CLONING AND SEQUENCING OF A BORDETELLA-PERTUSSIS SERUM RESISTANCE LOCUS

We have characterized a new virulence factor in Bordetella pertussis: serum resistance. Compared with Escherichia coil HB101, wild-type B. p ertussis was relatively resistant to classical-pathway, complement-dep endent killing by normal human serum. However, a mutant of B. pertussi s (BPM2041) which is less virulent in mice and which has Tn5'lac inser ted in a previously uncharacterized bvg-regulated gene was found to be at least 10-fold more susceptible to serum killing than the wild type . We have named this locus brk, for Bordetella resistance to killing. We have cloned and sequenced the brk locus, and it encodes two diverge ntly transcribed open reading frames (ORFs), termed BrkA and BrkB. Bot h ORPs are necessary for serum resistance. Within the 300 bases which separate the two ORPs and upstream of each ORF are putative sites for BvgA binding. BrkA shows 29% identity to pertactin and has two RGD mot ifs in addition to a conserved proteolytic processing site and an oute r membrane targeting signal. Like pertactin, BrkA is involved in adher ence and invasion. Despite the similarities, a pertactin mutant,vas fo und to be not as sensitive to serum killing as the BrkA or BrkB mutant s. BrkB is similar to ORFs in E. coli and Mycobacterium leprae and dis plays domains of homology to various transporters. On the basis of its hydropathy profile, BrkB is predicted to be a cytoplasmic membrane pr otein. By Southern biot, brk sequences were found in Bordetella bronch iseptica and Bordetella parapertussis but not in Bordetella avium.