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Brucella melitensis encounters a variety of conditions and stimuli during its life cycle—including environmental growth, intracellular infection, and extracellular dissemination—which necessitates flexibility of bacterial signaling to promote virulence. Cyclic-di-GMP is a bacterial secondary signaling molecule that plays an important role in adaptation to changing environments and altering virulence in a number of bacteria. To investigate the role of cyclic-di-GMP inB. melitensis , all 11 predicted cyclic-di-GMP-metabolizing proteins were separately deleted and the effect on virulence was determined. Three of these cyclic-di-GMP-metabolizing proteins were found to alter virulence. Deletion of thebpdA andbpdB genes resulted in attenuation of virulence of the bacterium, while deletion of thecgsB gene produced a hypervirulent strain. In aVibrio reporter system to monitor apparent alteration in levels of cyclic-di-GMP, both BpdA and BpdB displayed a phenotype consistent with cyclic-di-GMP-specific phosphodiesterases, while CgsB displayed a cyclic-di-GMP synthase phenotype. Further analysis found that deletion ofbpdA resulted in a dramatic decrease in flagellar promoter activities, and a flagellar mutant showed similar phenotypes to thebpdA andbpdB mutant strains in mouse models of infection. These data indicate a potential role for regulation of flagella inBrucella melitensis via cyclic-di-GMP.

Brucella melitensis Cyclic di-GMP Phosphodiesterase BpdA Controls Expression of Flagellar Genes