba-R detected all VIM (47) and 21/42 IMP. Aztreonam-avibactam was active against carbapenemase-producing isolates (percent susceptible [carbapenemases]): 90.7% (NDM), 92.9% (NDM+OXA-48-like), 100% (IMP, VIM, and KPC), and 88.9% (OXA-48-like). Cefiderocol susceptibility was as follows: 38.7% (NDM), 50.0% (NDM+OXA-48-like), 77.5% (IMP), 82.5% (VIM), 75.9% (KPC), and 100% (OXA-48-like). The most common carbapenemases in Enterobacterales include NDM, KPC, and OXA-48-like enzymes, all of which were accurately identified by the CARBA 5 and Carba-R kits. Aztreonam-avibactam demonstrated potent in vitro activity against isolates determined to be positive for carbapenemases by these kits, offering a potential therapeutic option when one of these isolates is encountered in the clinic. IMPORTANCE: In vitro diagnostic (IVD) kits exist that detect carbapenemases in clinical isolates of Enterobacterales. Their utility is hampered by the limited options to reliably treat infections caused by Enterobacterales that produce metallo-β-lactamases. Here, we test the utility of IVD kits for detecting carbapenemases in a recent collection of clinical Enterobacterales isolates and determine the in vitro activity of aztreonam-avibactam, cefiderocol, and other comparator agents against isolates that were positive for carbapenemases.

Objectives: Nacubactam is a novel diazabicyclooctane β-lactamase inhibitor with activity against PBP2 that is in clinical development with aztreonam or cefepime. The in vitro activities of aztreonam/nacubactam, cefepime/nacubactam and comparator agents were determined against carbapenemase-producing Enterobacterales, collected globally from 2021 to 2023. Methods: Isolates were collected as part of a surveillance study from 24 countries in Asia, Europe and North America. Susceptibility testing was by CLSI broth microdilution. Meropenem-non-susceptible Enterobacterales isolates were sequenced by nanopore technology to identify resistance mechanisms. Of 12 613 isolates collected, 630 carbapenemase-producing isolates were detected. Results: All KPC-producing (158) and OXA-48-like–producing (142) isolates were inhibited by ≤4/4 mg/L of aztreonam/nacubactam or ≤8/8 mg/L cefepime/nacubactam. Aztreonam/nacubactam at ≤4/4 mg/L was active against 98.7% of the 307 NDM-producing isolates, more than aztreonam/avibactam (95.1% susceptible) or cefiderocol [85.7% susceptible (CLSI breakpoints)], while cefepime/nacubactam was active against 94.1% at ≤8/8 mg/L. Insertions in PBP3 were present in 39/47 (83%) of NDM-positive Escherichia coli. Aztreonam/nacubactam was active against 97.4% of these at ≤4/4 mg/L and cefepime/nacubactam was active against 100% at ≤8/8 mg/L, while aztreonam/avibactam and cefiderocol activities were relatively poor [69.2% aztreonam/avibactam-susceptible and 46.2% cefiderocol-susceptible (CLSI)]. Conclusions: Aztreonam/nacubactam and cefepime/nacubactam demonstrated potent in vitro activity against Enterobacterales that carried serine-carbapenemases and MBLs, including those with chromosomal resistance mechanisms that compromise the current best-in-class β-lactam antibiotics for treating infections caused by MBL-positive organisms. The percentage of NDM-positive E. coli with insertions in PBP3 is alarming and warrants continued surveillance.