Gamma interferon test for the diagnosis of buffalo Tuberculosis

What is it about?

Bovine tuberculosis (bTB) is a worldwide zoonosis that affects many species of domestic and wild animals. Mycobaterium bovis is the main cause of infection in water buffalo (Bubalus bubalis) and bovines and is of great concern for human health and for buffalo producers in Italy. The bTB eradication programme is based on slaughterhouse surveillance and intradermal skin tests. Other in vivo diagnostic methods such as the interferon-gamma (IFN-γ) assay have been developed and are widely used in cattle to accelerate the elimination of bTB positive animals. The present study is the first to assess the use and performance of IFN-γ assays, which is used as an ancillary test for bTB diagnosis in water buffalo, and presents the results of a field-evaluation of the assay from 2012 to 2019 during the buffalo bTB eradication programme in Italy. The study involved 489 buffaloes with a positive result to the single intradermal tuberculin test (SITT). The IFN-γ assays and single intradermal comparative tuberculin test were used as confirmation tests. Then, a total of 458 buffaloes, reared on officially tuberculosis-free (OTF) herds, that were confirmed bTB-free for at least the last 6 years were subjected to IFN-γ testing. Furthermore, to evaluate the IFN-γ test in an OTF herd with Paratuberculosis (PTB) infection, 103 buffaloes were subjected to SITT and IFN-γ test simultaneously. Four interpretative criteria were used, and the IFN-γ test showed high levels of accuracy, with sensitivity levels between 75.3% (CI 95% 71.2–79.0%) and 98.4% (CI 95% 96.7–99.4%) and specificity levels between 94.3% (CI 95% 91.2–96.50%) and 98.5% (CI 95% 96.9–99.4%), depending on the criterion used. Finally, in the OTF herd with PTB infection, in buffalo, the IFN-γ test displayed high specificity values according to all 4 interpretative criteria, with specificity levels between 96.7% (CI 95% 88.4–99.5%) and 100% (CI 95% 96.2–100%), while SITT specificity proved unsatisfactory, with a level of 45.3% (CI 95% 35.0–55.7%). Our results showed that the IFN-γ test in the buffalo species could reach high Sensitivity and Specificity values, and that the level of Sensitivity and Specificity could be chosen based on the interpretative criterion and the antigens used depending on the health status of the herd and the epidemiological context of the territory. The IFN-γ test and the use of different interpretative criteria proved to be useful to implement bTB diagnostic strategies in buffalo herds, with the possibility of a flexible use of the assay.

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Why is it important?

The aim of this study was to evaluate the performance of the IFN-γ test in healthy buffaloes and naturally M. bovis-infected buffaloes. Therefore, we developed an IFN-γ test with combination of PPDs, a mixture of ESAT-6 and CFP-10 and four different interpretative criteria. The final goal was to verify the use of the IFN-γ test as an ancillary test to implement bTB diagnostic strategies in buffalo herds. The IFN-γ test has been proved as an objective method, as it utilizes a standard procedure and is not affected by the subjectivity of the operator, in contrast to SITT, which could be influenced by several factors that can interfere with Sp and Se (15, 30). Moreover, the IFN-γ test has a short execution time and can be repeated without time constraints. Unlike SITT, it does not interfere with the immune profile of the animal. In addition, it is not influenced by treatments with immunosuppressive drugs and is not affected, or at least is much less affected, by infection with mycobacteria other than M. bovis. Furthermore, its different interpretative criteria and antigens can be adopted according to the objective to be pursued and the epidemiological context (39). Our results in buffalo indicate that an IFN-γ-positive animal, especially if the test is applied in a bTB-infected herd, has a very high probability of really being infected.

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