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 H2O2 to protect cells from oxidative stress damage. Here, we identified and characterized the GpoA glutathione peroxidase of Streptococcus pneumoniae, one of the most important human bacterial pathogens. We demonstrated that recombinant GpoA exhibits glutathione peroxidase activity and that the Cys36 residue is essential for this function. We conducted a comparative analysis of the phenotypic characteristics linked to GpoA and those linked to other peroxidases, including the TpxD thiol peroxidase and the AhpD alkylhydroperoxidase. We analyzed a mutant strain lacking the gshT gene, which encodes the GSH-binding protein component of the glutathione ABC transporter 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 wild-type strain. These results indicate that GpoA, TpxD, and AhpD constitute a robust H2O2 detoxification system. Our findings further demonstrate that GpoA contributes to levofloxacin persistence under oxidative stress conditions, both in H2O2-treated cultures and during intracellular infection. Similar phenotypic patterns were observed in the ΔtpxD and 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 the wild-type strain, suggesting that GpoA contributes to pneumococcal pathogenesis. This study demonstrates that GpoA 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.