Immunization with a streptococcal multiple-epitope recombinant protein protects mice against invasive group A streptococcal infection

Abstract

Streptococcus pyogenes (group A Streptococcus; GAS) causes clinical diseases, including pharyngitis, scarlet fever, impetigo, necrotizing fasciitis and streptococcal toxic shock syndrome. A number of group A streptococcus vaccine candidates have been developed, but only one 26-valent recombinant M protein vaccine has entered clinical trials. Differing from the design of a 26-valent recombinant M protein vaccine, we provide here a vaccination using the polyvalence epitope recombinant FSBM protein (rFSBM), which contains four different epitopes, including the fibronectin-binding repeats domain of streptococcal fibronectin binding protein Sfb1, the C-terminal immunogenic segment of streptolysin S, the C3-binding motif of streptococcal pyrogenic exotoxin B, and the C-terminal conserved segment of M protein. Vaccination with the rFSBM protein successfully prevented mortality and skin lesions caused by several emm strains of GAS infection. Anti-FSBM antibodies collected from the rFSBM-immunized mice were able to opsonize at least six emm strains and can neutralize the hemolytic activity of streptolysin S. Furthermore, the internalization of GAS into nonphagocytic cells is also reduced by anti-FSBM serum. These findings suggest that rFSBM can be applied as a vaccine candidate to prevent different emm strains of GAS infection.

Streptolysin S induces mitochondrial damage and macrophage death through inhibiting degradation of glycogen synthase kinase-3β in Streptococcus pyogenes infection

“…RAW264.7 cells (4 × 10 5 cells/well in 24-well culture plate) were infected with wild type strain GAS at MOI of 25 for 1 h in presence of 1,000 μM of LiCl as previously described. Moreover, shGSK-3β-RAW264.7 cells and shLuc-RAW264.7 cells (4 × 10 5 cells/well in 24-well culture plate) were infected with wild type strain GAS at MOI of 25 for 1 h. After that, gentamicin (50 μg/ml) was added to kill the extracellular GAS for 1 h, and then cells were cultivated in antibiotic-free DMEM medium for an additional 16 h. Total viable bacteria were quantified by plating on THY agar plates as described previously 4 . For examination of initial GAS uptake, gentamicin (50 μg/ml) was added to kill the extracellular GAS for 0.5 h after GAS infection, and then cells were washed, and intracellular bacteria were quantified by plating on THY agar plates as described previously 4 .…”

“…SLS is the other potent cytolysin with a wide range of targets, but the immunogenicity of SLS is lower compared with SLO. Anti-SLS antibody induced by immunizing with the SLS peptide, coupled to keyhole limpet hemocyanin or with the SLS epitope-contained recombinant protein, can reduce the SLS-mediated hemolysis and mortality of GAS-infected mice 3,4 . The cytotoxic effects of SLS have been reported in different types of cells, in which it mediated apoptosis, oncosis, and activation of the proinflammatory signaling pathways during the invasive GAS infection 5–9 .…”

IL-33/ST2 Axis Plays a Protective Effect in Streptococcus pyogenes Infection through Strengthening of the Innate Immunity

“…Groups of six WT, IL - 33 -KO, and ST2 -KO mice were anesthetized by isoflurane inhalation and then injected subcutaneously with 1 mL of air to form an air pouch one day before the infection. One day later, 0.1 mL of bacterial suspension containing 3 × 10 8 S. pyogenes NZ131 cells was inoculated into the air pouch [ 48 ]. The animals were observed every day for a total of 13 days.…”

“…After that, cell suspensions were centrifuged at 6000 rpm for 10 min at 4 • C. The supernatants were discarded, and the sediment cells were cultured with RPMI medium containing gentamicin (50 µg/mL) at 37 • C for 30 min to kill extracellular bacteria. After several washes by RPMI medium, the cells were lysed, and intracellular viable bacteria were then quantified by plating on THY agar plates as described previously [48]. The bacterial numbers presented on this timepoint represent the phagocytosis of neutrophils.…”

“…Neutrophils (1.5 × 10 6 cells in 0.2 mL RPMI medium, Gibco) from WT, IL-33-KO, and ST2-KO mice were infected with GAS (50 µL) at a MOI of 0.01 for 4 h at 37 • C. After that, cell suspensions were centrifuged, the supernatants were collected, the sediment cells were lysed. The intracellular viable bacteria and remaining viable bacteria in the supernatants were further quantified by plating on THY agar plates as described previ-ously [48]. The bacterial numbers presented on this timepoint represent the bactericidal activity of neutrophils.…”