Reduction and inactivation of microorganisms with tolerance to quaternary ammonium compounds by chlorine dioxide

Introduction: Quaternary ammonium compounds (QACs) are globally used as active substances in bactericides. Since QACs are known to cause bacterial tolerances associated with increased tolerances to antibiotics, concerns about human health risks by QAC-tolerant bacteria are increasing. Once adapted to QAC, tolerant microorganisms can persist in tap water despite ongoing disinfection with QACs. For the inactivation of potential problematic QAC-tolerant microorganisms from tap water, the impact of chlorine dioxide as an alternative biocide was tested.

Goal: The aim of this study was to evaluate the susceptibility of QAC-tolerant gram-positive and gram-negative microorganisms to the oxidative disinfectant chlorine dioxide.

Material & Methods: The gram-negatives Escherichia coli, Pseudomonas aeruginosa and Stenotrophomonas maltophilia as well as the gram-positives Staphylococcus aureus und S. epidermidis were successively adapted to tolerance to benzalkonium chloride (BAC) representative for QACs by repeated inoculation and regrowth. Analyzing the susceptibility to oxidative disinfection, BAC tolerant bacteria were treated with chlorine dioxide at low (0.4 mg/L) and increased concentrations (1 mg/L) for 25 min followed by plating to determine the inactivation rates.

Results: The adaption to BAC resulted in a fast occurring tolerance, which was stable over weeks. The phenotype of BAC tolerance in tested bacteria remained even without BAC supplementation. The bacterial biofilm formation increased with the adaption to BAC. An exposure to 1 mg/L chlorine dioxide was adequate to remove densities of 10⁷ cells of E. coli, S. maltophilia, S. aureus and S. epidermidis. In contrast, treatment with 0.4 ml/L chlorine dioxide limited the bacterial inactivation.

Conclusion: BAC-tolerant bacteria demonstrated challenges towards a successful disinfection. Regarding a high bacterial load of BAC-tolerant bacteria, an increased concentration of chlorine dioxide inactivated the microorganisms successfully. A regular change of disinfectants with different modes of action is recommended to avoid formation of tolerances and resistances.