Streptococcus infantis Inhibits Streptococcus sanguinis H₂O₂ Production via Diffusible Signal

Objectives: We previously identified a bacterial consortium of Staphylococcus saprophyticus, Streptococcus infantis, and Streptococcus sanguinis that use regulated H₂O₂ production for oral cavity colonization resistance. S. infantis normally inhibits S. sanguinis’ H₂O₂ production. Detection of an invader, Escherichia coli, triggers signal cascades resulting in de-repression of H₂O₂ production. This study aimed to elucidate the underlying regulatory mechanisms for H₂O₂ production in S. sanguinis.
Methods: Kanamycin-resistant S. sanguinis was grown with S. infantis, and CFU was monitored to determine if S. infantis inhibits S. sanguinis growth. Catalase/peroxidase activity of S. infantis was measured. S. sanguinis, with an spxB promoter-controlled luciferase reporter gene, was used to study the impact of the presence of S. infantis on spxB expression. Proximity inhibition tests between S. sanguinis and S. infantis were performed. Lastly, an S. sanguinis mutant library was screened for mutants whose H₂O₂ production was not inhibited by S. infantis.
Results: Co-cultivation with S. infantis abolished H₂O₂ production in S. sanguinis. Our results showed that S. infantis does not kill S. sanguinis when grown in co-culture. S. infantis tested negative for catalase/peroxidase activity. Furthermore, S. sanguinis bearing an spxB-luc reporter gene displayed greatly reduced luciferase activity in the presence of S. infantis. The proximity inhibition tests revealed a gradient of inhibited H₂O₂ production in S. sanguinis colonies, with the largest reduction in cells closest to, but without physical contact with, S. infantis colonies. The mutant library screen resulted in several S. sanguinis SK36 mutants whose H₂O₂ production was not affected by S. infantis.
Conclusions: Together these data suggest that S. infantis inhibits S. sanguinis H₂O₂ production via diffusible signal(s), which could result in downregulation of spxB gene expression through a yet-to-be determined pathway. The mutant screen has provided us with several candidate genes potentially involved in this pathway.