Abstract
BackgroundChlorhexidine digluconate (CHG) is used as a disinfectant. The emergence of pathogens resistant to the biocide raises health concern. Information on specific efflux mechanisms utilised by bacteria to confer reduced susceptibility to the biocide, may be used to develop ways of preventing the efflux of the biocide from nosocomial pathogens resulting in higher disinfection activity. The aim of the study was to evaluate the role of ATP-binding cassette transporters on the transport of CHG in bacteria.MethodsClinical strains of Pseudomonas aeruginosa, Staphylococcus aureus and their respective laboratory strains ATCC 27853 and ATCC 9144 were used for susceptibility tests. The minimum inhibitory concentration (MIC) of CHG with or without an efflux pump inhibitor [reserpine or carbonyl cyanide m-chlorophenylhydrazone (CCCP)] was determined using the broth microdilution method. A spectrophotometric method to quantify the amount of chlorhexidine in a sample was developed, validated and used to quantify CHG within P. aeruginosa and S. aureus cells.ResultsIn the presence of reserpine, the MIC of CHG against the clinical strains of P. aeruginosa and S. aureus decreased from 6.3 to 3.2 µg/ml but showed no change against both ATCC isolates. The MIC of CHG in the presence of CCCP for both strains of P. aeruginosa remained unchanged but showed a reduction for both isolates of S. aureus. The suitability of the spectrophotometric method developed for quantifying the amount of CHG accumulated in microbial cells was validated and used successfully to quantify CHG accumulated within bacterial cells.ConclusionThe spectrophotometric determination of CHG within microbial cells may be used to quantify CHG in microbial cells. Only the clinical strain of P. aeruginosa showed significant efflux of CHG suggesting the participation of efflux transporters in the pumping out of CHG from this isolate. The use of efflux pump inhibitors together with the biocide may be explored to preventing the efflux of the biocide from P. aeruginosa resulting in order to increase disinfection activity.
Highlights
Chlorhexidine digluconate (CHG) is used as a disinfectant
A two-fold reduction in minimum inhibitory concentration (MIC) was observed when CHG was used together with 63 μg/ml reserpine against the clinical strain of P. aeruginosa
The MIC of CHG for the two S. aureus strains was below 0.1 μg/ml, which is lower than 6.3 μg/ml obtained when CHG was used on its own for both strains
Summary
Chlorhexidine digluconate (CHG) is used as a disinfectant. The emergence of pathogens resistant to the biocide raises health concern. Information on specific efflux mechanisms utilised by bacteria to confer reduced susceptibility to the biocide, may be used to develop ways of preventing the efflux of the biocide from nosocomial pathogens resulting in higher disinfection activity. Chlorhexidine digluconate is considered to be the “gold standard biocide” showing broad spectrum activity and is used both as a disinfectant and antiseptic [1] Both Grampositive and Gram-negative bacteria show susceptibility to chlorhexidine and the biocide displays bactericidal as well as bacteriostatic activity depending on concentration [6]. Due to the development of resistance of some microbes to biocides, nosocomial pathogens may not be completely eliminated, despite using antimicrobials in disinfecting hospital surfaces and equipment [4]. ABC pumps are primary transporters that use the energy of ATP binding and hydrolysis to export a variety of substrates across cellular membranes [12]
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