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Widely Used Benzalkonium Chloride Disinfectants Can Promote Antibiotic Resistance.

ABSTRACT: While the misuse of antibiotics has clearly contributed to the emergence and proliferation of resistant bacterial pathogens, with major health consequences, it remains less clear if the widespread use of disinfectants, such as benzalkonium chlorides (BAC), a different class of biocides than antibiotics, has contributed to this problem. Here, we provide evidence that exposure to BAC coselects for antibiotic-resistant bacteria and describe the underlying genetic mechanisms. After inoculation with river sediment, BAC-fed bioreactors selected for several bacterial taxa, including the opportunistic pathogen Pseudomonas aeruginosa, that were more resistant to several antibiotics than their counterparts in a control (no BAC) bioreactor. A metagenomic analysis of the bioreactor microbial communities, confirmed by gene cloning experiments with the derived isolates, suggested that integrative and conjugative elements encoding a BAC efflux pump together with antibiotic resistance genes were responsible for these results. Furthermore, the exposure of the P. aeruginosa isolates to increasing concentrations of BAC selected for mutations in pmrB (polymyxin resistance) and physiological adaptations that contributed to a higher tolerance to polymyxin B and other antibiotics. The physiological adaptations included the overexpression of mexCD-oprJ multidrug efflux pump genes when BAC was added in the growth medium at subinhibitory concentrations. Collectively, our results demonstrated that disinfectants promote antibiotic resistance via several mechanisms and highlight the need to remediate (degrade) disinfectants in nontarget environments to further restrain the spread of antibiotic-resistant bacteria.IMPORTANCE Benzalkonium chlorides (BAC) are biocides broadly used in disinfectant solutions. Disinfectants are widely used in food processing lines, domestic households, and pharmaceutical products and are typically designed to have a different mode of action than antibiotics to avoid interfering with the use of the latter. Whether exposure to BAC makes bacteria more resistant to antibiotics remains an unresolved issue of obvious practical consequences for public health. Using an integrated approach that combines metagenomics of natural microbial communities with gene cloning experiments with isolates and experimental evolution assays, we show that the widely used benzalkonium chloride disinfectants promote clinically relevant antibiotic resistance. Therefore, more attention should be given to the usage of these disinfectants, and their fate in nontarget environments should be monitored more tightly.

SUBMITTER: Kim M

PROVIDER: S-EPMC6102991 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature

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Widely Used Benzalkonium Chloride Disinfectants Can Promote Antibiotic Resistance.

Kim Minjae M Weigand Michael R MR Oh Seungdae S Hatt Janet K JK Krishnan Raj R Tezel Ulas U Pavlostathis Spyros G SG Konstantinidis Konstantinos T KT

Applied and environmental microbiology 20180817 17

While the misuse of antibiotics has clearly contributed to the emergence and proliferation of resistant bacterial pathogens, with major health consequences, it remains less clear if the widespread use of disinfectants, such as benzalkonium chlorides (BAC), a different class of biocides than antibiotics, has contributed to this problem. Here, we provide evidence that exposure to BAC coselects for antibiotic-resistant bacteria and describe the underlying genetic mechanisms. After inoculation with ...[more]

PMID: 29959242

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Project description:Disinfectants are routinely used in human environments to control and prevent the transmission of microbial disease, and this is particularly true during the current COVID-19 crisis. However, it remains unclear whether the increased disinfectant loadings to wastewater treatment plants facilitate the dissemination of antibiotic resistance genes (ARGs) in sewage sludge microbiomes. Here, we investigated the impacts of benzalkonium chlorides (BACs), widely used disinfectants, on ARGs profiles and microbial community structures in sewage sludge by using high-throughput quantitative PCR and Illumina sequencing. A total of 147 unique ARGs and 39 mobile genetic elements (MGEs) were detected in all sewage sludge samples. Our results show that exposure to BACs disinfectants at environmentally relevant concentrations significantly promotes both the diversity and absolute abundance of ARGs in sludge microbiomes, indicating the co-selection of ARGs by BACs disinfectants. The enrichment of ARGs abundance varied from 2.15-fold to 3.63-fold compared to controls. In addition, BACs exposure significantly alters bacterial and protistan communities, resulting in dysbiosis of the sludge microbiota. The Mantel test and Procrustes analysis confirm that bacterial communities are significantly correlated with ARGs profiles under BACs treatments. The structural equation model explains 83.8 % of the total ARGs variation and further illustrates that the absolute abundance of MGEs exerts greater impacts on the variation of absolute abundance of ARGs than microbial communities under BACs exposure, suggesting BACs may promote antibiotic resistance by enhancing the horizontal gene transfer of ARGs across sludge microbiomes. Collectively, our results provide new insights into the proliferation of antibiotic resistance through disinfectant usage during the pandemic and highlight the necessity to minimize the environmental release of disinfectants into the non-target environment for combating antibiotic resistance.

