Mazalovska Milena, Varadinov Nikola, Koynarski Tsvetoslav, Minkov Ivan, Teoharov Pavel, Lomonossoff George P, Zahmanova Gergana
Department of Plant Physiology and Molecular Biology, University of Plovdiv "Paisii Hilendarski", Plovdiv, Bulgaria.
Department of Animal Genetics, Faculty of Veterinary Medicine, Trakia University, Stara Zagora, Bulgaria.
Ann Lab Med. 2017 Jul;37(4):313-319. doi: 10.3343/alm.2017.37.4.313.
Hepatitis E virus (HEV) causes epidemics in developing countries and is primarily transmitted through the fecal-oral route. There have been recent reports on the zoonotic spread of the virus, and several animal species, primarily pigs, have been recognized as reservoirs of HEV. Because of its possible spread, there is an urgent need of a method for the cost-effective production of HEV proteins that can be used as diagnostic antigens for the serological detection of anti-HEV antibodies.
The HEV open reading frame (ORF)2 protein was purified from plant tissue by using immobilized metal-anion chromatography (IMAC). The recombinant protein was used to develop an in-house ELISA for testing anti-HEV antibodies in both human and swine sera. Thirty-six serum samples collected from patients with serologically proven HEV infection with commercial kits were tested for anti-HEV IgG antibodies by using the plant-expressed protein. Forty-five serum samples collected from apparently healthy pigs in Bulgarian farms were also tested.
We confirmed the transient expression and purification of a truncated version of the HEV genotype 3 capsid protein in Nicotiana benthamiana and its usefulness as a diagnostic antigen. ELISA showed the presence of anti-HEV IgG antibodies in 29 of the 36 human samples. The in-house ELISA showed anti-HEV IgG antibodies in 34 of the 45 pigs.
We describe a method for the production of HEV ORF2 protein in N. benthamiana and the usefulness of this protein for the serological detection of anti-HEV antibodies in both humans and swine.
戊型肝炎病毒（HEV）在发展中国家引发流行病，主要通过粪-口途径传播。最近有关于该病毒人畜共患传播的报道，几种动物物种，主要是猪，已被确认为HEV的宿主。由于其可能的传播，迫切需要一种经济高效生产HEV蛋白的方法，该蛋白可作为诊断抗原用于抗HEV抗体的血清学检测。
通过固定化金属阴离子色谱法（IMAC）从植物组织中纯化HEV开放阅读框（ORF）2蛋白。该重组蛋白用于开发一种内部ELISA，以检测人和猪血清中的抗HEV抗体。使用植物表达的蛋白对36份从经商业试剂盒血清学证实为HEV感染的患者收集的血清样本检测抗HEV IgG抗体。还检测了从保加利亚农场明显健康的猪收集的45份血清样本。
我们证实了HEV基因型3衣壳蛋白截短版本在本氏烟草中的瞬时表达和纯化，以及其作为诊断抗原的用途。ELISA显示36份人类样本中有29份存在抗HEV IgG抗体。内部ELISA显示45头猪中有34头存在抗HEV IgG抗体。
我们描述了一种在本氏烟草中生产HEV ORF2蛋白的方法，以及该蛋白在人和猪抗HEV抗体血清学检测中的用途。
