Antibodies toStaphylococcus aureuscapsular polysaccharides 5 and 8 perform similarlyin vitrobut are functionally distinctin vivo

Abstract

The capsular polysaccharide (CP) produced by Staphylococcus aureus is a virulence factor that allows the organism to evade uptake and killing by host neutrophils. Polyclonal antibodies to the serotype 5 (CP5) and type 8 (CP8) capsular polysaccharides are opsonic and protect mice against experimental bacteremia provoked by encapsulated staphylococci. Thus, passive immunotherapy using CP antibodies has been considered for the prevention or treatment of invasive antibioticresistant S. aureus infections. In this report, we generated monoclonal antibodies (mAbs) against S. aureus CP5 or CP8. Backbone specific mAbs reacted with native and O-deacetylated CPs, whereas Oacetyl specific mAbs reacted only with native CPs. Reference strains of S. aureus and a selection of clinical isolates reacted by colony immunoblot with the CP5 and CP8 mAbs in a serotype-specific manner. The mAbs mediated in vitro CP type-specific opsonophagocytic killing of S. aureus strains, and mice passively immunized with CP5 mAbs were protected against S. aureus bacteremia. Neither CP8-specific mAbs or polyclonal antibodies protected mice against bacteremia provoked by serotype 8 S. aureus clinical isolates, although these same antibodies did protect against a serotype 5 S. aureus strain genetically engineered to produce CP8. We detected soluble CP8 in culture supernatants of serotype 8 clinical isolates and in the plasma of infected animals. Serotype 5 S. aureus released significantly less soluble CP5 in vitro and in vivo. The release of soluble CP8 by S. aureus may contribute to the inability of CP8 vaccines or antibodies to protect against serotype 8 staphylococcal infections.

Release of Staphylococcus aureus extracellular vesicles and their application as a vaccine platform

“…We reported that S. aureus CP was shed from broth-grown S. aureus cells 59 , and it is feasible that EVs could serve as a vehicle to liberate CP from the cell envelope. The Streptococcus pneumoniae capsule was reported to hinder EV release in this pathogen 7 , whereas no effect was observed on EV yield in strains with or without the hyaluronic capsule of Streptococcus pyogenes 8 .…”

“…The membranes were immersed in PBST + 5% skim milk for 2 h before blocking the staphylococcal IgG binding proteins (Spa and Sbi) by overnight incubation at 4 °C with an irrelevant human IgG1 monoclonal antibody (10 µg ml −1 ) in PBST + 1% skim milk. The membrane was washed with PBST and incubated overnight at 4 °C with sera (diluted 1:1000 in PBST + 1% skim milk) pooled from mice immunized with EVs (see below) or murine mAbs 59 to CP5 (4C2; 1.2 µg ml −1 ) or CP8 (5A6; 1.2 µg ml −1 ). After washes with PBST, the membrane was incubated with alkaline phosphatase (AP)-conjugated goat anti-mouse antibody (1:15,000; Sigma; Cat.…”

MgrA Negatively Impacts Staphylococcus aureus Invasion by Regulating Capsule and FnbA

“…In the study, USA300 LAC and MW2 strains were used to show that MgrA has a positive effect on attachment to endothelial cells, which is in contrast to our results. USA300 strains are known to produce no capsule because of multiple mutations in the cap locus (45), whereas MW2 produces only a scant amount of capsule due to a frameshift mutation in one of the cap genes (46). As such, regulation of capsule by MgrA would not play a role in cell attachment in USA300 LAC and MW2, which may explain, in part, the difference between their and our studies.…”

The biogenesis of extracellular vesicles fromStaphylococcus aureusand their application as a novel vaccine platform

“…S. aureus CPs were associated with or packaged within EVs isolated from S. aureus strain Newman and MN8 in our study. We reported that S. aureus CP was shed from broth-grown S. aureus cells 78 , and it is feasible that EVs could serve as a vehicle to liberate CP from the cell envelope. The S. pneumonia capsule was reported to hinder EV release in this pathogen 7 , whereas no effect was observed on EV yield in strains with or without the hyaluronic capsule of S. pyogenes 8 .…”

“…To block the staphylococcal IgG binding proteins Spa and Sbi, the membranes were blocked with PBST + 5% skim milk and incubated overnight at 4°C with an irrelevant human IgG1 monoclonal antibody (10 μg/ml) in PBST + 1% skim milk. The membrane was washed with PBST and incubated overnight at 4°C with sera (diluted 1:1000 in PBST + 1% skim milk) pooled from mice immunized with EVs (see below) or murine mAbs 78 to CP5 (4C2; 1.2 μg/ml) or CP8 (5A6; 1.2 μg/ml). After washes with PBST, the membrane was incubated with alkaline phosphatase (AP)-conjugated goat anti-mouse antibody (1:15000 dilution in PBST + 1% skim milk) at RT for 2 h. The membrane was washed with PBST and developed with AP membrane substrate (KPL).…”