Disinfectants such as bleach, acids, phenols, and alcohols are important weapons against infectious diseases. During the COVID-19 pandemic, as the coronavirus ran rampant across the world, experts advised disinfecting frequently touched surfaces to prevent viral transmission. Despite this, scientists do not completely understand how these compounds kill microbes or how microbes become tolerant to them.
Now, a research team led by Xilin Zhao at Xiamen University has found that a mutation in an enzyme in Escherichia coli helps the bacterium survive exposure to disinfectants.1 The results, published in Proceedings of the National Academy of Sciences, provide insights into how bacteria develop tolerance to disinfectants and pinpoint genes that can be surveyed for disinfectant tolerance in pathogens.
The researchers started out by using a strain of E. coli engineered to carry a mutation in a gene involved in the carbohydrate phosphotransferase system (PTS), which causes the bacterium to exhibit tolerance to some disinfectants, including phenols.2 Although an initial exposure to phenol wiped out many of the ptsI mutants, some of the bacteria survived subsequent treatment.
Sequencing the genome of the phenol-tolerant bacteria revealed a mutation in a subunit of the enzyme phenylalanine-tRNA synthetase (PheS), which is essential for protein synthesis. This mutation caused the amino acid phenylalanine to be replaced by cysteine at the 158th position in the protein (F158C). Introducing the pheS F158C mutation to wild type E. coli using CRISPR/Cas9 editing tools rendered them tolerant to several other disinfectants.
When Zhao and his team replaced phenylalanine with other amino acids besides cysteine, they found that the mutants still tolerated phenol, suggesting that the trait depends on the specific position rather than the amino acids.
The researchers speculated that a mutation at the 158th position may reduce PheS’s ability to recognize its substrate, phenylalanine. This would mimic conditions of phenylalanine starvation, which can trigger the accumulation of alarmones, “antideath” signaling molecules produced by bacteria in response to such stress. One alarmone implicated in antimicrobial resistance is guanosine pentaphosphate (ppGpp).3 Introducing the mutation to an E. coli strain that cannot synthesize ppGpp eliminated phenol tolerance, confirming that the mutation acts through the alarmone.
Further experiments revealed that the alarmone triggers a cascade of events that culminates in the transcription of genes that suppress reactive oxygen species (ROS), which trigger bacterial death.4 Consistent with this, Zhao and his team observed that disinfectant exposure caused a surge in E. coli ROS levels and that this surge was reduced in the mutant strain.
Finally, the researchers investigated the clinical significance of their results. When they introduced an F158C mutation in the PheS of pathogenic Klebsiella pneumoniae and enterohemorrhagic E. coli, bacterial strains that commonly cause disease in people, the mutants exhibited tolerance to disinfectants.
Follow-up investigations should examine whether mutations in additional aminoacyl-tRNA synthetases result in similar disinfectant tolerance in bacteria and whether other cellular pathways are involved in conferring this effect, the authors noted.
- Chen M, et al. Broad-spectrum tolerance to disinfectant-mediated bacterial killing due to mutation of the PheS aminoacyl tRNA synthetase. Proc Nat Acad Sci USA. 2025
- Zeng J, et al. A broadly applicable, stress-mediated bacterial death pathway regulated by the phosphotransferase system (PTS) and the cAMP-Crp cascade. Proc Nat Acad Sci USA. 2022;119(23):e2118566119.
- Grucela PK, Zhang YE. Basal level of ppGpp coordinates Escherichia coli cell heterogeneity and ampicillin resistance and persistence. Microb Cell. 2023;10(11):248-260.
- Dapa T, et al. The SOS and RpoS regulons contribute to bacterial cell robustness to genotoxic stress by synergistically regulating DNA polymerase pol II. Genetics. 2017;206(3):1349-1360.
