Metabolic adaptations in antibiotic-resistant <i>Staphylococcus aureus</i> : understanding resistance mechanisms and enhancing antibiotic efficacy

ABSTRACT The global rise of antibiotic-resistant Staphylococcus aureus represents a serious clinical challenge, contributing substantially to the burden of antimicrobial resistance. This study explores the metabolic mechanisms of antibiotic-resistant S. aureus . Using LC-MS/MS metabolomics, we analyzed laboratory-evolved strains derived from the Newman strain: ciprofloxacin-resistant (R-CIP), cefazolin-resistant (R-CEF), and dual-resistant (R-CC). Metabolomic profiling revealed 41, 57, and 74 differentially abundant metabolites in R-CIP, R-CEF, and R-CC, respectively, with 21 common to all strains. Pathway analysis identified 12, 9, and 10 significantly perturbed pathways in R-CIP, R-CEF, and R-CC, respectively. Three pathways—glycine, serine, and threonine metabolism; alanine, aspartate, and glutamate metabolism; and taurine and hypotaurine metabolism—were consistently dysregulated across all resistant strains. R-CIP uniquely altered butanoate metabolism, while R-CC specifically changed glutamine and purine metabolism. iPath analysis further highlighted suppressed central carbon and energy metabolism as a shared feature of all resistant strains, accompanied by reduced pyruvate dehydrogenase and TCA cycle enzyme activities, as well as decreased cellular NADH, ATP, and reactive oxygen species levels. Supplementation with pyruvate, citrate, or fumarate significantly enhanced the bactericidal effects of ciprofloxacin and cefazolin. These findings demonstrate that S. aureus acquires resistance to ciprofloxacin and cefazolin through downregulation of central carbon and energy metabolism. Exogenous metabolites can potentiate antibiotic efficacy, offering insights into metabolism-targeted strategies to combat antibiotic-resistant S. aureus . IMPORTANCE The global rise of antibiotic-resistant Staphylococcus aureus poses a significant threat to public health, highlighting the urgent need for novel therapeutic strategies. Current understanding of antibiotic resistance mechanisms, particularly at the metabolic level, remains incomplete. This study investigates the metabolic alterations of resistance in S. aureus by analyzing laboratory-evolved strains resistant to ciprofloxacin (R-CIP), cefazolin (R-CEF), and both (R-CC). Through metabolomic profiling, we identified suppressed central carbon and energy metabolism as a shared resistance mechanism, with strain-specific adaptations further complicating the resistance landscape. Notably, supplementation with key metabolites like pyruvate, citrate, and fumarate enhanced antibiotic efficacy, suggesting a promising strategy to combat resistance. These findings provide critical insights into the metabolic vulnerabilities of resistant S. aureus , offering a novel avenue for the development of metabolism-targeted therapies to address the growing challenge of antibiotic resistance.