GpoA glutathione peroxidase links oxidative stress response, antibiotic persistence, and virulence in Streptococcus pneumoniae

Glutathione peroxidases are conserved enzymes found in prokaryotic and eukaryotic organisms that reduce H2O2to protect cells from oxidative stress damage. Here, we identified and characterized the GpoA glutathioneperoxidase of Streptococcus pneumoniae, one of the most important human bacterial pathogens. We demonstratedthat recombinant GpoA exhibits glutathione peroxidase activity and that the Cys36 residue is essential for thisfunction. We conducted a comparative analysis of the phenotypic characteristics linked to GpoA and those linkedto other peroxidases, including the TpxD thiol peroxidase and the AhpD alkylhydroperoxidase. We analyzed amutant strain lacking the gshT gene, which encodes the GSH-binding protein component of the glutathione ABCtransporter complex. The ΔgpoA, ΔtpxD, ΔahpD, and ΔgshT mutants exhibited increased susceptibility to H2O2,and also impaired intracellular survival in pneumocytes, macrophages, and neutrophils compared to the wildtypestrain.These results indicate that GpoA, TpxD, and AhpD constitute a robust H2O2 detoxification system. Our findingsfurther demonstrate that GpoA contributes to levofloxacin persistence under oxidative stress conditions, both inH2O2-treated cultures and during intracellular infection. Similar phenotypic patterns were observed in the ΔtpxDand ΔahpD mutants, indicating that antibiotic persistence depends on the oxidative stress response.In a murine model, the ΔgpoA mutant exhibited diminished survival across multiple organs relative to thewild-type strain, suggesting that GpoA contributes to pneumococcal pathogenesis. This study demonstrates thatGpoA functions as an efficient antioxidant enzyme integral to the oxidative stress response in S. pneumoniae,facilitating the bacterium's survival both extracellularly and intracellularly under oxidative stress conditions. Fil: Zappia, Victoria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Hernandez Morfa, Mirelys. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina