Department of Parasitology and Mycology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.
Infectious and Tropical Diseases Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
Iran Biomed J. 2022 Nov 1;26(5):374-9. doi: 10.52547/ibj.3801.
Cryptosporidium parvum is an important coccidian parasite infecting many mammals, including human. This parasite can manifest as chronic severe diarrhea in immunocompromised individuals, especially those with AIDS. The present study reports the recombinant production of recombinant (r)P2 and rP23 antigens of C. parvum as antigens for detecting human cryptosporidiosis using indirect ELISA tests.
The coding sequences of rP2 and rP23 proteins were codon-optimized, commercially synthesized and sub-cloned in the pET28a expression vector. The expressed proteins were purified by Ni-NTA column chromatography and confirmed by Western blotting. The efficacy of rP2/rP23 proteins for serodiagnosis was evaluated by positive (n = 20) and negative (n = 20) human sera, confirmed by the Ziehl-Neelsen staining as the gold standard test.
In ELISA test, the sera from C. parvum-infected patients reacted strongly to rP2/rP23. The sensitivity and specificity related to the diagnostic potential of rP2/rP23 in the ELISA assay were 100%.
Our results showed that combination of rP23 and rP2 antigens in ELISA significantly increases the performance of C. parvum serodiagnosis in human cryptosporidiosis.
微小隐孢子虫是一种重要的球虫寄生虫，感染许多哺乳动物，包括人类。这种寄生虫会在免疫功能低下的个体中表现为慢性严重腹泻，尤其是艾滋病患者。本研究报告了微小隐孢子虫重组（r）P2 和 rP23 抗原的重组生产，作为使用间接 ELISA 检测人类隐孢子虫病的抗原。
rP2 和 rP23 蛋白的编码序列经密码子优化后，商业合成并亚克隆到 pET28a 表达载体中。通过 Ni-NTA 柱层析纯化表达蛋白，并通过 Western blot 进行验证。通过 Ziehl-Neelsen 染色作为金标准检测，用阳性（n=20）和阴性（n=20）人血清评估 rP2/rP23 蛋白在血清学诊断中的效果。
在 ELISA 试验中，微小隐孢子虫感染患者的血清与 rP2/rP23 反应强烈。rP2/rP23 在 ELISA 检测中的诊断潜力的敏感性和特异性相关为 100%。
我们的结果表明，rP23 和 rP2 抗原在 ELISA 中的联合应用显著提高了微小隐孢子虫在人类隐孢子虫病中的血清学诊断性能。
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