Evaluation of the Dako IDEIA norovirus EIA assay for detection of norovirus using faecal specimens from Australian gastroenteritis outbreaks
Evaluation of the Dako IDEIA norovirus EIA assay for detection of norovirus using faecal specimens from Australian gastroenteritis outbreaks
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使用澳大利亚胃肠炎爆发的粪便样本检测诺如病毒的 Dako IDEIA 诺如病毒 EIA 测定法的评估
DOI：
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发表时间：
2006
期刊：
影响因子：
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通讯作者：
J. Marshall
中科院分区：
文献类型：
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作者：
A. Dimitriadis;L. Bruggink;J. Marshall
目的：检测诺沃克病毒的新技术需要不断评估，诺沃克病毒是胃肠炎的主要原因。这项研究的目的是使用澳大利亚维多利亚州胃肠炎暴发的材料，通过光度分析和目视分析来评估达科诺沃克病毒EIA检测的敏感性和特异性。方法：采用电子显微镜(EM)、两轮多重逆转录聚合酶链式反应(RT-PCR)和IDEIA诺如病毒检测方法，对41起胃肠炎暴发的130份粪便标本进行诺沃克病毒检测。对所有扩增产物充足的标本进行测序，以确定其诺如病毒的基因型别。此外，还使用六种成熟的RT-PCR方法检测了四个EIA阳性、多重RT-PCR/EM阴性的标本。此外，还使用了一系列RT-PCR方案来检测只有多重RT-PCR检测GII阳性，而EIA检测GI和GII阳性的标本。在另外7个样本上测试了多次冻融循环对EIA阳性的影响。另外七个已知含有胃肠炎病毒、腺病毒、星状病毒和轮状病毒的样本也进行了IDIA诺如病毒检测。结果：IDEIA诺如病毒检测单份标本的灵敏度和特异度分别为66%和85%(目测分析与多重RT-PCR相比)、63%和88%(光度分析与多重RT-PCR相比)、65%和87%(目测分析与多重RT-PCR和/或EM比较)和62%和90%(光度分析与多重RT-PCR和/或EM相比)。经多重RT-PCR和/或EM检测为阴性的4例EIA阳性标本，在6种可供选择的RT-PCR方法中无一例阳性。EIA检测GI和GII阳性，但仅多重RT-PCR检测GII阳性的标本，在任何替代RT-PCR方法中均未检测到GI。每次暴发至少要对三个样本进行环境影响评估，以确保诺沃克病毒阳性暴发(多重RT-PCR和/或EM)通过IDEIA诺沃克病毒检测(目测或光度分析)被归类为诺沃克病毒阳性。然而，一次诺沃克病毒阴性暴发(多重RT-PCR和/或EM)提供了四个样本，其中一个样本通过IDEIA诺沃克病毒检测呈诺沃克病毒阳性。通过开放阅读框架1测序鉴定了7种诺如病毒基因，并对所有7种诺沃克病毒基因(以及1份EM阳性/多重RT-PCR阴性的标本)进行了目测和光度分析。反复冷冻-解冻粪便样本(最多六次)不会降低EIA分析的灵敏度，但会使EIA阴性的样本EIA呈阳性。萨普病毒、腺病毒、星状病毒和轮状病毒阳性的标本EIA均为阴性。结论：IDEIA诺如病毒检测缺乏将特定结果归因于特定样本的敏感性和特异性，但在样本丰富的胃肠炎暴发中检测诺如病毒是有用的，尽管很难避免假阳性的风险。由于目测分析几乎可以与光度分析一样可靠地用于结果评估，因此，对于缺乏光度微板阅读器等专门设备的实验室来说，该试剂盒将是有用的。
Aims: New techniques for detection of norovirus, a major cause of gastroenteritis, require ongoing evaluation. The aim of this study was to use material from gastroenteritis outbreaks in Victoria, Australia, to evaluate the sensitivity and specificity of the Dako IDEIA norovirus EIA assay, using both photometric and visual analysis. Methods: A total of 130 faecal specimens from 41 gastroenteritis outbreaks were tested for norovirus by electron microscopy (EM), a two‐round multiplex reverse transcription‐polymerase chain reaction (RT‐PCR) method and the IDEIA norovirus assay. All specimens with sufficient amplified product were sequenced to determine their norovirus genotype. In addition, six well‐established RT‐PCR protocols were used to test four EIA‐positive, multiplex RT‐PCR/EM‐negative specimens. Also, a range of RT‐PCR protocols was used to test a specimen positive for GII only by the multiplex RT‐PCR but positive for GI and GII by the EIA. The effect of multiple freezing‐thawing cycles on EIA positivity was tested on seven additional specimens. A further seven specimens, known to contain the gastroenteritis viruses sapovirus, adenovirus, astrovirus and rotavirus were also tested by the IDEIA norovirus assay. Results: The IDEIA norovirus assay gave a single‐specimen sensitivity and specificity of 66% and 85%, respectively (visual analysis compared with the multiplex RT‐PCR), 63% and 88% (photometric analysis compared with the multiplex RT‐PCR), 65% and 87% (visual analysis compared with the multiplex RT‐PCR and/or EM) and 62% and 90% (photometric analysis compared with the multiplex RT‐PCR and/or EM). None of the four EIA‐positive specimens negative by the multiplex RT‐PCR and/or EM was positive by any of the six alternative RT‐PCR protocols. The specimen positive for GI and GII by EIA but for GII only by the multiplex RT‐PCR was not positive for GI by any of the alternative RT‐PCR protocols. A minimum of three specimens per outbreak had to be tested by the EIA to ensure that norovirus‐positive outbreaks (multiplex RT‐PCR and/or EM) were classified as positive for norovirus by the IDEIA norovirus assay (visual or photometric analysis). However, one specimen from a norovirus‐negative outbreak (multiplex RT‐PCR and/or EM) for which four specimens were provided was positive for norovirus by the IDEIA norovirus assay. Seven norovirus genotypes were identified by open reading frame 1 sequencing analysis and specimens from all seven norovirus genotypes (as well as an EM‐positive/multiplex RT‐PCR‐negative specimen) were detected by the IDEIA norovirus assay by both visual and photometric analysis. Repeated freezing‐thawing cycles (up to six) for faecal specimens did not reduce the sensitivity of the EIA assay but could render an EIA‐negative specimen EIA‐positive. The specimens positive for sapovirus, adenovirus, astrovirus and rotavirus were EIA‐negative. Conclusions: The IDEIA norovirus assay lacks the sensitivity and specificity to ascribe a particular result to a particular specimen, but could be useful for detecting norovirus in a gastroenteritis outbreak where specimens are plentiful, although it is difficult to avoid a risk of false positives. Since visual analysis can be used for result assessment almost as reliably as photometric analysis, the test kit would be useful for laboratories lacking specialist equipment such as a photometric microplate reader.
通过 Rotazyme 轮状病毒 ELISA 检测实验室啮齿动物标本中非特异性反应的鉴定。
DOI：
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发表时间：
1988
期刊：
Laboratory animal science
影响因子：
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作者：
Jure,MN;Morse,SS;Stark,DM
通讯作者：
Stark,DM