In-vitro characterisation of the phagocytosis and fate of anthrax spores in macrophages and the effects of anti-PA antibody

Poly-γ-Glutamate Capsule-Degrading Enzyme Treatment Enhances Phagocytosis and Killing of EncapsulatedBacillus anthracis

“…Alveolar phagocytes are thought to be involved in transporting inhaled spores to draining lymph nodes, where the spores are thought to germinate (42). Macrophages (60) and neutrophils (23,32) are reported to phagocytize and kill B. anthracis, and macrophages are required for resistance to infection in mice (7). Recent work suggests that antibody to the B. anthracis capsule enhances phagocytosis and killing of encapsulated bacilli (6,46,59) and that active (6) and passive (27) vaccination with capsule protects against experimental infection.…”

Analysis of a novel spore antigen in Bacillus anthracis that contributes to spore opsonization

“…To counteract the threat of such potential bioweapons, effective alternative vaccines and therapies should be developed. Better protection might be achieved, for instance, by targeting the early stages of infection by the spore as well as the toxins produced by the vegetative bacilli (Brossier et al, 2002;Cohen et al, 2000;Cote et al, 2005;Welkos et al, 2002). Thus, we hypothesized that a combination of PA and a defined set of spore-associated proteins could contribute to the development of a less reactogenic and more efficacious anthrax vaccine to provide broader coverage against emerging and/or engineered threats.…”

“…RAW264.7 cells were cultured in 24-well trays with coverslips and used in phagocytosis and intracellular spore viability assays as described previously (Cote et al, 2005;Welkos et al, 2001Welkos et al, , 2002. The cell cultures were infected with spores at an m.o.i.…”

“…As described by Banks et al (2005), they were then incubated for 30 min at 37 uC in medium containing 10 % fetal bovine serum and 5 mg gentamicin ml 21 then extensively washed to remove most of the unphagocytosed spores. The effects of antibodies on phagocytosis and intracellular spore germination and viability were assayed bacteriologically by viable-count determinations and by light and immunofluorescence microscopy of stained samples (Cote et al, 2005;Welkos et al, 2001Welkos et al, , 2002. Samples were stained with spore stain (malachite green) and counterstained with a Wright-Giemsa stain (Diff-Quik) (Welkos et al, 1989) for light microscopy.…”

“…Samples were stained with spore stain (malachite green) and counterstained with a Wright-Giemsa stain (Diff-Quik) (Welkos et al, 1989) for light microscopy. For immunofluorescence microscopy, coverslips were incubated with a rabbit antibody prepared against whole killed spores of the Ames strain that recognizes ungerminated and germinated spores as well as bacilli, and the spores were detected by doublelabelling with secondary antibodies, as described previously (Cote et al, 2005;Welkos et al, 2002).…”