Abstract：
Toxoplasma gondii is a protozoan parasite that may infect many mammals including humans. Cats are one of the main sources of infection for humans. Therefore, routine screening of cats with tests that are inexpensive, rapid, and do not require sophisticated laboratory equipment is important. In this study, a lateral flow assay (LFA) was designed to rapidly diagnose toxoplasmosis in cats. For this purpose, we selected GRA1 protein of T. gondii due to its high antigenicity in diagnostic and vaccine studies. We further analyzed the immunological properties of GRA1 protein using in silico tools. Then, we expressed and purified recombinant GRA1 (rGRA1) protein and used it during the development of LFA to detect toxoplasmosis in serum samples (n = 40) of cats. According to the results, rGRA1 protein has negative GRAVY value, high aliphatic index, alpha helix, random coil and 12 B cell epitopes. The in silico data supported the high antigenic properties of rGRA1 protein and showed that it can be a good antigen candidate for LFA. Among 30 cat positive serum samples, 27 were found positive by the LFA while seronegative sera (n = 10) were negative by the LFA. The preliminary data showed that the LFA has high sensitivity (90 %) and specificity (100 %). When we used high responsive cat sera (i.e. sera that have optical density > 0.5 with ELISA) the sensitivity value reached 100 %. These results showed that rGRA1 protein is a good candidate to develop a LFA for rapid diagnosis of toxoplasmosis in cats.
摘要：
弓形虫是一种原生动物寄生虫，可以感染包括人类在内的许多哺乳动物。猫是人类感染的主要来源之一。因此,用廉价的测试对猫进行常规筛查，快速,并且不需要精密的实验室设备很重要。在这项研究中,设计了一种侧流测定法（LFA）来快速诊断猫的弓形虫病。为此,我们选择了弓形虫的GRA1蛋白，因为它在诊断和疫苗研究中具有很高的抗原性。我们使用计算机工具进一步分析了GRA1蛋白的免疫学特性。然后，我们表达并纯化了重组GRA1（rGRA1）蛋白，并在LFA的开发过程中使用它来检测猫血清样品（n=40）中的弓形虫病。根据结果,rGRA1蛋白具有负GRAVY值，高脂肪指数，阿尔法螺旋,无规卷曲和12个B细胞表位。计算机上的数据支持rGRA1蛋白的高抗原特性，并表明它可以是LFA的良好抗原候选物。在30份猫阳性血清样本中，LFA发现27个为阳性，而LFA血清阴性血清（n=10）为阴性。初步数据显示LFA具有高灵敏度(90%)和特异性(100%)。当我们使用高响应猫血清(即用ELISA具有>0.5的光密度的血清)时,灵敏度值达到100%。这些结果表明，rGRA1蛋白是开发LFA以快速诊断猫弓形虫病的良好候选者。
