Streptococcus sanguinis induces neutrophil cell death by production of hydrogen peroxide

Streptococcus is the dominant bacterial genus in the human oral cavity and a leading cause of infective endocarditis. Streptococcus sanguinis belongs to the mitis group of streptococci and produces hydrogen peroxide (H 2 O 2 ) by the action of SpxB, a pyruvate oxidase. In this study, we investigated the involvement of SpxB in survival of S . sanguinis in human blood and whether bacterial H 2 O 2 exhibits cytotoxicity against human neutrophils. Results of a bactericidal test with human whole blood revealed that the spxB mutation in S . sanguinis is detrimental to its survival in blood. When S . sanguinis strains were exposed to isolated neutrophils, the bacterial survival rate was significantly decreased by spxB deletion. Furthermore, human neutrophils exposed to the S . sanguinis wild-type strain, in contrast to those exposed to an spxB mutant strain, underwent cell death with chromatin de-condensation and release of web-like extracellular DNA, reflecting induction of neutrophil extracellular traps (NETs). Since reactive oxygen species-mediated NET induction requires citrullination of arginine residues in histone proteins and subsequent chromatin de-condensation, we examined citrullination levels of histone in infected neutrophils. It is important to note that the citrullinated histone H3 was readily detected in neutrophils infected with the wild-type strain, as compared to infection with the spxB mutant strain. Moreover, decomposition of streptococcal H 2 O 2 with catalase reduced NET induction. These results suggest that H 2 O 2 produced by S . sanguinis provokes cell death of neutrophils and NET formation, thus potentially affecting bacterial survival in the bloodstream.