SUMMARYMice infected with lactate dehydrogenase-elevating virus (LDV) were found to produce high titres of IgG anti-LDV antibodies that remained elevated for more than l year. This response, which was T-dependent, showed a striking preponderance of IgG2a with, from one strain to another, variable proportions of IgG2b and IgG3 but always very little IgG1. The binding of these antibodies to viral protein blots showed a major reaction with VP3, the heterogeneous glycosylated material of the viral envelope. A minor reaction was also noted with VPI, the nucleocapsid protein, but no antibodies were detected against VP2, the non-glycosylated envelope protein of LDV. A similar preponderance of anti-VP3 antibodies was also observed in a large set of anti-LDV hybridomas. Analysis of VP3 with monoclonal antibodies suggested that, despite its heterogeneity, this material has a common polypeptide moiety that apparently carries two major epitopes.

BackgroundBovine tuberculosis (TB) control programs generally rely on the tuberculin skin test (TST) for ante-mortem detection of Mycobacterium bovis-infected cattle.ResultsPresent findings demonstrate that a rapid antibody test based on Dual-Path Platform (DPP®) technology, when applied 1-3 weeks after TST, detected 9 of 11 and 34 of 52 TST non-reactive yet M. bovis-infected cattle from the US and GB, respectively. The specificity of the assay ranged from 98.9% (n = 92, US) to 96.0% (n = 50, GB) with samples from TB-free herds. Multi-antigen print immunoassay (MAPIA) revealed the presence of antibodies to multiple antigens of M. bovis in sera from TST non-reactors diagnosed with TB.ConclusionsThus, use of serologic assays in series with TST can identify a significant number of TST non-reactive tuberculous cattle for more efficient removal from TB-affected herds.

Bovine tuberculosis (TB), caused by Mycobacterium bovis, remains an important zoonotic disease posing a serious threat to livestock and wildlife. The current TB tests relying on cell-mediated and humoral immune responses in cattle have performance limitations. To identify new serodiagnostic markers of bovine TB, we screened a panel of 101 recombinant proteins, including 10 polyepitope fusions, by a multiantigen print immunoassay (MAPIA) with well-characterized serum samples serially collected from cattle with experimental or naturally acquired M. bovis infection. A novel set of 12 seroreactive antigens was established. Evaluation of selected proteins in the dual-path platform (DPP) assay showed that the highest diagnostic accuracy (ϳ95%) was achieved with a cocktail of five best-performing antigens, thus demonstrating the potential for development of an improved and more practical serodiagnostic test for bovine TB.KEYWORDS antigen, antibody, Mycobacterium bovis, serodiagnosis, tuberculosis B ovine tuberculosis (TB) is an important zoonotic disease caused by Mycobacterium bovis, a member of the M. tuberculosis complex (1). Although M. bovis is most often isolated from tuberculous cattle, it has a broad range of susceptible host species, including humans (2-4). The control of TB in cattle is therefore difficult due to the existence of wildlife reservoirs of M. bovis, such as white-tailed deer in the United States, Eurasian badgers in Great Britain, wild boars in Spain, and brushtail possums in New Zealand (5-8).The antemortem diagnostic methods currently approved for use in cattle have limitations. The intradermal tuberculin test has suboptimal sensitivity and inconsistent performance (1, 9, 10), while the available blood-based assays, such as the Thermo Fisher Scientific Bovigam TB kit or the Idexx M. bovis antibody (Ab) test, lack the required accuracy and show a significant variability when used in different geographic areas (11-13). Given the ease of sample collection and the test procedure, antibody detection assays may be useful to identify M. bovis-infected cattle (1,14,15), but the existing methods require improvement.The primary goal of the present study was to identify novel seroreactive antigens of M. bovis. Using a multiantigen print immunoassay (MAPIA) and well-characterized serum samples serially collected from cattle with experimental or naturally acquired M. bovis infection, we screened a panel of 101 recombinant proteins of M. tuberculosis, including 10 polyepitope fusions. The performance of MAPIA-selected candidates was also evaluated in pilot experiments using the dual-path platform (DPP) technology to

Minute amounts of the anti-L3T4 antibody designated GK1.5 were found to deeply suppress in vivo antibody responses to T-dependent antigens. Primary responses to sheep erythrocytes were completely inhibited even when GK1.5 was administered up to 6 days before or 4 days after the antigen. Secondary responses to potent immunogens like sheep erythrocytes or keyhole limpet hemocyanin were also completely abolished by a single injection of GK1.5 just before the boost. This treatment had no effect on T-independent reactions such as the polyclonal activation of B lymphocytes with lipopolysaccharide or the anti-2,4,6-trinitrophenyl (TNP) response elicited by injection of TNP-Ficoll.