title: Enhancement of the sensitivity of the whole-blood gamma interferon assay for diagnosis of Mycobacterium bovis infections in cattle.
abstract: In this study, we determined if the sensitivity of the currently available in vitro test to detect bovine tuberculosis could be enhanced by adding the following immunomodulators: interleukin-2 (IL-2); granulocyte-macrophage colony-stimulating factor (GM-CSF); antibodies neutralizing IL-10 and transforming growth factor beta (TGF-beta); mono-methyl-l-arginine, which blocks nitric oxide production; and l-methyl-tryptophan, which interferes with the indoleamine dioxygenase pathway. Blood was obtained from uninfected control cattle, experimentally infected cattle, cattle responding positively to the skin test in tuberculosis-free areas (false positives), and cattle naturally infected with Mycobacterium bovis from New Zealand and Great Britain. Gamma interferon (IFN-gamma) responses to bovine purified protein derivative (PPD-b), avian purified protein derivative, and a fusion protein of ESAT-6 and CFP-10 were measured. Mono-methyl-l-arginine, l-methyl-tryptophan, or an antibody neutralizing TGF-beta had minimal impact on IFN-gamma production. IL-2 and GM-CSF promoted IFN-gamma release whether antigen was present or not. In contrast, adding an antibody against IL-10 enhanced only antigen-specific responses. In particular, addition of anti-IL-10 to ESAT-6/CFP-10-stimulated blood cultures enhanced the test sensitivity. Furthermore, whole blood cells from field reactors produced substantial amounts of IL-10 upon stimulation with PPD-b or ESAT-6/CFP-10. Testing "false-positive" cattle from tuberculosis-free areas of New Zealand revealed that addition of anti-IL-10 did not compromise the test specificity. Therefore, the use of ESAT-6/CFP-10 with anti-IL-10 could be useful to detect cattle potentially infected with tuberculosis, which are not detected using current procedures.
authors: AgResearch, Hopkirk Research Institute, Palmerston North, New Zealand.
authors: Michel
authors: M
authors: Denis
authors: D Neil
authors: DN
authors: Wedlock
authors: Allison R
authors: AR
authors: McCarthy
authors: Natalie A
authors: NA
authors: Parlane
authors: Paul J
authors: PJ
authors: Cockle
authors: H Martin
authors: HM
authors: Vordermeier
authors: R Glyn
authors: RG
authors: Hewinson
authors: Bryce M
authors: BM
authors: Buddle
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology; D000942:Antigens, Bacterial / Q000276:immunology; D001426:Bacterial Proteins / Q000276:immunology; D002417:Cattle; D016207:Cytokines / Q000276:immunology; D007371:Interferon-gamma / Q000097:blood*; D016753:Interleukin-10 / Q000276:immunology*; D009163:Mycobacterium bovis / Q000276:immunology*; D009500:Neutralization Tests; D011993:Recombinant Fusion Proteins / Q000276:immunology; D012680:Sensitivity and Specificity; D014380:Tuberculosis, Bovine / Q000175:diagnosis*
chemical_list: D000906:Antibodies; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C117940:CFP-10 protein, Mycobacterium tuberculosis; D016207:Cytokines; C525005:ESAT-6 antigen, Mycobacterium leprae; D011993:Recombinant Fusion Proteins; D016753:Interleukin-10; D007371:Interferon-gamma