Butyrate synergizes with glucose to promote anaerobic growth of Staphylococcus aureus via anaplerotic metabolism and stress response pathways

Short-chain fatty acids (SCFAs) like butyrate and propionate are abundant microbiota-derived metabolites that influence bacterial physiology in host-associated niches such as the gastrointestinal tract. However, their effects on Staphylococcus aureus under varying nutritional conditions remain incompletely understood. Here we investigated how SCFAs interact with glucose or galactose to regulate anaerobic growth, biofilm formation, and global transcription in S. aureus. Both SCFAs inhibit growth in a dose-dependent manner. Biofilm formation was differentially affected, with butyrate promoting and propionate suppressing biofilm formation. Glucose and galactose alleviated SCFA-mediated growth inhibition, with glucose exerting the strongest effect. Notably, glucose enhanced butyrate-associated growth and biofilm formation beyond glucose alone, whereas galactose produced more modest effects. Enzymatic and genetic analyses indicated that SCFA-sugar biofilms contain proteins and extracellular DNA and involve VraSR-dependent regulation. Transcriptomic profiling revealed broad metabolic reprogramming, including induction of urease genes, amino acid biosynthesis, and stress response pathways. Synergistic effects between butyrate and glucose were partially dependent on anaplerotic metabolism via pyruvate carboxylase, linking the TCA cycle to SCFA adaptation. Together these findings demonstrate that the nutritional environment dictates whether SCFAs impair S. aureus growth or reprogram its physiology, promoting metabolic adaptation and biofilm formation under sugar-replete conditions.