Serendipitous discovery of an immunoglobulin-binding autotransporter in Bordetella species.

Bordetella pertussis, human-derived Bordetella parapertussis (B. parapertussishu), and Bordetella bronchiseptica are gram-negative bacteria that cause respiratory infections in mammals. Multilocus enzyme electrophoresis, insertion sequence polymorphism, and genome sequence analyses have revealed that these bacteria are so closely related that they should be considered the same species (32, 34, 39). Despite their remarkable similarity, however, they differ in host specificity and disease manifestations. While B. bronchiseptica infects nearly all mammals, typically causing long-term asymptomatic or subclinical infections (4), B. pertussis and B. parapertussishu infect only humans and cause the acute disease known as whooping cough (2, 18, 20). Although many Bordetella factors with proven or postulated roles in pathogenesis have been identified and characterized, including some that differ substantially in amino acid sequence or expression profile among these subspecies, the molecular basis for differences in host specificity and disease is unknown. The autotransporter (AT) family comprises a large and diverse group of proteins that are exported across the outer membranes of gram-negative bacteria, where they either remain associated with the cell surface or are secreted into the extracellular milieu (see reference 21 for a review). All classical ATs consist of three domains: an N-terminal signal sequence, which directs the protein to the Sec secretion complex of the cytoplasmic membrane, a passenger domain, containing the effecter function, and a C-terminal β-domain, which facilitates the export of the passenger domain across the outer membrane. β-Domains are highly conserved and are the identifying feature of ATs. Passenger domains, however, are extremely diverse in both sequence and function. The majority of characterized passenger domains have roles in bacterial virulence through such actions as adherence to host cells, the proteolytic cleavage of host proteins, and the processing of other bacterial cell surface proteins (see reference 33 for a review). An analysis of the sequenced genomes of B. bronchiseptica, B. pertussis, and B. parapertussishu revealed the presence of 22 genes with the potential to encode classical ATs, of which the functions of only a few have been examined (34). These include pertactin (Prn), a putative adhesin that is included in B. pertussis acellular vaccines, BrkA, a protein that is involved in serum resistance and possibly adherence, and SphBI, a serine protease that is involved in the maturation of filamentous hemagglutinin (FHA), an adhesin of the two-partner secretion family (9, 14, 26, 29, 34, 36). Like all known protein virulence factors produced by Bordetella that have been identified so far, the expression of the genes encoding the characterized ATs is positively regulated by the BvgAS phosphorelay. We describe here the serendipitous discovery of BatB, a putative autotransporter produced by B. bronchiseptica. The characterization of a polyclonal antibody generated against an internal fragment of FHA revealed the presence of an unknown ∼180-kDa protein in whole-cell lysates (WCLs) of Bvg+-phase B. bronchiseptica. Since the protein was associated with the outer membrane and positively regulated by BvgAS, we sought to determine its identity. We determined that the protein was a previously uncharacterized putative Bordetella AT, BatB. Our results show that BatB binds immunoglobulin (Ig) and is necessary for B. bronchiseptica to resist inflammatory clearance from the respiratory tract. We also characterized differences in the sequence and expression of batB between B. bronchiseptica, B. pertussis, and B. parapertussishu. Our results suggest that BatB contributes to differences in disease and/or host specificity between the human- and non-human-adapted bordetellae.