Department of Food Science and Technology, School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai 200240, China.
State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510030, China.
Viruses. 2021 Jan 6;13(1):65. doi: 10.3390/v13010065.
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.
人类诺如病毒（HuNoVs）是全球食源性疾病的主要原因之一。病毒基因组是病毒溯源和病毒传播模式监测的最基本信息。然而，由于不同基因型之间的序列异质性和样本中低滴度，HuNoVs 的全基因组测序仍然具有挑战性。为了解决这一需求，本研究建立了转座酶辅助 RNA/DNA 杂交共标记（TRACE-seq）方法，用于 HuNoVs 的下一代测序文库制备。我们的数据表明，从所有 11 个临床样本中几乎可以获得整个 HuNoVs 基因组（>7 kb）。涉及 12 种基因型，包括 GI.3、GI.4、GI.5、GI.8、GII.2、GII.3、GII.4、GII.6、GII.12、GII.13、GII.14 和 GII.21。与传统的病毒宏基因组文库制备方法相比，优化后的 TRACE-seq 大大减少了宿主和细菌 RNA 的干扰。此外，通过使用更少的原始数据和足够的全基因组深度，可以组装病毒基因组序列。因此，该方法具有高通用性和可靠性，有望实现整个病毒基因组的获得。对于那些在体外难以培养、与复杂宿主背景混合且滴度较低的病毒，如本研究中使用的 HuNoVs，该方法特别适用。
