OmpK36 deficiency and inducible AmpC β-lactamase synergistically drive imipenem resistance in <i>Klebsiella aerogenes</i>

ABSTRACT This study systematically elucidates how ST88-KL186 Klebsiella aerogenes evolved carbapenem resistance through a dual synergistic mechanism involving porin deficiency and inducible AmpC β-lactamase, providing a mechanistic explanation for carbapenem resistance independent of carbapenemase production. From seven homologous strains (single-nucleotide polymorphism [SNP] difference 3–30) consecutively isolated from a patient with post-pancreaticoduodenectomy infection, three subsequent ascites isolates (R5–R7) developed carbapenem resistance after imipenem treatment. All strains carried the OmpK36 G138S mutation, which reduces pore diameter by ~15%, and displayed an inducible AmpC enzyme. However, an additional Q172Ter truncating mutation in OmpK36 was acquired exclusively in the resistant strains (R5–R7). Molecular dynamics simulations confirmed that the truncated protein loses conformational stability, resulting in a complete loss of functional porin. Although the susceptible strains already had a narrowed pore and inducible AmpC enzyme, residual porin function allowed enough drug entry to maintain susceptibility. In contrast, in resistant strains, loss of OmpK36 served as the primary barrier restricting drug entry, while the small amount of imipenem that entered was sufficient to induce high-level AmpC enzyme expression. The synergistic interplay between these two mechanisms: a “permeability barrier” coupled with “enzymatic hydrolysis”—ultimately overcame the carbapenem resistance threshold. This study underscores the importance of phenotypic screening for inducible AmpC enzymes in clinical practice and provides key evidence for optimizing the diagnosis and management of carbapenem-resistant Gram-negative bacteria. IMPORTANCE The synergistic resistance mechanism of OmpK36 deficiency and induced AmpC β-lactamase revealed in this study provides a critical theoretical supplement for understanding carbapenem resistance independent of carbapenemases. Structure-function analysis confirmed that the truncation mutation in OmpK36 in resistant strains leads to premature termination of translation and complete loss of the protein channel, thereby preventing drug entry into the cell while still triggering high expression of AmpC enzymes by the small amount of drug that does enter. The synergistic effect of these two mechanisms confers the carbapenem-resistant phenotype in Klebsiella aerogenes . This finding not only broadens our understanding of the evolution of adaptive bacterial resistance but also suggests that clinical microbiology laboratories should incorporate testing for this type of inducible AmpC enzyme into routine protocols, thereby providing direct evidence for precise identification of hidden resistance phenotypes and optimization of treatment regimens for severe infections.