Scientific Publications by FDA Staff

Infect Immun 2009 Jan;77(1):274-85

Four superoxide dismutases contribute to Bacillus anthracis virulence and provide spores with redundant protection from oxidative stress.

Cybulski RJ Jr, Sanz P, Alem F, Stibitz S, Bull RL, O'Brien AD

The Bacillus anthracis genome encodes four superoxide dismutases (SODs), enzymes capable of detoxifying oxygen radicals. That two of these SODs, SOD15 and SODA1, are present in the outer-most layers of the B.anthracis spore, is indicated by previous proteomic analyses of the exosporium. Given the requirement that spores must survive interactions with reactive oxygen species generated by cells such as macrophages during infection, we hypothesized that SOD15 and SODA1 protect the spore from oxidative stress and contribute to the pathogenicity of B. anthracis. To test these theories, we constructed a double knockout (Deltasod15DeltasodA1) of B. anthracis Sterne strain 34F2 and assessed its lethality in an A/J mouse intranasal infection model. The 50% lethal dose of Deltasod15DeltasodA1 was similar to that of wild-type (34F2), but, surprisingly, measurable whole spore SOD activity was greater than 34F2. A quadruple knockout strain (Deltasod15DeltasodA1DeltasodCDeltasodA2) was then generated, and, as anticipated, spore-associated SOD activity was diminished. Moreover, the quadruple knockout strain, when compared to wild-type, was attenuated more than 40-fold upon intranasal challenge of mice. Spore resistance to exogenously-generated oxidative stress and to macrophage-mediated killing correlated with virulence in A/J mice. Allelic exchange that restored sod15 and sodA1 to their wild-type state restored wild-type characteristics. We conclude that SOD molecules within the spore afford B .anthracis protection against oxidative stress and enhance the pathogenicity of B.anthracis in the lung. We also surmise that the presence of four SOD alleles within the genome provides functional redundancy for this key enzyme.