ABSTRACT
Multidrug resistance (MDR) is the ability of bacteria to withstand lethal doses of drugs diverse in their mechanism of action, which would be effective in the elimination of susceptible strains. Bacterial multidrug efflux pump is an important mechanism of antibiotic resistance and is required for many bacteria to cause infections. Routine antimicrobial sensitivity test fails to detect efflux pump mediated drug resistance. Early recognition of this mechanism would allow the use of alternative antibiotics and an efflux pump inhibitor in combination with the antibiotic therapy. This study was aimed at the detection of efflux pump-mediated drug resistance, which should be included in the diagnostic regimen to facilitate implementation of appropriate therapy to the ailing patient.
A total of 140 consecutive isolates (70 isolates each of Staphylococcus aureus and Pseudomonas aeruginosa) were obtained from various clinical specimens (urine, pus, catheter tip, high vaginal swab, wound, ear, throat and urethral swabs) sent to the Bacteriology Laboratory Central Hospital Benin City. These isolates were identified by standard bacteriological procedures and stored in agar slants for further analysis. All isolates were reactivated by subculturing onto nutrient agar and MacConkey agar. Phenotypic detection of some virulence factors namely hemagglutination test, capsule production and hemolysin production were done. Antibiotics susceptibility test of the isolates was carried out using the modified Kirby-Bauer disc diffusion method. Multidrug resistant isolates were screened for efflux activity using the Ethidium bromide agar cartwheel method. Efflux pump inhibitor based method was used to confirm efflux activity.
Hemolysin was the virulence factor produced by most isolates 71.4% (n = 50) for S. aureus isolates and 75.7% (n = 53) for P. aeruginosa isolates. A percentage of 38.6% (n = 27) of S. aureus isolates were positive for hemagglutination test while 74.3% (n = 52) of P. aeruginosa isolates were able to agglutinate red cells. Only about 15.7% (n = 11) of S. aureus isolates and 24.3% (n = 17) of P. aeruginosa isolates were positive for possession of capsules. Antibiotics susceptibility pattern of S. aureus revealed that the isolates were mainly susceptible to Ofloxacin (92.9%), Gentamicin (85.7%) and Erythromycin (51.4%) with the least susceptibility to Cloxacillin (4.3%). Similarly, P. aeruginosa isolates were mainly susceptible to Ofloxacin (92.9%), Gentamicin (90%) and Erythromycin (41.4%), with least susceptibility to Augmentin (1.4%). Multidrug resistance was observed in 41. 4% (n=29) of S. aureus isolates and 44.3% (n = 31) of P. aeruginosa isolates. The ethidium bromide agar cartwheel method of screening revealed efflux activity in 48.3% (n = 14) of the multidrug resistant S. aureus isolates and 58.1% (n = 18) of the multidrug resistant P. aeruginosa isolates. A higher efflux activity was observed in P. aeruginosa in comparison to S. aureus with 12.9% (n=4) of the MDR P.aeruginosa isolates yet to fluoresce at a concentration as high as 2.5mg/l ethidium bromide. The confirmatory efflux pump inhibitor based method showed efflux activity in 28% (n =8) of the multidrug S. aureus isolates with 24% (n=7) active against more than one antibiotic. Omeprazole as an efflux pump inhibitor was not effective for isolates of P. aeruginosa, as the presence of omeprazole did not give a four-fold decrease in MIC of antibiotics against P. aeruginosa isolates. This study has revealed the uprise of multidrug resistance among isolates of S. aureus and P. aeruginosa studied and has revealed that as much as 28% of multidrug resistance among isolates of S. aureus in this study is attributable to efflux pump related mechanisms; hence the need for inclusion of efflux related test in the diagnostic regimen for multidrug resistant bacteria.
