Restoration of β-lactam activity by nacubactam combinations against extensively drug-resistant K. pneumoniae harbouring diverse carbapenem resistance gene

Background: Carbapenem-resistant K. pneumoniae (CRKP) has emerged as a critical global health threat due to limited therapeutic options and high rates of multidrug resistance. Novel β-lactamase inhibitors such as Nacubactam may enhance the activity of conventional agents against carbapenemase-producing strains. Methods: This experimental study investigated 200 CRKP isolates identified using the VITEK 2.0 system and subjected to antimicrobial susceptibility testing. Carbapenemase production was assessed phenotypically by mCIM and eCIM assays, and molecular detection of resistance genes (NDM-1, OXA-48, KPC, VIM, IMP) was performed by multiplex PCR. Synergistic antimicrobial properties displayed when nacubactam is administered together with meropenem (MRP) and cefepime (CFM) was evaluated using checkerboard microdilution and time-kill spot assays. Results: Of 200 isolates, 150 (75%) demonstrated carbapenemase production. Among these, 65 (43.3%) were metallo-β-lactamase producers while 85 (56.6%) produced non-MBL carbapenemase (Serine type). Genotypic profiling revealed NDM-1 in 33.3% of isolates, OXA-48 in 31.8%, and KPC in 9.41%, with frequent co-harboring of multiple genes. None carried IMP or VIM gene. All isolates exhibited multidrug resistance, with 100% resistance to most β-lactams, aminoglycosides, fluoroquinolones, and carbapenems. Colistin remained universally active. Checkerboard assays demonstrated pronounced synergy of Nacubactam with meropenem against NDM-1, OXA-48, and KPC producers (FICI ≤ 0.285), while combinations with cefepime showed synergistic or additive activity (FICI 0.41–0.58). Time-kill assays confirmed bactericidal synergy, with ≥ 3 log₁₀ CFU reduction relative to single agents. Conclusion: This study highlights a high prevalence of NDM-1 and OXA-48 carbapenemase among CRKP isolates with extensive multidrug resistance. Nacubactam exhibited significant synergistic activity with meropenem, and moderate synergy with cefepime, across a range of genotypic variants. These findings suggest that Nacubactam–β-lactam combinations may represent a promising therapeutic strategy against CRKP infections, warranting further clinical evaluation.