Proteomic comparisons of opaque and transparent variants of <i>Streptococcus pneumoniae</i> by two dimensional-differential gel electrophoresis

Streptococcus pneumoniae (the pneumococcus) is a human pathogen, accounting for massive global morbidity and mortality. Although asymptomatic colonization of the nasopharynx almost invariably precedes disease, the critical determinants enabling pneumococcal progression from this niche to cause invasive disease are poorly understood. One mechanism proposed to be central to this transition involves opacity phase variation, whereby pneumococci harvested from the nasopharynx are typically transparent, while those simultaneously harvested from the blood are opaque. Here, we used two dimensional-differential gel electrophoresis (2D-DIGE) to compare protein expression profiles of transparent and opaque variants of 3 pneumococcal strains, D39 (serotype 2), WCH43 (serotype 4) and WCH16 (serotype 6A) in vitro. One spot comprising a mixture of capsular polysaccharide biosynthesis protein and other proteins was significantly up-regulated in the opaque phenotype in all 3 strains; other proteins were differentially regulated in a strain-specific manner. We conclude that pneumococcal phase variation is a complex and multifactorial process leading to strain-specific pathogenicity.Melissa H. Chai, Florian Weiland, Richard M. Harvey, Peter Hoffmann, Abiodun D. Ogunniyi, James C. Pato

Interplay between manganese and iron in pneumococcal pathogenesis: role of the orphan response regulator RitR

Streptococcus pneumoniae (the pneumococcus) is a major human pathogen that is carried asymptomatically in the nasopharynx by up to 70% of the human population. Translocation of the bacteria into internal sites can cause a range of diseases, such as pneumonia, otitis media, meningitis, and bacteremia. This transition from nasopharynx to growth at systemic sites means that the pneumococcus needs to adjust to a variety of environmental conditions, including transition metal ion availability. Although it is an important nutrient, iron potentiates oxidative stress, and it is established that in S. pneumoniae, expression of iron transport systems and proteins that protect against oxidative stress are regulated by an orphan response regulator, RitR. In this study, we investigated the effect of iron and manganese ion availability on the growth of a ritR mutant. Deletion of ritR led to impaired growth of bacteria in high-iron medium, but this phenotype could be suppressed with the addition of manganese. Measurement of metal ion accumulation indicated that manganese prevents iron accumulation. Furthermore, the addition of manganese also led to a reduction in the amount of hydrogen peroxide produced by bacterial cells. Studies of virulence in a murine model of infection indicated that RitR was not essential for pneumococcal survival and suggested that derepression of iron uptake systems may enhance the survival of pneumococci in some niches

SpxA1 Involved in Hydrogen Peroxide Production, Stress Tolerance and Endocarditis Virulence in Streptococcus sanguinis

Streptococcus sanguinis is one of the most common agents of infective endocarditis. Spx proteins are a group of global regulators that negatively or positively control global transcription initiation. In this study, we characterized the spxA1 gene in S. sanguinis SK36. The spxA1 null mutant displayed opaque colony morphology, reduced hydrogen peroxide (H2O2) production, and reduced antagonistic activity against Streptococcus mutans UA159 relative to the wild type strain. The ΔspxA1 mutant also demonstrated decreased tolerance to high temperature, acidic and oxidative stresses. Further analysis revealed that ΔspxA1 also exhibited a ∼5-fold reduction in competitiveness in an animal model of endocarditis. Microarray studies indicated that expression of several oxidative stress genes was downregulated in the ΔspxA1 mutant. The expression of spxB and nox was significantly decreased in the ΔspxA1 mutant compared with the wild type. These results indicate that spxA1 plays a major role in H2O2 production, stress tolerance and endocarditis virulence in S. sanguinis SK36. The second spx gene, spxA2, was also found in S. sanguinis SK36. The spxA2 null mutant was found to be defective for growth under normal conditions and showed sensitivity to high temperature, acidic and oxidative stresses

Hydrogen peroxide release by bacteria suppresses inflammasome-dependent innate immunity

Hydrogen peroxide (H2O2) has a major function in host-microbial interactions. Although most studies have focused on the endogenous H2O2 produced by immune cells to kill microbes, bacteria can also produce H2O2. How microbial H2O2 influences the dynamics of host-microbial interactions is unclear. Here we show that H2O2 released by Streptococcus pneumoniae inhibits inflammasomes, key components of the innate immune system, contributing to the pathogen colonization of the host. We also show that the oral commensal H2O2-producing bacteria Streptococcus oralis can block inflammasome activation. This study uncovers an unexpected role of H2O2 in immune suppression and demonstrates how, through this mechanism, bacteria might restrain the immune system to co-exist with the host