Francisella tularensis subsp. tularensis Schu S4 Disulfide Bond Formation Protein B, but Not an RND-Type Efflux Pump, Is Required for Virulence

Francisella tularensis subsp.tularensis is a highly virulent bacterium that is a CDC select agent. Despite advancements in the understanding of its biology, details pertaining to virulence are poorly understood. In previous work, we identified a transposon insertion mutant in the FTT0107c locus that was defective in intracellular survival in HepG2 and J77A.1 cells. Here, we report that this mutant was also highly attenuated in vivo. The FTT0107c locus is predicted to encode an ortholog of the disulfide bond formation B protein (DsbB). This designation was confirmed by complementation of anEscherichia coli dsbB mutant. ThisdsbB mutant of Schu S4 was highly attenuated in mice, but unlike what has been reported forFrancisella novicida , intranasal immunization with a sublethal dose did not induce protection against wild-type challenge.dsbB was found to be transcribed in an operon withacrA andacrB , which encode an RND-type efflux pump. However, this pump did not make a significant contribution to virulence because strains with nonpolar deletions inacrA andacrB behaved like wild-type strain Schu S4 with respect to intracellular growth and in vivo virulence. This result is in contrast to a report that anacrB mutant of a live vaccine strain ofF. tularensis has decreased virulence in mice. Overall, these results demonstrate key differences between the virulence requirements of Schu S4 and less virulent subspecies ofFrancisella . We have shown that DsbB is a key participant in intracellular growth and virulence, and our results suggest that there are critical virulence factors that contain disulfide bonds.