Changes in Outer Membrane Proteins of Benzalkonium Chloride Adapted Pseudomonas Aeruginosa and Mutations in GyrA and ParC Genes (original) (raw)
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Antibiotic Resistance Induction by Benzalkonium Chloride Exposure in Nosocomial Pathogens
International Journal of Infection, 2016
Background: Biocides (disinfectants) are crucial for controlling various infections and are widely used in environments for the control of microorganisms. Exposure of bacteria to biocides can select for mutants with decreased biocide susceptibility that often display a decrease in susceptibility to antibiotics. Objectives: The present work was done during 14 months from February 2015, at the microbiology laboratory of the Veterinary college of Shahrekord university with the aim of investigating the impact of benzalkonium chloride exposure on antibiotic resistance in some common nosocomial pathogens. Methods: Standard strains of Pseudomonas aeruginosa, Acinetobacter baumannii, Escherichia coli and Staphylococcus aureus were used, and in parallel to each standard strain four hospital isolates collected from Shahrekord and Esfahan hospitals were examined. Tube double serial dilution method was used for determination of minimum inhibitory concentration (MIC) of antibiotics and Benzalkonium Chloride (BKC). Spontaneous mutants were developed by exposure of examined species to BKC, and their mean MICs to examined drugs were evaluated. Results: The mean MICs of the BKC and antibiotics used in this study were not similar between the parent and mutant strains of the examined isolates. In E. coli, A. baumannii and S. aureus isolates, differences between the mean MICs of BZK and ciprofloxacin had a similar pattern and were statistically significant (P < 0.05). In isolates of P. aeruginosa, differences between parent and mutant isolates for all of the tested drugs were significant. Conclusions: Vast use of BKC in various environments and their accumulation represents a potential risk for selective pressure towards selection of bacteria with decreased antibiotic susceptibility.
Study of cross-resistance to other antimicrobial agents and efflux pump (EP) resistance mechanism in benzalkonium chloride (BC) adapted Pseudomonas aeruginosa isolates. Minimum inhibitory concentration (MIC) of BC of 88 P. aeruginosa isolates that collected from clinical settings in Egypt and cross-resistance of BC-adapted isolates to other antimicrobial agents were determined by agar dilution method. EP regulatory genes (i.e., MexR, NfxB, MexT, and MexZ) were detected by PCR in BC adapted isolates, and then EP activity of isolates which have these four genes was determined by cartwheel method followed by sequencing for the isolate which showed the highest EP activity. Finally; the expression of EP genes (MexA, MexC, MexE, and MexX) was determined in the same isolate by realtime polymerase chain reaction (RT-PCR) in presence and absence of the most potent efflux pump inhibitor (EPI). Twenty isolates were BC-adapted with MIC 2,048 mg/l, showed cross-resistance to cetrimide and ciprofloxacin but not to other disinfectants used. The isolate no. 87 selected to be sequenced. Mutation founded in MexR (V126E) and MexZ (L138R) genes. By RT-PCR, sertraline (the most potent EPI used) decreased the expression of the EP genes to three-folds. These results demonstrate that EP was the major mechanism of resistance to BC.
Journal of Basic …, 2011
The main goal of this work was to examine whether the continuous exposure of single and binary P. aeruginosa and E. coli biofilms to sub-lethal benzalkonium chloride (BC) doses can induce adaptive response of bacteria. Biofilms were formed during 24 h and then put continuously in contact with BC for more 5 days. The six-day-old adapted biofilms were then submitted to BC challenge, characterized and inspected by SEM. Both single and binary adapted biofilms have clearly more biomass, polysaccharides and proteins and less activity even though the number of cells was identical. After BC treatment, adapted biofilms maintained their mass and activity. SEM examination revealed that those adapted biofilms had a slimier and denser matrix that became thicker after BC treatment. Continuous exposure of bacteria to antimicrobials can lead to development of biofilms encompassing more virulent and tolerant bacteria. This adaptive resistance can be the result of a phenotypic adaptation, a genetic acquired resistance or both. Instead of eradicating biofilms and kill microorganisms, the use of a disinfectant can, favour biofilm formation and tolerance. This must be a genuine concern as it can happen in clinical environments, where the use of antimicrobials is unavoidable.
