Immunization with LytB protein of Streptococcus pneumoniae activates complement-mediated phagocytosis and induces protection against pneumonia and sepsis

Abstract

The cell wall glucosaminidase LytB of Streptococcus pneumoniae is a surface exposed protein involved in daughter cell separation, biofilm formation and contributes to different aspects of the pathogenesis process. In this study we have characterized the antibody responses after immunization of mice with LytB in the presence of alhydrogel as an adjuvant. Enzyme-linked immunosorbent assays measuring different subclasses of immunoglobulin G, demonstrated that the antibody responses to LytB were predominantly IgG1 and IgG2b, followed by IgG3 and IgG2a subclasses. Complement-mediated immunity against two different pneumococcal serotypes was investigated using sera from immunized mice. Immunization with LytB increased the recognition of S. pneumoniae by complement components C1q and C3b demonstrating that anti-LytB antibodies trigger activation of the classical pathway. Phagocytosis assays showed that serum containing antibodies to LytB stimulates neutrophil-mediated phagocytosis against S. pneumoniae. Animal models of infection including invasive pneumonia and sepsis were performed with two different clinical isolates. Vaccination with LytB increased bacterial clearance and induced protection demonstrating that LytB might be a good candidate to be considered in a future protein-based vaccine against S. pneumoniae.

Novel Immunoprotective Proteins of Streptococcus pneumoniae Identified by Opsonophagocytosis Killing Screen

“…Further immunoblotting detection yielded 186 antisera that reacted with 119 unique pneumococcal proteins (see Table S2 in the supplemental material; also data not shown). This list included several proteins that have been shown to be vaccine candidates, such as PspA (23,24), PcsB (20), LytA (21,25), and LytB (21,26).…”

“…1B and Table S2). Eight of the 15 OPK-positive proteins identified by this screen are known or predicted to be associated with cell wall or membrane, including PspA (23), PcsB (20), LytA (21,25), LytB (21,26), enolase, penicillin-binding protein 2b (PBP2b), the putative transporter permease ProWX, and the putative transporter substrate-binding lipoprotein SP148. The other seven proteins do not have obvious signal sequence for secretion or transmembrane segments and thus appear to be cytoplasmic proteins (i.e., the asparagine synthetase AsnA, the transcriptional regulator CodY, the phosphomevalonate kinase MvaK2, the putative segregation and condensation protein ScpB, the pyruvate oxidase SpxB, the small heat shock protein SP7, and the alcohol dehydrogenase SP285).…”

“…Numerous conserved proteins have been shown to confer protection in animal models against lethal infection by multiple serotypes of virulent S. pneumoniae strains. These protection-eliciting proteins include pneumococcal surface protein A (PspA) (23,24), peptidoglycan hydrolase PcsB (20), autolysin LytA (21,25), and endo-beta-N-acetylglucosaminidase LytB (21,26). Immunization with certain combinations of pneumococcal proteins proves to be safe in humans (19,(27)(28)(29).…”

“…This principle allows the vaccine efficacy in the immunized individuals to be quantitatively assessed by antibody response using the opsonophagocytosis killing (OPK) assay (33,34). Previous studies revealed that antibodies against pneumococcal proteins also promote bacterial clearance from the blood through opsonophagocytosis in a complement-and phagocyte-dependent manner (20,24,26,35). Daniels et al have recently described a modified version of the OPK assay to evaluate the level of protective antibody against PspA by increasing the incubation time (36).…”

The Versatility of Opportunistic Infections Caused by Gemella Isolates Is Supported by the Carriage of Virulence Factors From Multiple Origins

“…encode orthologs of the S. pneumoniae LytB glucosaminidase (Bai et al, 2014). LytB, a member of the CBP family of proteins, is a chain-dispersing enzyme and an important pneumococcal virulence factor (López and García, 2004;Ramos-Sevillano et al, 2011;Corsini et al, 2016). The presence of pCho-TAs in Gemella was fully sustained by sequence comparisons (Supplementary Tables S7, S8).…”