Chen Xingwen, Zheng Chunfu, Wu Fuqing
Department of Environmental and Occupational Health Sciences, University of Texas Health Science Center at Houston School of Public Health, Houston, TX, USA.
Department of Epidemiology, University of Texas Health Science Center at Houston School of Public Health, Houston, TX, USA.
Methods Mol Biol. 2025;2940:319-328. doi: 10.1007/978-1-0716-4615-1_27.
The surveillance of SARS-CoV-2 is significantly limited by the presence of numerous variants with distinct behaviors driven by rapid evolution. Although the conventional clinical genome sequencing method provides comprehensive data, it has limitations in capturing the epidemiological dynamics of the variants circulating in the community. In this chapter, we describe a protocol using RBD amplicon sequencing of wastewater samples. We present the detailed experimental procedure for wastewater sample processing, robust nested-PCR-based amplification of the target RBD sequence, and library preparation. Moreover, we describe a general pipeline of sequencing data processing and interpretation, showing the epidemiological and evolutionary trends of SARS-CoV-2 variants in the community.
严重急性呼吸综合征冠状病毒2（SARS-CoV-2）的监测受到众多具有不同行为的变体的显著限制，这些变体由快速进化驱动。尽管传统的临床基因组测序方法可提供全面的数据，但在捕捉社区中传播的变体的流行病学动态方面存在局限性。在本章中，我们描述了一种使用废水样本的受体结合域（RBD）扩增子测序的方案。我们展示了废水样本处理、基于稳健的巢式聚合酶链反应（PCR）扩增目标RBD序列以及文库制备的详细实验步骤。此外，我们描述了测序数据处理和解读的一般流程，展示了社区中SARS-CoV-2变体的流行病学和进化趋势。
