Boonham N, Walsh K, Smith P, Madagan K, Graham I, Barker I
Central Science Laboratory, Sand Hutton, YO41 1LZ, York, UK.
J Virol Methods. 2003 Mar;108(2):181-7. doi: 10.1016/s0166-0934(02)00284-7.
Currently, most diagnostic methodology is geared towards detection of a very specific target species and often a number of assays need to be run in parallel to reach a result. The generic methods that are available for virus testing tends to give identification to the genus level only. The method described in this paper addresses this problem by exploiting a technology that has potential to test for a large number of targets in a single assay. Using the array constructed, the method was able to detect several common potato viruses (PVY, PVX, PVA, PVS) in single and mixed infections. The method was shown to be able to discriminate sequences with less than 80% sequence identity but was able to detect sequence variants with greater than 90% sequence identity. Thus the method should be useful for discriminating at the species level, but able to cope well with the intrinsic variability found within the genomes of RNA viruses. The sensitivity of the assay was found to be comparable with ELISA. The paper illustrates a significant step forward in the development of diagnostic methodologies by presenting for the first time a method that could theoretically be used not just for viruses, but for all the plant pathogens and pests that a modern diagnostic laboratory would want to test for, in a single completely generic and highly parallel format.
目前,大多数诊断方法都针对检测非常特定的目标物种,而且通常需要同时进行多项检测才能得出结果。现有的病毒检测通用方法往往只能鉴定到属的水平。本文所述方法通过利用一种有潜力在单次检测中检测大量目标的技术解决了这一问题。使用构建的阵列,该方法能够检测单种感染和混合感染中的几种常见马铃薯病毒(马铃薯Y病毒、马铃薯X病毒、马铃薯A病毒、马铃薯S病毒)。结果表明该方法能够区分序列同一性低于80%的序列,但能够检测序列同一性高于90%的序列变体。因此,该方法在物种水平的区分上应该是有用的,并且能够很好地应对RNA病毒基因组中存在的内在变异性。该检测方法的灵敏度与酶联免疫吸附测定法相当。本文首次提出了一种理论上不仅可用于病毒,还可用于现代诊断实验室想要检测的所有植物病原体和害虫的方法,以单一完全通用且高度并行的形式呈现,这说明诊断方法的发展向前迈出了重要一步。