Commonly used non-antibiotic medications promote mutation frequency and antimicrobial resistance in Escherichia coli

Abstract Antimicrobial resistance (AMR) poses a global threat to public health. The excessive use of antibiotics significantly contributed to the rise of resistance. Recent evidence suggests that non-antibiotic medications (NAMs) also play a role in antimicrobial resistance development, although this aspect remains less explored and understood. This issue is particularly relevant in residential aged care facilities (RACFs) where both NAMs and antibiotics are frequently used, and AMR is prevalent. We investigated the propensity of NAMs that are commonly used in RACFs and contribute to polypharmacy including non-steroidal anti-inflammatory drugs (NSAIDs) such as ibuprofen and diclofenac, acetaminophen (antipyretic), metformin (glucose-lowering medication), atorvastatin (lipid-lowering agent), tramadol (opioid analgesic), temazepam (hypnotic), and pseudoephedrine (sympathomimetic) to promote bacterial antibiotic resistance by increasing the acquisition of mutations. Escherichia coli was exposed to different NAMs at their gut concentration, combined with ciprofloxacin and the mutation frequency was determined. Additionally, we explored the simultaneous effect of two NAMs as a starting point for studying polypharmacy. Ibuprofen and acetaminophen significantly increased mutation frequency, and conferred high levels of ciprofloxacin resistance, especially when E. coli was exposed to two NAMs. Whole genome sequencing revealed that these changes correlated with mutations in DNA gyrase GyrA, the multiple antibiotic resistance regulator, MarR, and the drug efflux pump expression suppressor, AcrR. Consequently, an increase in transcription of the acrA gene from the AcrAB-TolC drug efflux pump was observed. The combination of two NAMs increased the mutation rate. These multiple mutations caused the higher levels of ciprofloxacin resistance that were observed. Given the risk of polypharmacy to induce AMR, and the results observed in this study, the assessment NAMs in their ability to promote bacterial resistance warrants special attention in future studies into prescribing practices.