الفهرس 
Cover Page Title in EnglishOnly 14 pages are availabe for public view
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Abstract
Standardization of Interferon-gamma assay for diagnosis of bovine tuberculosis in EgyptBovine tuberculosis (bTB) is a chronic disease of animals caused primarily by Mycobacterium bovis (M. bovis), a member of the M. tuberculosis complex. In Egypt, bTB continues to cause significant losses in the cattle industry and a major public health problem. There are critical gaps in the current Egyptian national bTB surveillance program (NbTBS) as it based only on the use of single cervical tuberculin test (SCT) with a mammalian type of PPD. Despite of its world wide application, the IFN-γ assay has not been applied in Egypt until today. This assay is a fully validated test by the OIE, EU and USDA as an ancillary test to the tuberculin skin tests. The objective of this study was to determine appropriate cut-off values for protein purified derivatives (PPD) and ESAT6-CFP10 antigens to obtain optimal results of IFN-γ assay in terms of sensitivity and specificity to complement official skin-test screening in Egypt. Both bacterial culture and PCR were used as gold standard tests to verify the bTB infectious status of animals included in the present study. The sensitivity and specificity (Ser and Spr) of PPD and antigen cocktail-based IFN-γ assays (IFN-γ-BA and IFN-γ-EC) were analyzed by different cut-off points, relative to gold standards bTB positive confirmation, retrospectively using blood samples collected from 34 SCT reactors from recognized bTB-infected zones in five high bTB prevalence governorates during the 2011, 2012 and 2013. Bovine TB infection is verified for each reactor by the disclosure of typical visible lesions during post-mortem examination in the slaughterhouses; culture of M. bovis in primary isolation medium and molecular identification of M. tuberculosis complex (MTC) and M. bovis using PCR. The absolute specificity (Sp) was studied using blood samples collected from 14 cattle from one bTB-free herd from a bTB-free zone. Collectively, the infection rate was decreased from 2009 to 2012, and then dramatically increased in 2013. Based on the localities of SCT reactors, bTB is extensively distributed among cattle in the Nile Delta. Most of slaughtered reactors that were included in the present study showed typical VL. In SCT-reactors with VL (n = 23), the tuberculosis infection was confirmed in 22 slaughtered animals based on the gross lesions, bacterial culture and PCR. Collectively, the animal that was confirmed negative for bTB, was negative by both IFN-γ assays. 22 animals were positive for the IFN-γ-EC assays by one or more applied cut-off points. In SCT-reactors with NVL (n = 11), the tuberculosis infection was confirmed only in 7 slaughtered animals based on the bacterial culture and PCR. Of these animals, 5 and 6 animals were positive by IFN-γ-BA and IFN-γ-EC respectively by one or more applied cut-off points. Results revealed the use of IFN-γ-EC provided high sensitivity but equal specificity, comparable to the estimates obtained for IFN-γ-BA. Data analysis showed the use of (PPDbOD>0.1, PPDbOD–NILOD>0.05 and PPDbOD>PPDaOD) strategy to get optimal IFN-γ-BA results {PPD-Ser (90%) and PPD-Spr (100.0%)}, and (ECOD–NILOD≥0.1) strategy to get optimal IFN-γ-EC results {EC-Ser (97%) and EC-Spr (100.0%)}. In the present study, in case of confirmed bTB infected animals, five animals were SCT false positive; one animal was IFN-γ-EC false negative and three animals were IFN-γ-BA false negative. These findings suggest that removal of all animals reacting positive to either of the two tests to control bovine TB outbreaks should be economically evaluated especially in developing countries. Taken together, the study suggests retesting skin-test positive/ IFN-γ assay negative animals with the IFN-γ assay to increase the Se of the IFN-γ test. Finally, in the present study the suggested cut-off values that revealed excellent test specificity (Sp of 100%) with the two IFN-γ assays, and acceptable test sensitivity (Ser of 90% and 97% with IFN-γ-BA and IFN-γ-EC respectively). Thus, the IFN-γ assay appears to be a useful complementary diagnostic tool for the detection of bTB in Egypt when used with specific peptides and an adapted protocol. In conclusion, this is the first report pointing out the appropriate cut-off points to optimize IFN-γ assay as a routine ancillary test for diagnosis of bTB in Egypt.