Ip Hon S, Wiley Michael R, Long Renee, Palacios Gustavo, Shearn-Bochsler Valerie, Whitehouse Chris A
U.S. Geological Survey, National Wildlife Health Center, Madison, WI, USA.
Center for Genomic Sciences, United States Army Medical Research Institute of Infectious Diseases, Frederick, MD, USA.
J Virol Methods. 2014 Sep;206:42-5. doi: 10.1016/j.jviromet.2014.05.018. Epub 2014 May 29.
Advances in massively parallel DNA sequencing platforms, commonly termed next-generation sequencing (NGS) technologies, have greatly reduced time, labor, and cost associated with DNA sequencing. Thus, NGS has become a routine tool for new viral pathogen discovery and will likely become the standard for routine laboratory diagnostics of infectious diseases in the near future. This study demonstrated the application of NGS for the rapid identification and characterization of a virus isolated from the brain of an endangered Mississippi sandhill crane. This bird was part of a population restoration effort and was found in an emaciated state several days after Hurricane Isaac passed over the refuge in Mississippi in 2012. Post-mortem examination had identified trichostrongyliasis as the possible cause of death, but because a virus with morphology consistent with a togavirus was isolated from the brain of the bird, an arboviral etiology was strongly suspected. Because individual molecular assays for several known arboviruses were negative, unbiased NGS by Illumina MiSeq was used to definitively identify and characterize the causative viral agent. Whole genome sequencing and phylogenetic analysis revealed the viral isolate to be the Highlands J virus, a known avian pathogen. This study demonstrates the use of unbiased NGS for the rapid detection and characterization of an unidentified viral pathogen and the application of this technology to wildlife disease diagnostics and conservation medicine.
大规模平行DNA测序平台(通常称为下一代测序(NGS)技术)的进展,已极大地减少了与DNA测序相关的时间、人力和成本。因此,NGS已成为发现新型病毒病原体的常规工具,并且在不久的将来可能会成为传染病常规实验室诊断的标准。本研究展示了NGS在快速鉴定和表征从一只濒危的密西西比沙丘鹤大脑中分离出的病毒方面的应用。这只鸟是种群恢复计划的一部分,于2012年艾萨克飓风席卷密西西比州的保护区几天后,被发现处于瘦弱状态。尸检确定毛圆线虫病可能是死因,但由于从这只鸟的大脑中分离出一种形态与披膜病毒一致的病毒,因此强烈怀疑是虫媒病毒病因。由于针对几种已知虫媒病毒的单独分子检测均为阴性,因此使用Illumina MiSeq进行无偏差NGS来明确鉴定和表征致病病毒病原体。全基因组测序和系统发育分析表明,该病毒分离株是高地J病毒,一种已知的禽类病原体。本研究展示了使用无偏差NGS快速检测和表征未鉴定的病毒病原体,以及该技术在野生动物疾病诊断和保护医学中的应用。