Microbiology, 2007
Benzalkonium chloride (BC) is a commonly used disinfectant and preservative. This study describes changes in expression level at the transcriptomic and proteomic level for Escherichia coli K-12 gradually adapted to a tolerance level to BC of 7-8 times the initial MIC. Results from DNA arrays and two-dimensional gel electrophoresis for global gene and protein expression studies were confirmed by real-time quantitative PCR. Peptide mass fingerprinting by MALDI-TOF MS was used to identify differentially expressed proteins. Changes in expression level in adapted cells were shown for porins, drug transporters, glycolytic enzymes, ribosomal subunits and several genes and proteins involved in protection against oxidative stress and antibiotics. Adapted strains showed increased tolerance to several antibiotics. In conclusion, E. coli K-12 adapted to higher tolerance to BC acquired several general resistance mechanisms, including responses normally related to the multiple antibiotic resistance (Mar) regulon and protection against oxidative stress. The results revealed that BC treatment might result in superoxide stress in E. coli.
Benzalkonium chloride antagonises aminoglycoside antibiotics and promotes evolution of resistance
EBioMedicine, 2021
Background: Biocide disinfectants are essential tools in infection control, but their use can inadvertently contribute to emergence of antibiotic-resistant bacteria. In this study we systematically examine the effect of the biocide benzalkonium chloride, which is primarily used for surface disinfection but is also present as a preservative in many consumer products, on the activity of aminoglycoside antibiotics in Acinetobacter baumannii. Methods: The effect of subinhibitory BAC on aminoglycoside treatment of A. baumannii ATCC17978 was investigated using time-to-kill assays, MIC determination, directed evolution experiments, fluctuation tests and labelled gentamicin accumulation assays. Further MIC determinations and directed evolution experiments were performed with additional Gram-negative ESKAPE pathogens. Findings: In A. baumannii ATCC17978, BAC prevents gentamicin killing and drastically increases the frequency at which resistant mutants emerge, through reducing intracellular antibiotic accumulation. BAC also increases the MIC of multiple aminoglycoside antibiotics (kanamycin, tobramycin, streptomycin, gentamicin and amikacin). BAC promotes the emergence of mutants with reduced gentamicin susceptibility in other Gram-negative ESKAPE pathogens but does not always alter the MIC. These effects occur at BAC concentrations which are similar to residual levels in high-use environments, and just below the concentration range for BAC when used as a preservative in eye drops and ear drops. Interpretation: Our results suggest that subinhibitory BAC has the potential to antagonise aminoglycoside activity and promote the emergence of bacterial mutants with reduced susceptibility. We suggest that the extremely widespread use of BAC in clinical and home settings and its long half-life mean there is potential for these interactions to occur in the environment, or in patients who use BAC-containing products while taking aminoglycosides to treat skin, eye or ear infections, although such co-exposure is likely to be rare. We suggest that biocide stewardship is needed to prevent the types of exposure that can contribute to antibiotic resistance.
Excessive discharge of quaternary ammonium disinfectants such as benzalkonium chloride (BAC) into aquatic systems can trigger several physiological responses in environmental microorganisms. In this study, we isolated a low susceptible strain ofAeromonas hydrophila, namely INISA09, to from a wastewater treatment plant in Costa Rica. We characterized its phenotypic response upon exposure to three different concentrations of BAC and determined the primary mechanisms related to its resistance using genomic and proteomic tools. The genome of the strain is 4.6-Mb with 4,273 genes, and we found a propound genome rearrangement and thousands of missense mutations when compared to the reference strainA. hydrophilaATCC 7966. We identified 15,762 missense mutations mainly associated with transport, antimicrobial resistance, and outer membrane proteins. In addition, quantitative proteomic analysis revealed the significant upregulation of several efflux pumps and the downregulation of porins whe...
