National Veterinary Research Institute in Puławy, Department of Parasitology and Invasive Diseases, Aleja Partyzantów 56, 24-100 Puławy, Poland.
USDA-Agricultural Research Service, Animal Parasitic Diseases Lab, BARC-East Building 1040, 10300 Baltimore Avenue, 10705 Beltsville, MD, USA.
Int J Parasitol. 2022 Feb;52(2-3):145-155. doi: 10.1016/j.ijpara.2021.08.002. Epub 2021 Sep 17.
Molecular epidemiology using traditional sequencing has been notoriously difficult in inbred parasites due to a lack of genetic variation available for discriminating among parasites. Next generation sequencing techniques offer a solution to this problem by increasing the number of loci that can be sequenced. Here, we introduce Trich-tracker, a tool that makes efficient use of diagnostic variation distributed throughout the genome of Trichinella spiralis to more rapidly, and conclusively, resolve connections and distinctions among focal outbreaks of T. spiralis. In particular, we rapidly characterised genetic variation among a sample of parasites from Polish farms and wildlife, sampling genomic variation using double digest restriction site-associated DNA sequencing (ddRADseq). Approximately 400,000 bases of sequence were generated from each sample and shown to be distributed across the genome with single nucleotide polymorphisms occurring at a frequency of approximately one base in 10,000. Both phylogenetic and Bayesian clustering analyses indicated that ddRADseq genotypes formed distinct clusters for specific outbreaks and were quite distinct from wild boar samples. Two of the investigated outbreaks were more similar to each other than to other outbreak samples, suggesting a link between these outbreaks. Hence, the Trich-tracker procedure identified informative genomic variation which afforded unprecedented epidemiological resolution. Trich-tracker is very flexible tool, quickly and inexpensively mining genomes of even highly inbred populations of T. spiralis to support outbreak investigations. The simplicity of the entire procedure, and time and cost effectiveness of Trich-tracker support its practical application in ongoing Trichinella outbreaks. The discriminating power of this tool is tunable and scalable, allowing application in a variety of epidemiological contexts, and is easily adapted to other parasite systems.
由于缺乏可用于区分寄生虫的遗传变异，传统测序的分子流行病学在近交寄生虫中一直是一个难题。下一代测序技术通过增加可测序的基因座数量为解决这个问题提供了一种解决方案。在这里，我们介绍了 Trich-tracker，这是一种工具，可以有效地利用分布在旋毛虫基因组中的诊断变异，更快速、更明确地解决旋毛虫焦点暴发之间的联系和区别。特别是，我们快速描述了来自波兰农场和野生动物的寄生虫样本的遗传变异，使用双酶切限制位点相关 DNA 测序（ddRADseq）对基因组变异进行采样。从每个样本中生成了大约 40 万个碱基的序列，并证明这些序列分布在基因组中，单核苷酸多态性的发生频率约为每 10000 个碱基一个。系统发育和贝叶斯聚类分析都表明，ddRADseq 基因型为特定暴发形成了独特的聚类，与野猪样本有很大的不同。两个被调查的暴发比其他暴发样本更相似，这表明这些暴发之间存在联系。因此，Trich-tracker 程序确定了具有信息性的基因组变异，从而提供了前所未有的流行病学分辨率。Trich-tracker 是一种非常灵活的工具，可以快速、廉价地挖掘旋毛虫高度近交种群的基因组，以支持暴发调查。整个程序的简单性以及 Trich-tracker 的时间和成本效益支持了它在正在进行的旋毛虫暴发中的实际应用。该工具的区分能力是可调的和可扩展的，允许在各种流行病学背景下应用，并易于适应其他寄生虫系统。
