Contributions of Pneumolysin, Pneumococcal Surface Protein A (PspA), and PspC to Pathogenicity ofStreptococcus pneumoniaeD39 in a Mouse Model

Abstract

Successful colonization of the upper respiratory tract by Streptococcus pneumoniae is an essential first step in the pathogenesis of pneumococcal disease. However, the bacterial and host factors that provoke the progression from asymptomatic colonization to invasive disease are yet to be fully defined. In this study, we investigated the effects of single and combined mutations in genes encoding pneumolysin (Ply), pneumococcal surface protein A (PspA), and pneumococcal surface protein C (PspC, also known as choline-binding protein A) on the pathogenicity of Streptococcus pneumoniae serotype 2 (D39) in mice. Following intranasal challenge with D39, stable colonization of the nasopharynx was maintained over a 7-day period at a level of approximately 10 5 bacteria per mouse. The abilities of the mutant deficient in PspA to colonize the nasopharynx and to cause lung infection and bacteremia were significantly reduced. Likewise, the PspC mutant and, to a lesser extent, the Ply mutant also had reduced abilities to colonize the nasopharynx. As expected, the double mutants colonized less well than the parent to various degrees and had difficulty translocating to the lungs and blood. A significant additive attenuation was observed for the double and triple mutants in pneumonia and systemic disease models. Surprisingly, the colonization profile of the derivative lacking all three proteins was similar to that of the wild type, indicating virulence gene compensation. These findings further demonstrate that the mechanism of pneumococcal pathogenesis is highly complex and multifactorial but ascribes a role for each of these virulence proteins, alone or in combination, in the process.

Pneumococcal Interactions with Epithelial Cells Are Crucial for Optimal Biofilm Formation and Colonization In Vitro and In Vivo

“…Similarly, we found that deletion of PspA, which protects against complement attack in the bloodstream but shows no role in early colonization (4), had no effect on biofilm formation over 48 h, whereas PspC-, autolysin-, and pneumolysin-negative strains, which colonize poorly in vivo (4,8,53,71), formed lessstructured biofilms on epithelial cells than the wild-type strain. By quantifying biomass and biofilm-specific resistance to antibiotics after biofilm formation over a prefixed epithelial substratum, we could accurately predict virulence phenotypes using our in vitro model, whereas no difference was observed when biofilms were formed on abiotic surfaces.…”

“…To further validate the important role for epithelial-bacterial cell interactions in biofilm formation and the usefulness of biofilm formation on epithelial surfaces to predict colonization efficiency, we used the wild-type strain D39 and deletion mutants lacking each of five virulence-associated molecules (autolysin, pneumolysin, PspC, PspA, and capsule). The D39 strain background rather than the EF3030 strain background was chosen both because these strains have been used previously in biofilm experiments on abiotic surfaces, which would allow comparisons with previous literature in the field, and because they are well studied for their ability to interact with host structures and impact colonization or invasive disease (4,8,9,28,53). The strains were grown on glass or prefixed NCI-H292 epithelial cells for various times and evaluated by SEM, as well as for biomass and resistance to gentamicin.…”

Identification of a novel pneumococcal vaccine antigen preferentially expressed during meningitis in mice

“…Other single mutants were not significantly attenuated in either challenge model (Figure 2, A and B, and Supplemental Figure 2, A and B). However, we showed previously that while mutagenesis of single mutants may not have a significant impact on pneumococcal virulence, multiple mutations resulted in significant attenuation of virulence (28,29). Therefore, as a result of the marginal attenuation of virulence observed with the loss of expression of the single genes α-glycerophosphate oxidase (GlpO), PTS system, galactitol-specific IIA component (ptsGal), and voltage-gated chloride channel family protein (vgcl) (denoted ΔglpO, ΔptsGal, and ΔvgcL mutants, respectively) in the i.n.…”

Pneumococcal Adhesins PavB and PspC Are Important for the Interplay with Human Thrombospondin-1

“…The most promising candidates were pneumococcal surface protein C (PspC) and pneumococcal adherence and virulence factor B (PavB). Both proteins possess repeating structures and were previously shown to bind various human extracellular/matricellular matrix proteins such as fibronectin or Vn (11,31), thereby mediating pneumococcal colonization and invasion of host cells (11,(32)(33)(34)(35)(36).…”