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比较不同的 RT-qPCR 检测方法对人源和牛源 A 组轮状病毒的检测效果,并对编码 VP4 和 VP7 衣壳蛋白的基因序列分析进行特征描述。

Comparison of different RT-qPCR assays for the detection of human and bovine group A rotaviruses and characterization by sequences analysis of genes encoding VP4 and VP7 capsid proteins.

机构信息

Agriculture and Agri-Food Canada, Food Research and Development Centre, Saint-Hyacinthe, QC, Canada.

出版信息

J Appl Microbiol. 2013 May;114(5):1435-48. doi: 10.1111/jam.12165. Epub 2013 Mar 1.

Abstract

AIMS

The aim of this study was to compare the performance of four RT-qPCR assays for the detection of human and bovine group A rotaviruses and to characterize the positive samples by sequence analysis of VP4 and VP7 genes.

METHODS AND RESULTS

RNA extracted from eight human rotavirus strains, and a panel of 33 human and 25 bovine faecal samples was subjected to different RT-qPCR detection systems. Among these assays, only RT-qPCR primers and probe systems B and C were able to detect all human rotavirus strains from cell culture solutions and faecal samples. However, the results showed that the system C was generally more sensitive by one or two logs than the other RT-qPCR assays tested. With the bovine faecal samples, the most efficient RT-qPCR systems were B and A with the detection in 100 and 92% of samples tested, respectively. Human group A rotavirus G1P[8] and bovine G6P[11] were the most frequently used strains identified in this study. A G3P[9] strain, closely related to a feline rotavirus isolated in the USA, was also discovered in a human rotavirus infection.

CONCLUSION

The RT-qPCR system B was the only TaqMan assay evaluated in this study able to detect rotavirus RNA in all positive human and bovine faecal samples.

SIGNIFICANCE AND IMPACT OF THE STUDY

Utilization of only one RT-qPCR for the detection of human and bovine group A rotaviruses and the possibility of human infection by a feline rotavirus strain.

摘要

目的

本研究旨在比较四种实时 RT-qPCR 检测方法在检测人源和牛源 A 组轮状病毒中的性能,并通过 VP4 和 VP7 基因序列分析对阳性样本进行特征分析。

方法与结果

从 8 株人轮状病毒株和 33 个人源和 25 个牛源粪便样本中提取 RNA,然后采用不同的 RT-qPCR 检测系统进行检测。在这些检测方法中,只有 RT-qPCR 引物和探针系统 B 和 C 能够从细胞培养液和粪便样本中检测到所有的人轮状病毒株。然而,结果表明,系统 C 通常比其他测试的 RT-qPCR 检测方法灵敏 1 到 2 个对数级。对于牛源粪便样本,最有效的 RT-qPCR 系统是 B 和 A,其检测效率分别为 100%和 92%。本研究中最常发现的人源 A 组轮状病毒 G1P[8]和牛源 G6P[11]。还在人轮状病毒感染中发现了一株与美国分离的猫源轮状病毒密切相关的 G3P[9]株。

结论

在本研究中评估的 RT-qPCR 系统 B 是唯一能够检测所有阳性人源和牛源粪便样本中轮状病毒 RNA 的 TaqMan 检测方法。

研究意义与影响

该研究结果提示仅使用一种 RT-qPCR 即可同时检测人源和牛源 A 组轮状病毒,以及人类感染猫源轮状病毒的可能性。

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