Antimicrobial Agents and Chemotherapy, 2003
The adaptation mechanism of Pseudomonas aeruginosa ATCC 10145 to quaternary ammonium compounds (QACs) was investigated. A P. aeruginosa strain with adapted resistance to QACs was developed by a standard broth dilution method. It was revealed that P. aeruginosa exhibited remarkable resistance to N -dodecylpyridinium iodide (P-12), whose structure is similar to that of a common disinfectant, cetylpyridinium chloride. Adapted resistance to benzalkonium chloride (BAC), which is commonly used as a disinfectant, was also observed in P. aeruginosa . Moreover, the P-12-resistant strain exhibited cross-resistance to BAC. Analysis of the outer membrane protein of the P-12-resistant strain by two-dimensional polyacrylamide gel electrophoresis showed a significant increase in the level of expression of a protein (named OprR) whose molecular mass was approximately 26 kDa. The actual function of OprR is not yet clear; however, OprR was expected to be an outer membrane-associated protein with homo...
Adaptation of Pseudomonas aeruginosa to 2,2'-methylenebis (4-chlorophenol
Journal of Applied Microbiology, 1993
A culture of Pseudomonas aeruginosa, isolated from a cooling water system, was grown in the presence of sub-inhibitory concentrations of 2,2′-methylenebis(4-chlorophenol) (MBC). It adapted to increasing concentrations from an initial minimum inhibitory concentration of 36 μg ml-1 to the highest, 80 μg ml-1. Resistant cultures exhibited a higher survival rate when exposed to 320 μg ml-1 than did the original strain. Lipopolysaccharide and outer membrane protein profiles were determined by SDS PAGE. No changes were detected in lipopolysaccharide profiles. The quantity of OprP, the phosphate uptake protein in the outer membrane, decreased to a low level correlating with decreased phosphate (Pi) uptake during growth. It is proposed that OprP is the place of entry for MBC and that the cell can adapt by decreasing the level of OprP in the outer membrane.
Journal of Babol University of Medical Sciences, 2019
BACKGROUND AND OBJECTIVE: The resistance genes of quaternary ammonium compounds(qac) play an important role in the resistance of gram-negative bacteria producing broad-spectrum beta-lactamases to disinfectants. The aim of this study was detection of qacEΔ1, qacG, qacE, qacF resistance genes in Escherichia coli producing broadspectrum beta-lactamases to benzalkonium chloride. METHODS: This study cross sectional-descriptive was conducted on 150 clinical samples of selected hospitals in Arak. ESBL strains were identified by using phenotypic methods of disc diffusion and combinatory disc method and evaluating the SHV, TEM, CTXM1 genes by genotyping method. The PCR was performed to determine the resistance genes qacEΔ1, qacG, qacE and qacF.The electrophoresis of PCR products and the MIC of benzalkonium chloride were relative to E. coli producing ESBL. Antibiotic pattern of Escherichia coli (ESBL), quadruple ammonium resistance genes and benzalkonium chloride MIC were also investigated. FINDINGS: This study showed that 60% of Escherichia coli were ESBL producer. The qacEΔ1 genes were observed in all of them and qacE, qacF, qacG genes were not found in any of the strains. The strains had MIC range from 32 to 64 mg/l for benzalkonium chloride. Resistance to carbapenems (33.33%) was observed. CONCLUSION: This study showed that qacEΔ1 resistance gene and resistance to disinfectant benzalkonium chloride increased. Also increased resistance to the antibiotics studied were observed in E. coli ESBL strains.
Detection of benzalkonium chloride resistance in community environmental isolates of staphylococci
We isolated a total of 653 strains from 64 community environmental samples in Massachusetts, USA. Among these isolates, 9.65 % (63 strains) were benzalkonium chloride (BC)-resistant staphylococci. All BC-resistant strains were collected from surfaces upon which antibacterial wipes or antibacterial sprays containing 0.02–0.12 % BC had frequently been used in the fitness centres. However, isolates from surfaces upon which antibacterial wipes or antibacterial sprays had not been used were all sensitive to BC. All BC-resistant strains were also resistant to erythromycin, penicillin and ampicillin. In addition, 51 strains showed resistance to cetyltrimethylammonium bromide (CTAB), 15 strains showed resistance to chloramphenicol, 12 strains showed resistance to ciprofloxacin and four strains showed resistance to meticillin. Resistance gene analysis demonstrated that 41 strains contained qacA/B, 30 strains had qacC, 25 strains contained qacG, 16 strains had qacH and eight strains contained qacJ. These data indicate that application of BC is associated with environmental staphylococcal antimicrobial resistance.