Phenotypic and molecular detection of efflux pump and expression profiling genes in Klebsiella pneumoniae exposed to ciprofloxacin and amoxicillin/clavulanic acid.

BACKGROUND: One of the main strategies Gram-negative bacteria employ to lower internal antibiotic concentrations is efflux. By identifying and eliminating a variety of harmful substances, including antibiotics, these active transport systems help bacteria become less susceptible to drugs and develop new resistance mechanisms. Efflux pumps are involved in bacterial stress response, pathogenicity, biofilm formation, and host physiology alteration in addition to offering an escape from antibacterial chemicals.

OBJECTIVE: This research is designed to study the prevalence of efflux pump genes and the measure expression of )AcrAB, MarA, MdtK OmpK35 and OmpK36 genes before and after treatment with combination of Ciprofloxacin (CIP) and Amikacin (AMC) antibiotic CIP of Klebsiella pneumoniae isolated from hospitalized patients in Iraq hospitals.

MATERIALS AND METHODS: During a six-month cross-sectional survey (January-June 2025), twenty K. pneumoniae strains were collected from patients admitted to several teaching hospitals. Their antimicrobial sensitivity was determined by applying the agar-based disk diffusion assay. The ethidium bromide (EtBr) agar cartwheel method can be used to phenotypically detect the activity of efflux pumps in K. pneumoniae isolates. The PCR method was used to determine the frequency of genes encoding AcrAB, marA, mdtK, ompK35, and ompK36 in K. pneumoniae, and the Real-time PCR method was used to determine the expression of the efflux pump gene exposed to ciprofloxacin and amoxicillin.

RESULTS: The findings indicate that 90% of K. pneumoniae phenotypically displayed resistance to tetracycline, 90% amoxicillin/clavulanic acid, 90% piperacillin, 90% ciprofloxacin, and 89%. All the Klebsiella isolates show fluorescence under UV light at varying concentrations of ethidium bromide, indicating efflux pump activity. The higher prevalence of AcrAB and MarA genes100%. A multidrug efflux pump-associated gene is identified among the isolates. Moreover, the porin genes OmpK35 and OmpK36 are observed in 66.66% of the samples. In contrast, only 50% of the K. pneumoniae strains harbored the MdtK efflux pump gene. However, the analysis show that 50% of the isolates carry the MarkD gene, which play crucial roles in biofilm formation. The expression of AcrAB, MarA, MdtK ompK35 and OmpK36 genes is higher and exposed to ciprofloxacin and amoxicillin.

CONCLUSIONS: The efficacy of synergistic antibiotic and efflux pump inhibitor combinations in eradicating MDR bacterial resistance is an encouraging discovery.