Vogels Chantal B F, Hill Verity, Breban Mallery I, Chaguza Chrispin, Paul Lauren M, Sodeinde Afeez, Taylor-Salmon Emma, Ott Isabel M, Petrone Mary E, Dijk Dennis, Jonges Marcel, Welkers Matthijs R A, Locksmith Timothy, Dong Yibo, Tarigopula Namratha, Tekin Omer, Schmedes Sarah, Bunch Sylvia, Cano Natalia, Jaber Rayah, Panzera Charles, Stryker Ian, Vergara Julieta, Zimler Rebecca, Kopp Edgar, Heberlein Lea, Morrison Andrea M, Michael Scott F, Grubaugh Nathan D
Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
Yale Institute for Global Health, Yale University, New Haven, Connecticut, United States of America.
medRxiv. 2023 Oct 13:2023.10.13.23296997. doi: 10.1101/2023.10.13.23296997.
The increasing burden of dengue virus on public health due to more explosive and frequent outbreaks highlights the need for improved surveillance and control. Genomic surveillance of dengue virus not only provides important insights into the emergence and spread of genetically diverse serotypes and genotypes, but it is also critical to monitor the effectiveness of newly implemented control strategies. Here, we present DengueSeq, an amplicon sequencing protocol, which enables whole-genome sequencing of all four dengue virus serotypes.
We developed primer schemes for the four dengue virus serotypes, which can be combined into a pan-serotype approach. We validated both approaches using genetically diverse virus stocks and clinical specimens that contained a range of virus copies. High genome coverage (>95%) was achieved for all genotypes, except DENV2 (genotype VI) and DENV 4 (genotype IV) sylvatics, with similar performance of the serotype-specific and pan-serotype approaches. The limit of detection to reach 70% coverage was 10-10 RNA copies/μL for all four serotypes, which is similar to other commonly used primer schemes. DengueSeq facilitates the sequencing of samples without known serotypes, allows the detection of multiple serotypes in the same sample, and can be used with a variety of library prep kits and sequencing instruments.
DengueSeq was systematically evaluated with virus stocks and clinical specimens spanning the genetic diversity within each of the four dengue virus serotypes. The primer schemes can be plugged into existing amplicon sequencing workflows to facilitate the global need for expanded dengue virus genomic surveillance.
登革病毒对公共卫生造成的负担日益加重，疫情爆发愈发频繁且剧烈，这凸显了加强监测与控制的必要性。对登革病毒进行基因组监测不仅能深入了解基因多样的血清型和基因型的出现与传播情况，对于监测新实施的控制策略的效果也至关重要。在此，我们介绍了DengueSeq，一种扩增子测序方案，可实现对所有四种登革病毒血清型进行全基因组测序。
我们针对四种登革病毒血清型开发了引物方案，这些方案可组合成一种泛血清型方法。我们使用基因多样的病毒株和含有不同病毒拷贝数的临床标本对这两种方法进行了验证。除了登革病毒2型（基因型VI）和登革病毒4型（基因型IV）的丛林型毒株外，所有基因型均实现了高基因组覆盖率（>95%），血清型特异性方法和泛血清型方法的性能相似。所有四种血清型达到70%覆盖率的检测限均为10-10 RNA拷贝/μL，这与其他常用引物方案相似。DengueSeq便于对血清型未知的样本进行测序，能够检测同一样本中的多种血清型，并且可与多种文库制备试剂盒和测序仪器配合使用。
我们使用涵盖四种登革病毒血清型各自遗传多样性的病毒株和临床标本对DengueSeq进行了系统评估。这些引物方案可接入现有的扩增子测序工作流程，以满足全球对扩大登革病毒基因组监测的需求。
