Involvement of NADH Oxidase in Competition and Endocarditis Virulence in Streptococcus sanguinis

Abstract

c Here, we report for the first time that the Streptococcus sanguinis nox gene encoding NADH oxidase is involved in both competition with Streptococcus mutans and virulence for infective endocarditis. An S. sanguinis nox mutant was found to fail to inhibit the growth of Streptococcus mutans under microaerobic conditions. In the presence of oxygen, the recombinant Nox protein of S. sanguinis could reduce oxygen to water and oxidize NADH to NAD ؉ . The oxidation of NADH to NAD ؉ was diminished in the nox mutant. The nox mutant exhibited decreased levels of extracellular H 2 O 2 ; however, the intracellular level of H 2 O 2 in the mutant was increased. Furthermore, the virulence of the nox mutant was attenuated in a rabbit endocarditis model. The nox mutant also was shown to be more sensitive to blood killing, oxidative and acid stresses, and reduced growth in serum. Thus, NADH oxidase contributes to multiple phenotypes related to competitiveness in the oral cavity and systemic virulence.

Contribution of NADH oxidase to oxidative stress tolerance and virulence ofStreptococcus suisserotype 2

“…NADH oxidase contributes to the pathogenesis of S. pneumoniae, Group B Streptococcus and Streptococcus sanguinis. 12,13,15,29 In this study, we demonstrated that the virulence of S. suis 2 in a murine infection model was completely abolished by deletion of nox. The survival rates were significantly higher in mice infected with the Dnox mutant than in those infected with the WT and CDnox strains.…”

“…This is not surprising, as S. suis NADH oxidase shows considerable identity to its orthologs from various streptococcal species, which have been shown to be involved in oxidative stress response and virulence. [12][13][14][15]29 The Dnox mutant exhibits reduced resistance to oxidative stress generated by environmental oxygen and hydrogen peroxide. This result is in agreement with the observed effects of spxA1 deletion in S. suis and nox2 deletion in Group B Streptococcus.…”

“…NADH oxidase activity was measured by monitoring the decrease in NADH absorbance (A 340 ) at 25 C, as described previously. 29,47 The reaction mixture contained 50 mM potassium phosphate buffer (pH 7.0), 0.2 mM EDTA, 0.2 mM NADH, and purified NADH oxidase. One unit of enzyme activity corresponds to the oxidation of 1 mmol of NADH per min at 25 C.…”

“…11 These included nox (SSUSC84_0648, encoding NADH oxidase), tpx (SSUSC84_1246, encoding thiol peroxidase), copA (SSUSC84_1247, encoding copper-transporting ATPase), and sodA (SSUSC84_1386, encoding superoxide dismutase). 11 Genes nox, tpx and sodA have been well studied and reported to be involved in oxidative stress responses and/or virulence in various streptococci and other bacteria, [12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29] while copA has been shown to be implicated in copper resistance. [30][31][32] Additionally, an undefined gene, 0350 (SSUSC84_0350, encoding a hypothetical protein) is potentially involved in oxidative stress and/or virulence, as it was downregulated 8.7-fold in DspxA1.…”

Genetics ofsanguinis-Group Streptococci in Health and Disease

“…Mutations in the genes encoding lipoprotein SsaB (a putative manganese transport protein) (38), spxA1 (encoding a global regulator involved in H2O2 production) (138) and nox (encoding an NADH oxidase that also influences H2O2 production) (139), together with the anaerobic ribonucleotide reductase already mentioned, have all been shown to impair the capacity of S. sanguinis to cause IE. This suggests that the ability of S. sanguinis to adapt to differences in oxygen tension and the production of and resistance to reactive oxygen species are important in the development of IE, although molecular details have yet to be explained.…”