The spread of antibiotic resistance in health care settings and in the water environment is a formidable human health concern that could render antibiotics useless against infection. Disinfectants applied in (waste)water treatment and healthcare settings can effectively inactivate antibiotic resistant bacteria (ARB), thereby preventing vertical gene transfer and conjugation. However, genes derived from dead cells have the potential to remain biologically active and, through natural transformation, confer antibiotic resistance to nonresistant bacteria. Therefore, to ensure resistance is not spread to downstream bacteria, the transformation activity of antibiotic resistance genes (ARGs) should also be deactivated. However, little is known about the effectiveness of disinfectants to destroy ARGs. This study evaluates the ability of ozone (widely used in water treatment) and hydrogen peroxide (commonly applied in healthcare settings) to inactivate vegetative cells of a multidrug-resistant ARB strain, Bacillus subtilis 1A189, and destroy the ARG encoding its resistance traits. ARG deactivation in both intracellular and extracellular forms was assessed with a culture-based transformation assay. Double-stranded DNA concentration, DNA size distribution, mononucleotide concentrations, and gene copy number were measured using fluorescence, pulsed-field gel electrophoresis, HPLC, and quantitative polymerase chain reaction (qPCR), respectively. These chemical analyses were used to determine damage mechanisms and evaluated as potential surrogates for analysis of ARG transformation activity. At disinfectant exposures (CTs) relevant to drinking water treatment for ozone (�1 mg/L*min) and healthcare settings for hydrogen peroxide (�30,000 mg/L*min), each yielded �6 log inactivation of B. subtilis 1A189 cells and =3.5 log elimination of ARG transforming activity. The primary DNA damage mechanisms for ozone and hydrogen peroxide appeared to be nucleobase degradation and DNA strand scission, respectively. qPCR showed promise as a surrogate for transformation activity when DNA was exposed to ozone but less so with hydrogen peroxide, due to differences in the DNA damage mechanisms for each disinfectant.

Deactivation of antibiotic resistance genes with ozone and hydrogen peroxide

248th National Meeting (2014)

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