Abstract
Human noroviruses (HuNoVs) are one of the leading causes of foodborne illnesses globally. The viral genome is the most essential information for viral source tracing and viral transmission pattern monitoring. However, whole genome sequencing of HuNoVs is still challenging due to the sequence heterogeneity among different genotypes and low titer in samples. To address this need, in this study, the Transposase assisted RNA/DNA hybrid Co-tagmentation (TRACE-seq) method was established for next generation sequencing library preparation of HuNoVs. Our data demonstrated that almost the whole HuNoVs genome (>7 kb) could be obtained from all of the 11 clinical samples tested. Twelve genotypes including GI.3, GI.4, GI.5, GI.8, GII.2, GII.3, GII.4, GII.6, GII.12, GII.13, GII.14, and GII.21 were involved. Compared with the traditional method for viral metagenomics library preparation, optimized TRACE-seq greatly reduced the interference from the host’s and bacterial RNAs. In addition, viral genome sequences can be assembled by using less raw data with sufficient depth along the whole genome. Therefore, for the high versatility and reliability, this method is promising for whole viral genome attainment. It is particularly applicable for the viruses with a low titer that are mixed with a complicated host background and are unable to be cultured in vitro, like the HuNoVs utilized in this study.
Highlights
Human noroviruses (HuNoVs) are the leading cause of non-bacterial epidemic gastroenteritis globally and cause an estimated 684 million illnesses of acute gastroenteritis worldwide, resulting in approximately $60 billion in societal costs [1]
All the sequences were above 70.0, indicating that the assembled sequences are of high similarity with the norovirus sequences in the database
Metagenomic sequencing with spiked primer enrichment (MSSPE) enriches targeted viral RNA sequences by using spiked primers [38]
Summary
Human noroviruses (HuNoVs) are the leading cause of non-bacterial epidemic gastroenteritis globally and cause an estimated 684 million illnesses of acute gastroenteritis worldwide, resulting in approximately $60 billion in societal costs [1]. A norovirus (NoV) has a single-stranded, positive-sense genomic RNA of approximately 7.5 kb, that have a 5 -terminal virus protein, are genome-linked (VPg), and possess 3 -terminal poly (A) [2]. Based on phylogenetic analysis of major capsid protein (VP1) amino acid sequences, at least 10 norovirus genogroups (GI-GX) and 48 genotypes have been recognized [3]. The uncorrected pairwise distance ranges for strain, genotype, and genogroup were 0–14.1%, 14.3–43.8%, and 44.9–61.4%, respectively [4]. To prevent and control the HuNoVs outbreaks, the key is to identify the viral pollution source and monitor viral transmission patterns based on the viral genome sequences [6]
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