Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
Infectious Diseases and Tropical Medicine Research Center, Shahid Beheshti University of Medial Sciences, Tehran, Iran.
Semin Dial. 2021 Sep;34(5):332-337. doi: 10.1111/sdi.12981. Epub 2021 May 3.
Toxoplasmosis causes serious complications in immunocompromised and pregnant women. Serological tests for the detection of toxoplasmosis are often designed from parasitic tachyzoites antigens. The process of producing these antigens is very difficult. The purpose of this study was evaluation of T. gondii-rGRA5 for the immunodiagnosis and molecular detection of Toxoplasma infection using enzyme-linked immunosorbent assay (ELISA) and LAMP methods in hemodialysis patients. The GRA5 gene was successfully expressed and purified by affinity chromatography assay and evaluated by western blot. Then it was used to design an ELISA assay. A total of 260 samples were tested for anti-Toxoplasma IgG and IgM antibodies using a commercial ELISA kit and designed ELISA kit. Finally, the LAMP method was used to evaluate the precision and reliability of the results obtained by commercial and designed ELISA kits. The consistency of the results of two methods was analyzed using the Kappa coefficient of agreement. The rGRA5 revealed higher immunoreactivity with 1:100 dilution of sera from toxoplasmosis patients. The specificity and sensitivity of the assay were 93% and 96%, respectively. According to the Kappa coefficient, there was a substantial correlation between the results of ELISA and LAMP based on rGRA5 (≈98%, p < 0.001). Also it showed that rGRA5 protein can be used as an antigenic protein for designing sero-diagnostic tests to identify Toxoplasma infection especially in hemodialysis patients.
弓形虫病可导致免疫功能低下和孕妇出现严重并发症。用于检测弓形虫病的血清学检测通常基于寄生虫速殖子抗原设计。这些抗原的生产过程非常困难。本研究旨在通过酶联免疫吸附试验（ELISA）和环介导等温扩增（LAMP）方法评估 T. gondii-rGRA5 对血液透析患者弓形虫感染的免疫诊断和分子检测。GRA5 基因通过亲和层析试验成功表达和纯化，并通过 Western blot 进行评估。然后，它被用于设计 ELISA 检测。使用商业 ELISA 试剂盒和设计的 ELISA 试剂盒对 260 份样本进行抗弓形虫 IgG 和 IgM 抗体检测。最后，使用 LAMP 方法评估商业和设计 ELISA 试剂盒获得结果的精度和可靠性。使用一致性系数分析两种方法结果的一致性。rGRA5 与弓形虫病患者血清的 1:100 稀释液具有更高的免疫反应性。该检测的特异性和灵敏度分别为 93%和 96%。根据 Kappa 系数，基于 rGRA5 的 ELISA 和 LAMP 结果之间存在显著相关性（≈98%，p<0.001）。此外，结果表明 rGRA5 蛋白可用作设计血清学诊断检测的抗原蛋白，以鉴定弓形虫感染，尤其是在血液透析患者中。
