The glutathione pathway is required for biofilm formation in Acinetobacter baumannii

Acinetobacter baumannii is a Gram-negative nosocomial bacterium that is a member of the ESKAPE group of pathogens, notable for its virulence and intrinsic antibiotic resistance. It causes diverse infections, including respiratory and soft tissue disease, that are increasingly difficult to treat. Glutathione (GSH), the major intracellular redox buffer, is known in other bacteria to protect against stress and influence physiological processes such as biofilm formation. To investigate the roles of GSH in A. baumannii , we analyzed transposon mutant strains lacking gshA and gshB genes , which encode the enzymes that catalyze the two steps of GSH biosynthesis. Both mutant strains failed to produce GSH, exhibited impaired growth, and were hypersensitive to oxidative stress, nitrosative stress, toxins, and ferric chloride, compared to the wildtype strain. They also showed pronounced defects in biofilm formation and motility. Transcriptomic analysis of the gshA mutant strain relative to the wildtype strain revealed upregulation of genes involved in phenylacetate degradation and fimbrial biogenesis, while genes involved in iron and sulfur uptake and metabolism were downregulated. Moreover, mutant strains lacking GSH-dependent S-nitrosoglutathione reductases (GSNORs) displayed similar biofilm and nitrosative stress defects. Together, these findings demonstrate that GSH and GSNORs play central roles in stress resistance, biofilm development, and metabolic regulation in A. baumannii , highlighting their importance in the physiology and pathogenesis of this clinically relevant pathogen.