Roles of the TonB1 and TonB2 proteins in haemin iron acquisition and virulence in Riemerella anatipestifer.

Two TonB systems in Riemerella anatipestifer were found and characterized as ExbB1-ExbD1-TonB1 and ExbB2-ExbD2-ExbD2'-TonB2, but the significance of two sets of TonB complexes in R. anatipestifer is not clear. In this study, by deleting the tonB1 or tonB2 gene of R. anatipestifer strain OHS, we investigated the roles of the TonB1 and TonB2 proteins in iron acquisition and virulence. The results showed that strain CH3 could utilize haemin as the sole iron source in the presence of L-cysteine, but haemin iron acquisition was defective in the CH3 Delta tonB1 mutant, and the deletion of either tonB1 or tonB2 significantly reduced adhesion to and invasion of Vero cells. Animal experiments indicated that the LD50 of the CH3 Delta tonB1 and CH3 Delta tonB2 mutants in ducklings was similar to 224- and similar to 87-fold, respectively, higher than that of the WT OHS strain. Additional analysis indicated that blood bacterial loading of ducklings infected with CH3 Delta tonB1 or CH3 Delta tonB2 decreased significantly compared with that found for WT OHS-infected ducklings. Thus, our results indicated that the TonB1, but not TonB2 protein, is involved in haemin iron acquisition and that both TonB proteins are necessary for optimal bacterial virulence.