Detection of field isolates of human and animal group C rotavirus by reverse transcription-polymerase chain reaction and digoxigenin-labeled oligonucleotide probes
Detection of field isolates of human and animal group C rotavirus by reverse transcription-polymerase chain reaction and digoxigenin-labeled oligonucleotide probes
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DOI：
10.1016/s0166-0934(99)00104-4
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发表时间：
1999-12-01
影响因子：
3.1
通讯作者：
Gentsch, JR
中科院分区：
文献类型：
--
作者：
Alfieri, AA;Leite, JPG;Gentsch, JR
轮状病毒（RV）是婴幼儿急性胃肠炎的重要病原体，也是世界范围内各种动物的主要病原体。这些病毒属于呼肠孤病毒科，含有11个双链RNA（dsRNA）片段的基因组。分别由基因组区段4和7、8或9编码的两种主要蛋白质VP 4和VP 7引起中和抗体应答，并形成目前将（gp）A组轮状病毒分为P（VP 4）和G（VP 7）血清型的基础。并且牛菌株不能广泛获得。在这项研究中，多重逆转录-聚合酶链反应（RT-PCR）和地高辛标记的寡核苷酸探针，使用化学发光已被开发用于检测和区分VP 7基因的培养适应和现场分离的人，猪和牛的gp C RV。多重RT-PCR和地高辛探针对人、猪和牛gp C RV的VP 7基因具有特异性，并允许检测和表征猪gp C RV与猪gp A或gp B轮状病毒的单一和混合感染。与聚丙烯酰胺凝胶电泳法相比，gp C RV的检出率大于50%。这些新的诊断方法可能有助于确定这些病毒在人类和动物感染中的流行病学重要性。(C)1999年由Elsevier Science B. V.出版，版权所有。
Rotaviruses (RV) are important etiological agents of acute gastroenteritis in infants and young children, as well as the young of a variety of animals worldwide. These viruses belong to Reoviridae family and contain a genome of 11 segments of double-stranded RNA (dsRNA). Two major proteins, VP4 and VP7, encoded by genome segments 4 and 7, 8 or 9, respectively, evoke a neutralizing antibody response and form the basis for the current classification of group (gp) A rotavirus into P (VP4) and G (VP7) serotypes, Although much recent progress has been made on the molecular biology of gp C RV, routine methods to detect and discriminate human, porcine, and bovine strains are not available widely. In this study, a multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) and digoxigenin-labeled (dig) oligonucleotide probes using chemiluminescence has been developed to detect and discriminate VP7 genes from culture-adapted and field isolates of human, porcine and bovine gp C RV. The multiplex RT-PCR and dig-probes were specific for the VP7 genes of human, porcine and bovine gp C RV and allowed detection and characterization of single and mixed infections of porcine gp C RV with porcine gp A or gp B rotaviruses. Detection rates for gp C RV were more than 50% when compared with polyacrylamide gel electrophoresis. These new diagnostic assays may help determine the epidemiological importance of these viruses in human and animal infections. (C) 1999 Published by Elsevier Science B.V. All rights reserved.