|Wannemuehler, Mike - IOWA STATE UNIV., AMES|
Submitted to: Pig Veterinary Society International Congress Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: September 20, 2000
Publication Date: N/A
Technical Abstract: Brachyspira pilosicoli is the agent of porcine intestinal spirochetosis. The organism also colonizes a range of animal species including humans. A unique feature associated with infection with B. pilosicoli is the attachment of large numbers of spirochetes by one end to the colonic epithelium. The major objective of this study was to identify unique B. pilosicoli outer membrane proteins (OMPs) that may be involved in the host parasite relationship. Membrane vesicles were obtained from B. pilosicoli strain 95-1000 by osmotic lysis and centrifugation. The vesicles were separated by density into four fractions: high density membrane vesicle fractions A (1.18 g/cm3) and B (1.16 g/cm3) and low density membrane vesicle fractions A (1.14 g/cm3) and B (1.12 g/cm3). The application of cellular markers showed that each band contained outer membrane and was free of cytoplasmic and flagella contamination. However, inner membrane contamination could not be completely eliminated. Low density membrane vesicle fraction B contained the lowest amount of penicillin binding proteins and was therefore selected as antigen for the production of monoclonal antibodies (MAbs). Five MAbs reacting with B. pilosicoli proteins of 23, 23/24 (doublet), 35, 61, and 79 kDa were produced. The antigens were not present in whole cell lysates of the type strains of B. hyodysenteriae, B. innocens, B. intermedia, B. alvinipulli or B. murdochii. Immunogold labeling with concentrated MAb that was directed against the 23 kDa protein confirmed that the protein was located on the outer membrane. The MAbs will be useful reagents for identifying unique B. pilosicoli membrane proteins and for elucidating the role of OMPs in interactions between B. pilosicoli cells and the host.