cytochalasin-d and Anthrax

Previous reports suggested that lethal toxin (LT)-induced caspase-1 activity and/or IL-1beta accounted for Bacillus anthracis (BA) infection lethality. In contrast, we now report that caspase-1-mediated IL-1beta expression in response to BA spores is required for anti-BA host defenses. Caspase-1(-/-) and IL-1beta(-/-) mice are more susceptible than wild-type (WT) mice to lethal BA infection, are less able to kill BA both in vivo and in vitro, and addition of rIL-1beta to macrophages from these mice restored killing in vitro. Non-germinating BA spores induced caspase-1 activity, IL-1beta and nitric oxide, by which BA are killed in WT but not in caspase-1(-/-) mice, suggesting that the spore itself stimulated inflammatory responses. While spores induced IL-1beta in LT-susceptible and -resistant macrophages, LT induced IL-1beta only in LT-susceptible macrophages. Cooperation between MyD88-dependent and -independent signaling pathways was required for spore-induced, but not LT-induced, IL-1beta. While both spores and LT induced caspase-1 activity and IL-1beta, LT did not induce IL-1beta mRNA, and spores did not induce cell death. Thus different components of the same bacterium each induce IL-1beta by distinct signaling pathways. Whereas the spore-induced IL-1beta limits BA infection, LT-induced IL-1beta enables BA to escape host defenses.

Topics: Animal Structures; Animals; Anthrax; Antigens, Bacterial; Apoptosis; Bacillus anthracis; Bacterial Toxins; Caspase 1; Cycloheximide; Cytochalasin D; Interleukin-1beta; Lipopolysaccharides; Macrophages, Peritoneal; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; Myeloid Differentiation Factor 88; Nitric Oxide; Phagocytosis; Phosphorylation; Signal Transduction; Spores, Bacterial; STAT1 Transcription Factor; Staurosporine; Survival Analysis

Carbohydrate analyses of whole-spore extracts have confirmed the presence of rhamnose in the spore of the fully virulent Ames strain of Bacillus anthracis. A gene cluster containing loci with high homology to the rhamnose biosynthetic genes, rmlACBD, was identified within the B. anthracis chromosome. The first gene of this cluster, rmlA, was inactivated by forming a merodiploid cointegrate using an internal fragment of the gene within the Ames strain of B. anthracis to construct the mutant strain Ames-JAB1. Carbohydrate analysis of spores from this mutant demonstrated the loss of rhamnose. When assaying for spore infection of macrophages, we detected a significant decrease in the recovery with the Ames-JAB1 strain compared to the recovery with the Ames wild-type strain. When pre-treating macrophages with cytochalasin-D, spores of the mutant were further hindered in recovery, indicating that the spores were not able to bind as well to the macrophages. However, in guinea pigs challenge experiments, no difference in virulence was observed between the mutant and wild-type strains. These results suggest that the incorporation of rhamnose into the spore coat of B. anthracis is required for optimal interaction with macrophages but is not required for full virulence in this animal model.

Topics: Animals; Anthrax; Bacillus anthracis; Bacterial Adhesion; Cytochalasin D; Disease Models, Animal; Female; Guinea Pigs; Macrophages; Multigene Family; Mutation; Rhamnose; Sequence Deletion; Spores, Bacterial; Virulence