Protective Role of Bacillus anthracis Exosporium in Macrophage-Mediated Killing by Nitric Oxide

Abstract

The ability of the endospore-forming, gram-positive bacterium Bacillus anthracis to survive in activated macrophages is key to its germination and survival. In a previous publication, we discovered that exposure of primary murine macrophages to B. anthracis endospores upregulated NOS 2 concomitant with an · NO-dependent bactericidal response. Since NOS 2 also generates O 2 · ؊ , experiments were designed to determine whether NOS 2 formed peroxynitrite (ONOO ؊ ) from the reaction of · NO with O 2 · ؊ and if so, was ONOO ؊ microbicidal toward B. anthracis. Our findings suggest that ONOO ؊ was formed upon macrophage infection by B. anthracis endospores; however, ONOO ؊ does not appear to exhibit microbicidal activity toward this bacterium. In contrast, the exosporium of B. anthracis, which exhibits arginase activity, protected B. anthracis from macrophage-mediated killing by decreasing · NO levels in the macrophage. Thus, the ability of B. anthracis to subvert · NO production has important implications in the control of B. anthracis-induced infection.

Bacillus anthracis spores and lethal toxin induce IL‐1β via functionally distinct signaling pathways

“…We previously observed that murine peritoneal macrophages were capable of killing BA in vitro through an NO-mediated bactericidal mechanism that targets the emerging vegetative bacilli [8,9,13,14]. Therefore, we determined whether the susceptibility of caspase-1 -/-mice to lethal infection was due to an inability of its macrophages to kill the BA.…”

“…No germination of DgerH spores was evident either by direct inspection or by susceptibility of spores in culture supernatants to heating. Since macrophages kill the emerging vegetative bacilli within macrophages in an NO-dependent manner [8,13], we measured whether BA could induce NO, a key microbicidal effector mechanism of murine macrophages, in the absence of caspase-1. While DgerH BA spores induced the expression of NO in macrophages of caspase-1 +/+ mice, there was minimal NO generated following the same stimulation in macrophages of caspase-1 -/-mice (Fig.…”

“…Germination-proficient BA Sterne 34F2 and a congenic germination-deficient mutant strain, DgerH, were used during this study [13,14]. The spores were prepared as previously described [8].…”

Surface architecture of endospores of the Bacillus cereus/anthracis/thuringiensis family at the subnanometer scale

“…For example, in B. anthracis, BclA-integrin interaction promotes spore uptake by macrophages in vitro (22). The exosporium contributes protection against macrophage-mediated damage (21), whereas enzymes in the exosporium (23) moderate responses to germinants. However, although the exosporium may have a role in natural anthrax infections, a BclA-containing exosporium is not required for anthrax infection in animal models (24).…”

“…The exosporium is likely to contain a number of other structural proteins (many reviewed in ref. 4) as well as loosely bound proteins such as immune inhibitor A (20) and arginase (21).…”