title: Development of diagnostic reagents to differentiate between Mycobacterium bovis BCG vaccination and M. bovis infection in cattle.
abstract: In Great Britain a recent independent scientific review for the government has concluded that the development of a cattle vaccine against Mycobacterium bovis holds the best long-term prospect for tuberculosis control in British herds. A sine qua non for vaccination is the development of a complementary diagnostic test to differentiate between vaccinated animals and those infected with M. bovis so that test-and-slaughter-based control strategies can continue alongside vaccination. In order to assess the feasibility of developing a differential diagnostic test for a live vaccine, we chose M. bovis BCG Pasteur as a model system. Recombinant forms of antigens which are expressed in M. bovis but not, or only at low levels, in BCG Pasteur (ESAT-6, MPB64, MPB70, and MPB83) were produced. These reagents were tested either alone or in combination by using peripheral blood mononuclear cells from M. bovis-infected, BCG-vaccinated, and Mycobacterium avium-sensitized calves. All four antigens induced in vitro proliferation and gamma interferon responses only in M. bovis-infected animals. A cocktail composed of ESAT-6, MPB64, and MPB83 identified infected animals but not those vaccinated with BCG. In addition, promiscuous T-cell epitopes of ESAT-6, MPB64, and MPB83 were formulated into a peptide cocktail. In T-cell assays with this peptide cocktail, infected animals were identified with frequencies similar to those obtained in assays with the protein cocktail, while BCG-vaccinated or M. avium-sensitized animals did not respond. In summary, our results suggest that peptide and protein cocktails can be designed to discriminate between M. bovis infection and BCG vaccination.
authors.affiliation: TB Research Group, Bacteriology Department, Veterinary Laboratories Agency-Weybridge, New Haw, Addlestone, KT15 3NB, United Kingdom. mvordermeier.vla@gtnet.gov
authors.forename: H M
authors.initials: HM
authors.lastname: Vordermeier
authors.forename: P C
authors.initials: PC
authors.lastname: Cockle
authors.forename: A
authors.initials: A
authors.lastname: Whelan
authors.forename: S
authors.initials: S
authors.lastname: Rhodes
authors.forename: N
authors.initials: N
authors.lastname: Palmer
authors.forename: D
authors.initials: D
authors.lastname: Bakker
authors.forename: R G
authors.initials: RG
authors.lastname: Hewinson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry / Q000276:immunology; D001500:BCG Vaccine / Q000737:chemistry / Q000276:immunology*; D015415:Biomarkers; D002417:Cattle; D003937:Diagnosis, Differential; D007371:Interferon-gamma / Q000276:immunology; D007376:Interleukin-2 / Q000032:analysis / Q000276:immunology; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000382:microbiology; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology; D008969:Molecular Sequence Data; D009164:Mycobacterium Infections / Q000175:diagnosis* / Q000276:immunology; D009162:Mycobacterium avium / Q000276:immunology; D009163:Mycobacterium bovis / Q000276:immunology / Q000302:isolation & purification*; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology; D014374:Tuberculin Test; D014380:Tuberculosis, Bovine / Q000175:diagnosis* / Q000276:immunology / Q000517:prevention & control*
chemical_list: D000942:Antigens, Bacterial; D001500:BCG Vaccine; D015415:Biomarkers; D007376:Interleukin-2; D010446:Peptide Fragments; D011994:Recombinant Proteins; D007371:Interferon-gamma