Role of Bacillus anthracis spore structures in macrophage cytokine responses

The innate immune response of macrophages (M phi) to spores, the environmentally acquired form of Bacillus anthracis, is poorly characterized. We therefore examined the early M phi cytokine response to B. anthracis spores, before germination. M phi were exposed to bacilli and spores of Sterne strain 34F2 and its congenic nongerminating mutant (Delta gerH), and cytokine expression was measured by real-time PCR and an enzyme-linked immunosorbent assay. The exosporium spore layer was retained (exo(+)) or removed by sonication (exo(-)). Spores consistently induced a strong cytokine response, with the exo(-) spores eliciting a two- to threefold-higher response than exo(+) spores. The threshold for interleukin-1 beta (IL-1 beta) production by wild-type M phi was significantly lower than that required for tumor necrosis factor alpha expression. Cytokine production was largely dependent on MyD88, suggesting Toll-like receptor involvement; however, the expression of beta interferon in MyD88(-/-) M phi suggests involvement of a MyD88-independent pathway. We conclude that (i) the B. anthracis spore is not immunologically inert, (ii) the exosporium masks epitopes recognized by the M phi, (iii) the M phi cytokine response to B. anthracis involves multiple pattern recognition receptors and signaling pathways, and (iv) compared to other cytokines, IL-1 beta is expressed at a lower spore concentration.