Molecular mechanism of antibiotic resistance induced by mono- and twin-chained quaternary ammonium compounds

The usage of quaternary ammonium compounds (QACs) as disinfectants has increased dramatically since the outbreak of COVID-19 pandemic, leading to potentially accelerated emergence of antibiotic resistance. Long-term exposure to subinhibitory level QACs can lead to multidrug resistance, but the contribution of mutagenesis to resistance evolution is obscure. In this study, we subcultured E. coli K-12 under subinhibitory (0.25 x and 0.5 x Minimum Inhibitory Concentration, MIC) or inhibitory (1 x and 2 x MIC) concentrations of benzalkonium chloride (BAC, mono-chained) or didecyldimethylammonium chloride (DDAC, twin-chained) for 60 days. The sensitivity of QAC-adapted cells to five typical antibiotics decreased significantly, and in particular, the MIC of rifampicin increased by 85 times. E. coli adapted faster to BAC but developed 20-167% higher antibiotic resistance with 56% more mutations under DDAC exposure. The broader mutations induced by QACs, including negative regulators (acrR , marR , soxR , and crp), outer membrane proteins and transporters (mipA and sbmA), and RNA polymerase (rpoB and rpoC), potentially contributed to the high multi-drug resistance. After QACs stresses were removed, the phenotypic resistance induced by subinhibitory concentrations of QACs was reversible, whereas that induced by inhibitory concentrations of QACs was irreversible. The different patterns and molecular mechanism of antibiotic resistance induced by BAC and DDAC is informative to estimating the risks of broader QACs present at varied concentrations in the environment.

Automated summary

The usage of quaternary ammonium compounds (QACs) as disinfectants has increased dramatically during the COVID-19 pandemic, potentially leading to accelerated antibiotic resistance. Long-term exposure to subinhibitory level QACs can lead to multidrug resistance, but the role of mutagenesis in resistance evolution is not clear. This study found that E. coli adapted differently to benzalkonium chloride (BAC) and didecyldimethylammonium chloride (DDAC), with DDAC exposure resulting in higher antibiotic resistance and more mutations. The mutations induced by QACs potentially contributed to high multi-drug resistance. The reversible and irreversible phenotypic resistance induced by subinhibitory and inhibitory concentrations of QACs, respectively, highlight the importance of understanding the mechanisms and risks of different QACs in the environment.