Investigation Of Pseudomonas Aeruginosa Genes Associated With Intrinsic Antibiotic Resistance

Several antiobiotics resistant Gram-negative pathogens are responsibie for the major of nosocomial infections, Pseudomonas aeruginosa is one of them. It exhibits high intrinsic and acquired resistance to antimicrobial agents. Ttreatment of serious infections is often compli-cated by pathogens'antibiotic resistance. So investigate antiobiotics resistence mechanism is reguired in order to control bacterial infections.In our previous work, we used seven antibiotics to screen a P. aeruginosa transposon insertion library and identified 43 genes that are involved in intrinsic antibiotic resistance. Among them, PA2580 showed important effect on the pathogen's resistance to several antibiotics and PA4747 was identified by Dotsch et al had similarity characteristic. So they have been chosen as the subjects of the current research.In order to determine the mechanisms of antibiotics resistence, several experiments have been carried out, including construction of mutant strain, antimicrobial susceptibility assays, monitoring gene expression,in vitro expression and purification of PA2580 protein and measurement of virulence changes.The results shows that subinhibitory concentrations of chloramphenicol, spectinomycins and tetracycline upregulater PA2580 gene expression, indicating it could be induced by antibiotics. PAO(APA2580) mutant strain had elevated susceptibility to ciprofloxacin, carbenicillin and chloramphenicol. Moreover, this mutant exhibited higher tolerance to H2O2 than the wild-type strain, on account of the expression of catalase genes was downregulated. A series of virulence factors genes including the type III secretory system and motility ability were also downregulated in the mutant.Susceptibility test of PAO(APA4747) strain shows that PA4747 gene is involved in intrinsic antibiotic resistence. This mutant was resistant to two antibiotics and the MIC of carbenicillin increased 5 times and the MIC of ampicillin increased 2.5 times compared to the wild type strain. Higher (3-lactamase activity in the mutant Intracellular is probably the cause of the observed antibiotics resistance. In addition, mutation of PA4747 altered pyocyanin production and twitching ability in P. aeruginosa.