Degradation and Deactivation of Intracellular Bacterial Antibiotic Resistance Genes by Commonly Used Healthcare and Personal Care Disinfectants

Abstract

This work investigated efficacies of commonly used healthcare and personal care disinfectants, including glutaraldehyde, chlorhexidine, ethanol, povidone-iodine, benzalkonium chloride, phenol, free chlorine, hydrogen peroxide (H₂O₂), and 254 nm UV light, in degrading (as measured by qPCR analyses of ∼1000 bp amplicon loss) and deactivating (as measured by transforming activity loss) bacterial antibiotic resistance genes (ARGs) during inactivation of antibiotic-resistant bacteria (ARB) on inanimate surfaces or in aqueous suspension. Intracellular ARGs (iARGs) blt, mecA, and ampC, within vegetative cells of Bacillus subtilis, Staphylococcus aureus, and Pseudomonas aeruginosa, respectively, were treated on PTFE and/or stainless-steel surfaces or in aqueous phosphate buffer (PB; H₂O₂ only), to simulate potential healthcare and personal care cleaning applications under representative disinfectant exposure conditions. No chemical disinfectant yielded more than limited (≤1.9log₁₀) iARG degradation/deactivation under the conditions investigated, even when ARB cells were extensively inactivated (at levels from 3.1log₁₀ to ≥6log₁₀). In contrast, UV irradiation yielded up to ∼2.8-3.2log₁₀ iARG degradation/deactivation at corresponding ARB inactivation levels up to ∼4log₁₀ in the case of the blt gene within B. subtilis cells on PTFE surfaces, though levels of iARG degradation/deactivation and ARB inactivation were generally lower than expected based on prior aqueous-phase results, likely due to light-shielding effects at the typical ∼10⁸-10⁹ CFU/mL cell inoculum densities used for surface disinfection tests. During exposure to H₂O₂ in PB, iARG deactivation and ARB inactivation reached up to 1.7log₁₀ and >3.5log₁₀, respectively, while iARG degradation was minimal (≤0.2log₁₀); this appears to be driven by DNA-strand fragmentation (as observed by pulsed-field gel electrophoresis analysis) likely resulting from reaction with endogenous HO^• (or Fe(IV)) generated via intracellular iron-catalyzed H₂O₂ decomposition. While all investigated disinfectants were able to effectively inactivate ARB cells themselves, these results demonstrate that most are ineffective in simultaneously degrading and deactivating iARGs, highlighting the potential benefits of employing disinfectants such as 254 nm UV light, that selectively target bacterial DNA, to improve mitigation of antibiotic resistance dissemination.

Keywords: ARB; ARGs; disinfection; environmental surfaces; natural transformation; pathogens.