The boaB mutation in B. pseudomallei DD503 decreased attachment to A549 and HEp2 cells by ~50% (Fig 5A and 5B, respectively) and caused a 62% reduction in adherence to NHBE cultures (Fig 5C). As expected, the double mutant strain DD503.boaA.boaB exhibited significantly lower attachment to epithelial cells compared to the parent strain DD503 (Fig

buy AZD1480 5A, B, and 5C). The adherence levels of the double mutant, however, did not differ significantly from that of the single mutants in any of the cell types tested. One possible explanation for this apparent lack of synergistic effect is that other adhesins expressed by the double mutant strain DD503.boaA.boaB provide a high background level of adherence. Taken together, these Dibutyryl-cAMP results demonstrate that the boaA and boaB gene products contribute to the adherence of B. mallei and B. pseudomallei Obeticholic manufacturer to epithelial cells of the human respiratory tract. Figure 5 Adherence of B. mallei and B. pseudomallei strains to human respiratory epithelial cells. The effects of boaA and boaB mutations on the adherence

of B. pseudomallei (Bp) DD503 and B. mallei (Bm) ATCC23344 to monolayers of A549 (panels A and D) and HEp2 (panels B and E) cells and cultures of NHBE (panels C and F) was measured in duplicate on at least 3 separate occasions. The results are expressed as the mean percentage (± standard error) of inoculated bacteria adhering to epithelial cells. Asterisks indicate that the difference between the adherence of the mutant and that of the parental strain is statistically significant (P < 0.05). As previously stated, autotransporter adhesins often specify

Urease additional biological functions including survival within host cells [72]. In addition, B. pseudomallei and B. mallei are facultative intracellular pathogens that are particularly proficient at replicating inside professional phagocytic cells. For these reasons, we measured the ability of our panel of Burholderia mutant and parent strains to replicate within J774A.1 murine macrophages. In B. pseudomallei DD503, inactivation of the boa genes had no effect on phagocytosis of the organism (Fig 6A). Once inside macrophages, the boaA (DD503.boaA) and boaB (DD503.boaB) single mutants replicated at rates equivalent to that of the progenitor strain DD503 (Fig 6B). However, when both boaA and boaB genes were disrupted (DD503.boaA.boaB), intracellular growth was diminished by 60% (Fig 6B). To verify that this reduced intracellular fitness was not due to a global growth defect, we measured the growth of strains DD503 and DD503.boaA.boaB in broth as well as in tissue culture medium. We found that both strains grew at equivalent rates under both conditions (data not shown). Interestingly, the double mutant did not exhibit a growth defect in epithelial cells (data not shown). These results suggest a role for the BoaA and BoaB proteins in B. pseudomallei’s ability to grow inside professional phagocytes.