Project description:Extensive use of colistin in food animals is deemed a major driving force for the emergence and transmission of mcr-1 However, a non-colistin usage factor(s) contributing to mobile colistin resistance may also exist in animal production systems. Given that polymyxin, a bacterium-derived peptide antibiotic, has been successfully used as a surrogate to study bacterial resistance to antimicrobial peptides (AMPs), acquisition of MCR-1 may confer cross-resistance to the unrelated AMPs implicated in practical applications. To test this, we first constructed Escherichia coli recombinant strains differing only in the presence or absence of functional MCR-1. Among diverse tested AMPs, MCR-1 was observed to confer cross-resistance to bacitracin, an in-feed antibiotic widely used in animal industry. The significantly (2-fold) increased bacitracin MIC was confirmed by using different bacitracin products, broth media, and laboratory host strains for susceptibility tests. Subsequently, an original mcr-1 gene-bearing plasmid, pSLy21, was conjugatively transferred to eight clinical E. coli recipient strains isolated from diarrheic pigs, which also led to significantly increased MICs of both colistin (4-fold to 8-fold) and bacitracin (2-fold). Growth curve examination further demonstrated that MCR-1 provides a growth advantage to various E. coli strains in the presence of bacitracin. Given that bacitracin, a feed additive displaying low absorption in the intestine, can be used in food animals with no withdrawal required, imprudent use of bacitracin in food animals may serve as a risk factor to enhance the ecological fitness of MCR-1-positive E. coli strains, consequently facilitating the persistence and transmission of plasmid-mediated colistin resistance in agricultural ecosystem.IMPORTANCE Polymyxins (e.g., colistin) are the drugs of last resort to treat multidrug-resistant infections in humans. To control mobile colistin resistance, there is a worldwide trend to limit colistin use in animal production. However, simply limiting colistin use in animal production may still not effectively mitigate colistin resistance due to an overlooked non-colistin usage factor(s). Using controlled systems, in this study, we observed that MCR-1 confers cross-resistance to bacitracin, a popular in-feed antibiotic used in food animals. Thus, imprudent and extensive usage of bacitracin in food animals may serve as a non-colistin usage risk factor for the transmissible colistin resistance. Further comprehensive in vitro and in vivo studies are highly warranted to generate science-based information for risk assessment and risk management of colistin resistance, consequently facilitating the development of proactive and effective strategies to mitigate colistin resistance in animal production system and protect public health.

Project description:Pharmaceutical products that are contaminated with Burkholderia cepacia complex (BCC) bacteria may pose serious consequences to vulnerable patients. Benzyldimethylalkylammonium chloride (BZK) cationic surfactants are extensively used in medical applications and have been implicated in the coselection of antimicrobial resistance. The ability of BCC to degrade BZK, tetradecyldimethylbenzylammonium chloride (C14BDMA-Cl), dodecyldimethylbenzylammonium chloride (C12BDMA-Cl), decyldimethylbenzylammonium chloride (C10BDMA-Cl), hexyldimethylbenzylammonium chloride, and benzyltrimethylammonium chloride was determined by incubation in 1/10-diluted tryptic soy broth (TSB) to determine if BCC bacteria have the ability to survive and inactivate these disinfectants. With BZK, C14BDMA-Cl, and C12BDMA-Cl, inhibition of the growth of 20 BCC strains was observed in disinfectant solutions that ranged from 64 to 256 µg/ml. The efflux pump inhibitor carbonyl cyanide m-chlorophenylhydrazone increased the sensitivity of bacteria to 64 µg/ml BZK. The 20 BCC strains grew well in 1/10-diluted TSB medium with BZK, C12BDMA-Cl, and C10BDMA-Cl; they absorbed and degraded the compounds in 7 days. Formation of benzyldimethylamine and benzylmethylamine as the initial metabolites suggested that the cleavage of the C alkyl-N bond occurred as the first step of BZK degradation by BCC bacteria. Proteomic data confirmed the observed efflux activity and metabolic inactivation via biodegradation in terms of BZK resistance of BCC bacteria, which suggests that the two main resistance mechanisms are intrinsic and widespread.ImportanceBenzyldimethylalkylammonium chloride is commonly used as an antiseptic in the United States. Several recent microbial outbreaks were linked to antiseptics that were found to contain strains of the Burkholderia cepacia complex. Burkholderia species survived in antiseptics, possibly because of the degradation of antiseptic molecules or regulation of relevant gene expression. In this study, we assessed the efflux pump and the potential of B. cepacia complex bacteria to degrade benzyldimethylalkylammonium chloride and improved our understanding of the resistance mechanisms, by using proteomic and metabolic information. To our knowledge, this is the first systematic report of the intrinsic mechanisms of B. cepacia complex strain resistance to benzyldimethylalkylammonium chloride, based on the metabolic and proteomic evidence for efflux pumps and the complete biodegradation of benzyldimethylalkylammonium chloride.

Project description:BackgroundBiocide 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.MethodsThe 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.FindingsIn 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.InterpretationOur 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.FundingThis work was funded by the National Health and Medical Research Council of Australia. The funders had no role in study design, interpretation or decision to publish.

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Widely Used Benzalkonium Chloride Disinfectants Can Promote Antibiotic Resistance.

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Widely Used Benzalkonium Chloride Disinfectants Can Promote Antibiotic Resistance.

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