Reversible decrease in swimming motility accompanies emergent β-lactam antibiotic resistance in Escherichia coli

Antimicrobial resistance (AMR) threatens global health, food security, and development; however, the evolutionary and molecular pathways that drive AMR remain poorly understood, limiting our ability to advance diagnostics and rapidly detect newly emerging resistance. In this report, we employed laboratory evolution of an environmental Escherichia coli strain (NIST 0056) under B-lactam antibiotic pressure (ampicillin and cephalexin) to specifically investigate the relationship between resistance genotype and motility phenotype. Within 10 passages, populations evolved varying levels of B-lactam resistance (including high-level resistance > 32 ug/mL) and showed consistent, dose-dependent reductions in swimming motility. Whole-population genome sequencing revealed large deletions in marR, SNPs in efflux pump gene acrB and fumarate reductase gene frdD, and mutations with evidence of amplification in the chromosomal class C B-lactamase gene ampC, collectively linking these efflux-mediated, B-lactam resistance mechanisms to loss of motility. Notably, motility reductions were more pronounced in individual clones than in the mixed populations (p < 0.005, Mann-Whitney; two-tailed test). Following removal of antibiotic pressure, motility was restored and AMR-associated mutations decreased in frequency, suggesting fitness costs associated with resistance. In aggregate, our results demonstrate that reduced swimming motility accompanies specific efflux-driven, B-lactam resistance mechanisms. Further work will investigate if the behavior is generalizable across flagellated Enterobacteriaceae. Because these rapid changes are amenable to high-throughput measurement, swimming motility may offer a tractable physiological metric for AMR detection in environmental surveillance.