Sunlight Inactivation of Human Norovirus and Bacteriophage MS2 Using a Genome-Wide PCR-Based Approach and Enzyme Pretreatment
Sunlight Inactivation of Human Norovirus and Bacteriophage MS2 Using a Genome-Wide PCR-Based Approach and Enzyme Pretreatment
复制标题
DOI：
10.1021/acs.est.1c01575
复制
发表时间：
2021-06-08
影响因子：
11.4
通讯作者：
Boehm, Alexandria B.
中科院分区：
文献类型：
--
作者：
Loeb, Stephanie K.;Jennings, Wiley C.;Boehm, Alexandria B.
人诺如病毒（hNoV）是胃肠道疾病的重要病原体，可通过摄入受污染的水传播。目前培养hNoV是不切实际的，hNoV检测依赖于基于实时聚合酶链反应（RT-PCR）的方法。这种方法无法区分感染性病毒和灭活病毒，因为即使基因组的其他区域存在损伤，RNA基因组的完整区域也可以扩增，或者因为感染性病毒体中不包含完整的遗传物质。在本文中，我们采用多个长扩增子RT-qPCR外推方法来测定全基因组损伤和酶预处理，以研究模拟阳光对hNoV在无致敏剂的清澈水中的感染性的影响。使用MS 2大肠杆菌噬菌体作为内部控制，全基因组损伤外推法，以前成功地应用于UV-254灭活，大大高估了阳光灭活，表明在不同的光谱条件下的光灭活的关键差异。与MS 2基因组RNA相比，hNoV基因组RNA对每个碱基的模拟日光降解更敏感，而酶促预处理表明hNoV经历了比MS 2更多的衣壳损伤。这项工作提供了实际和机制的洞察到内源性的阳光灭活单链RNA噬菌体MS 2，广泛使用的替代品，和hNoV GII.4悉尼，一个重要的健康相关的病毒，在明确的无敏化剂的水。
Human norovirus (hNoV) is an important etiology of gastrointestinal illness and can be transmitted via ingestion of contaminated water. Currently impractical to culture, hNoV detection is reliant on real-time polymerase chain reaction (RT-PCR)-based methods. This approach cannot distinguish between infective and inactivated viruses because intact regions of the RNA genome can amplify even if the damage is present in other regions of the genome or because intact genetic material is not contained within an infectious virion. Herein, we employ a multiple long-amplicon RT-qPCR extrapolation approach to assay genome-wide damage and an enzymatic pretreatment to study the impact of simulated sunlight on the infectivity of hNoV in clear, sensitizer-free water. Using MS2 coliphage as an internal control, the genome-wide damage extrapolation approach, previously successfully applied for UV-254 inactivation, vastly overestimated sunlight inactivation, suggesting key differences in photoinactivation under different spectral conditions. hNoV genomic RNA was more susceptible to simulated sunlight degradation per base compared to MS2 genomic RNA, while enzymatic pretreatment indicated that hNoV experienced more capsid damage than MS2. This work provides practical and mechanistic insight into the endogenous sunlight inactivation of single-stranded RNA bacteriophage MS2, a widely used surrogate, and hNoV GII.4 Sydney, an important health-relevant virus, in clear sensitizer-free water.