Filamentous Hemagglutinin ofBordetella bronchisepticaIs Required for Efficient Establishment of Tracheal Colonization

Abstract

Adherence to ciliated respiratory epithelial cells is considered a critical early step in Bordetella pathogenesis. ForBordetella pertussis, the etiologic agent of whooping cough, several factors have been shown to mediate adherence to cells and cell lines in vitro. These putative adhesins include filamentous hemagglutinin (FHA), fimbriae, pertactin, and pertussis toxin. Determining the precise roles of each of these factors in vivo, however, has been difficult, due in part to the lack of natural-host animal models for use with B. pertussis. Using the closely related species Bordetella bronchiseptica, and by constructing both deletion mutation and ectopic expression mutants, we have shown that FHA is both necessary and sufficient for mediating adherence to a rat lung epithelial (L2) cell line. Using a rat model of respiratory infection, we have shown that FHA is absolutely required, but not sufficient, for tracheal colonization in healthy, unanesthetized animals. FHA was not required for initial tracheal colonization in anesthetized animals, however, suggesting that its role in establishment may be dedicated to overcoming the clearance action of the mucociliary escalator.

Contribution of Bordetella bronchiseptica Filamentous Hemagglutinin and Pertactin to Respiratory Disease in Swine

“…B. bronchiseptica strain KM22, a virulent swine isolate, and strains TN27 and TN28, fhaB and prn deletion mutants of KM22, respectively, were maintained on Bordet-Gengou (BG) agar (Difco, Sparks, MD) supplemented with 10% sheep blood. B. bronchiseptica strains RB50 and RBX9 have been previously described (16). Liquid culture bacteria were grown at 37°C to mid-log phase in Stainer-Scholte broth.…”

“…Using the anti-FHA antibody, a ϳ240-kDa polypeptide was detected in the whole-cell lysates of all three strains. To ensure that this polypeptide was not a truncated form of FHA, we additionally evaluated all of the strains along with the previously well-characterized strains RB50 and RBX9 (RB50 containing an in-frame deletion of the fhaB gene) (16) by Western blot analysis. A similar ϳ240-kDa polypeptide was also detected in the whole-cell lysates of RBX9 (data not shown), indicating that this polypeptide was not a truncated form of FHA.…”

“…To test this, we compared the ability of TN27 (⌬fhaB) and TN28 (⌬prn) to KM22 in mediating adherence in a standard adherence assay. L2 cells were chosen to be evaluated due to routine use in attachment assays involving Bordetella (16,40). Attachment to a porcine epithelial cell line was also included in this assay.…”

“…This RGD domain also plays an important role in the upregulation of CR3 binding activity by interacting with the leukocyte response integrin/ integrin-associated protein located on monocytes and macrophages (27). In addition, FHA of B. bronchiseptica has been shown to be required for colonization of the rat trachea (16). PRN belongs to the type V autotransporter protein family and, similar to FHA, contains an RGD domain as well (18).…”

“…Swine isolates of B. bronchiseptica possess unique genetic and phenotypic traits relative to isolates from other host species (20,44,55). Recent experiments in rats demonstrate that attachment is a multifactorial process (16,40), and this is also likely to be true in swine. However, no definitive data exist with respect to swine, either in vivo or with swine tissue or cells in vitro.…”