Detection of MexR and MexZ efflux pump genes and assessment of the synergistic effect of amikacin and gentamycin with some efflux pump inhibitors in antibiotic-resistant Pseudomonas aeruginosa

Background: A bacterium called Pseudomonas aeruginosa causes infections with high mortality rates and has the ability to rapidly acquire resistance to a variety of antibiotics.Methods: A total of 118 clinical specimens from both males and females of different age groups were gathered between early September 2022 and the end of December 2022 from several hospitals in Al-Anbar city, Iraq. The sample sources were wounds, burns, urinary tract infections (UTI), and sputum. P. aeruginosa was identified through its microscopical properties, confirmed using the VITEK 2 automated system, followed by molecular detection using the 16S rRNA gene. The Kirby-Bauer disk diffusion method was used based on CLSI guidelines (CLSI, 2022) for the aminoglycoside antibiotics class. DNA extraction from the twenty isolates was conducted using Presto™ Mini gDNA Bacteria Kit. After that, molecular analysis of MexR and MexZ was performed using the PCR technique.Results: Within this, 65 isolates were determined as P. aeruginosa from 118 samples. The proportion of P. aeruginosa isolates from wound samples was 27/65 (41.5%), while in the burn wound, Urinary tract infection (UTI), and sputum were 20/65 (30.8%), 14/65(21.5%), and 4/65(6.2%), respectively. The findings of the test for antibiotic resistance were Amikacin (73.84%), Gentamycin (81.53%), Netilmicin (33.84%), and Tobramycin (66.15%). The molecular analysis of MexZ genes from the twenty isolates showed the highest prevalence, 20/20 (100%), and the MexR gene 14/20 (70%). The antagonism of aminoglycoside antibiotics by divalent cations was found, and it is noted that the presence of divalent cations increased the antibiotic Minimum Inhibitory Concentration (MIC) values.Conclusion: It was found that MexR and MexZ genes are widespread in P. aeruginosa isolates, which indicated elevated mexXY-OprM and MexAB-OprM gene expression in P. aeruginosa. These elevated efflux pumps of P. aeruginosa may influence antibiotic ion fluxes at the bacterial cell membrane, contributing to the improved resistance to aminoglycosides. The aminoglycoside resistance isolates expressed decreased efflux pumps when inhibited by CCCP and Conessine.Keywords: Pseudomonas aeruginosa, Efflux pump inhibitors, MexXY-OprM efflux system, Conessin