Clinically relevant mutations in regulatory regions of metabolic genes facilitate early adaptation to ciprofloxacin in Escherichia coli

The genomic landscape associated with early adaptation to ciprofloxacin is poorly understood. Although the interplay between core metabolism and antimicrobial resistance is being increasingly recognized, mutations in metabolic genes and their biological role remain elusive. Here, we exposed Escherichia coli to increasing gradients of ciprofloxacin with intermittent transfer-bottlenecking and identified mutations in three non-canonical targets linked to metabolism including a deletion (tRNA-ArgΔ414-bp) and point mutations in the regulatory regions of argI (ARG box) and narU. Our findings suggest that these mutations modulate arginine and carbohydrate metabolism, facilitate anaerobiosis and increased ATP production during ciprofloxacin stress. Furthermore, mutations in the regulatory regions of argI and narU were detected in over 70% of sequences from clinical E. coli isolates and were overrepresented among ciprofloxacin-resistant isolates. In sum, we have identified clinically relevant mutations in the regulatory regions of metabolic genes as a central theme that drives physiological changes necessary for adaptation to ciprofloxacin stress.