Parsons Sven D C, Menezes Angela M, Cooper David, Walzl Gerhard, Warren Robin M, van Helden Paul D
DST/NRF Centre of Excellence for Biomedical TB Research/MRC Centre for Molecular and Cellular Biology/Division of Molecular Biology and Human Genetics, Stellenbosch University, P.O. Box 19063, Tygerberg, 7505, South Africa.
Vet Immunol Immunopathol. 2012 Aug 15;148(3-4):337-42. doi: 10.1016/j.vetimm.2012.04.025. Epub 2012 May 19.
The development of diagnostic tests for tuberculosis (TB) in exotic species is constrained by host biology and the limited availability of suitable assay reagents. As such, we evaluated a gene expression assay (GEA) which is easily modified for novel species and allows for initial sample processing under field conditions. African buffaloes (Syncerus caffer) were categorized using the single comparative intradermal tuberculin test, and blood from test-positive and test-negative animals was incubated for 20 h in "Nil" tubes (containing saline) and "TB Antigen" tubes (containing Mycobacterium tuberculosis complex (MTC)-specific antigens) of a commercial human TB test, the QuantiFERON(®)-TB Gold (In-Tube) (QFT) assay. Blood samples were then stabilized in RNAlater(®) and transported to the laboratory for RNA extraction. A Custom TaqMan GEA was used to calculate the relative abundance of interferon-gamma (IFN-γ) mRNA in the TB Antigen tube compared to that in the Nil tube as a marker of immune activation in response to MTC antigen recognition. The GEA results from the two buffalo groups were compared and a cutoff value of 2.85 was calculated to differentiate between animals from these groups with a sensitivity of 80% (95% C.I.: 56-94%) and a specificity of 95% (95% C.I.: 75-100%). Further optimization of this assay could provide a highly useful tool for the diagnosis of MTC infection in exotic species.
外来物种结核病（TB）诊断测试的发展受到宿主生物学特性以及合适检测试剂供应有限的限制。因此，我们评估了一种基因表达检测方法（GEA），该方法易于针对新物种进行修改，并允许在野外条件下进行初始样本处理。使用单一比较皮内结核菌素试验对非洲水牛（非洲野水牛）进行分类，将试验阳性和试验阴性动物的血液在商用人类结核病检测试剂盒QuantiFERON(®)-TB Gold（In-Tube）（QFT）的“Nil”管（含生理盐水）和“TB抗原”管（含结核分枝杆菌复合群（MTC）特异性抗原）中孵育20小时。然后将血样用RNAlater(®)稳定，并运至实验室进行RNA提取。使用定制的TaqMan GEA计算TB抗原管中干扰素-γ（IFN-γ）mRNA相对于Nil管中的相对丰度，作为对MTC抗原识别的免疫激活标志物。比较了两组水牛的GEA结果，计算出区分这些组动物的临界值为2.85，灵敏度为80%（95%置信区间：56-94%），特异性为95%（95%置信区间：75-100%）。进一步优化该检测方法可为外来物种MTC感染的诊断提供非常有用的工具。
