A novel multiplex RT-PCR for identification of VP6 subgroups of human and porcine rotaviruses
A novel multiplex RT-PCR for identification of VP6 subgroups of human and porcine rotaviruses
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DOI：
10.1016/j.jviromet.2010.05.013
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发表时间：
2010-09-01
影响因子：
3.1
通讯作者：
Maneekarn, Niwat
中科院分区：
文献类型：
--
作者：
Thongprachum, Aksara;Chaimongkol, Natthawan;Maneekarn, Niwat
VP6 蛋白抗原允许将轮状病毒分为至少四个亚组，具体取决于 SG 特异性表位的存在或不存在：SG I、SG II、SG (I+II) 和 SG non-(I+II)。然而，针对人和猪轮状病毒VP6蛋白上表位的MAb有时不识别SG特异性表位或识别不相关的SG表位，因此导致亚组分配不正确。为了解决这个问题，开发了一种新型多重 RT-PCR 作为替代工具，使用新设计的基因组 I 或 II 特异性引物来鉴定 VP6 基因组。通过对已知 SG I、SG II、SG (I+II) 和 SG non-(I+II) 株的人和猪轮状病毒进行测试，并与单一 RT-PCR 和 VP6 序列分析进行比较，评估了新开发的多重 RT-PCR 方法的灵敏度和特异性。结果表明，新的多重RT-PCR方法测定的人和猪轮状病毒的基因组与单一RT-PCR和VP6序列分析测定的基因组100％一致。该方法被证明特异、灵敏、耗时少，并且成功地对患有急性腹泻的儿童和仔猪中传播的轮状病毒临床分离株进行基因分型。 (C) 2010 Elsevier B.V. 保留所有权利。
VP6 protein antigens allow classification of rotaviruses into at least four subgroups, depending on the presence or absence of SG-specific epitopes: SG I, SG II, SG (I+II), and SG non-(I+II). However, MAbs against epitopes on the VP6 protein of human and porcine rotaviruses, sometimes, do not recognize SG-specific epitopes or recognize irrelevant-SG epitopes and therefore result in the incorrect assignment of subgroups. In order to solve this problem, a novel multiplex RT-PCR was developed as an alternative tool to identify VP6 genogroups using newly designed primers which are specific for genogroup I or II. The sensitivity and specificity of the newly developed multiplex RT-PCR method was evaluated by testing with human and porcine rotaviruses of known SG I, SG II, SG (I+II), and SG non-(I+II) strains in comparison with monoplex RT-PCR and VP6 sequence analysis. The results show that the genogroups of both human and porcine rotaviruses as determined by the new multiplex RT-PCR method were in 100% agreement with those determined by monoplex RT-PCR and VP6 sequence analysis. The method was shown to be specific, sensitive, less-time consuming, and successful in genogrouping clinical isolates of rotaviruses circulating in children and piglets with acute diarrhea. (C) 2010 Elsevier B.V. All rights reserved.