IDENTIFICATION OF CAMPYLOBACTER-JEJUNI ON THE BASIS OF A SPECIES-SPECIFIC GENE THAT ENCODES A MEMBRANE-PROTEIN

To facilitate discrimination between the closely related enteropathoge ns Campylobacter jejuni and C. coli, unique differences in antigenic s urface structure were examined. A genomic library of C. jejuni 81116 w as constructed in plasmid pBluescriptIISK(-) and expressed in Escheric hia coli K-12. Rabbit hyperimmune serum raised against C. jejuni ATCC 29428 recognized a clone expressing a C. jejuni 24-kDa membrane-associ ated protein. Antiserum raised against sonicated recombinant E. coli e xpressing the 24-kDa protein reacted with C. jejuni, whereas C. coli d id not react specifically. Determination of the nucleotide sequence of the DNA insert of this recombinant plasmid revealed an open reading f rame encoding 214 amino acids; the gene was designated mapA; and its g ene product was designated MAPA. The 18 N-terminal amino acid residues constitute a signal sequence characteristic of prokaryotic membrane l ipoproteins. In a dot blot hybridization assay with a mapA probe, 120 clinical isolates of C. jejuni were unequivocally discriminated from 1 26 other campylobacters, including 34 C. coli isolates. A PCR test bas ed on the mapA sequence was developed for identification of C. jejuni. A PCR product was obtained with all of the clinical isolates of C. je juni tested from human, dog, cat, bovine calf, and chicken sources. Re combinant MAPA with an added C-terminal six-histidine tail was affinit y purified and used to immunize rabbits. The rabbit anti-MAPA serum sp ecifically recognized the protein in whole cells of C. jejuni on Weste rn blots (immunoblots). The MAPA protein was present in all of the C. jejuni strains tested and was absent in C. coli and related campylobac ters.