基于中国不同毒力的1型弓形虫虫株二代测序的变异检测
Variation detection based on next-generation sequencing of type Chinese 1 strains of Toxoplasma gondii with different virulence from China.
作者信息
Cheng Weisheng, Liu Fang, Li Man, Hu Xiaodong, Chen He, Pappoe Faustina, Luo Qingli, Wen Huiqin, Xing Tian, Xu Yuanhong, Shen Jilong
机构信息
Department of Clinical Laboratory, the First Affiliated Hospital of Anhui Medical University, Hefei, 230022, People's Republic of China.
Department of Microbiology and Parasitology, Anhui Provincial Laboratory of Pathogen Biology and Anhui Key Laboratory of Zoonoses, Anhui Medical University, Hefei, 230022, People's Republic of China.
出版信息
BMC Genomics. 2015 Oct 30;16:888. doi: 10.1186/s12864-015-2106-z.
BACKGROUND
Toxoplasma gondii is an intracellular protozoan that affects most species of endothermic animals including humans with a great infection rate. The vertical transmission of T. gondii causes abortion, constituting a serious threat to humans and leading to great losses in livestock production. Distinct from population structure of T. gondii in North America and Europe, Chinese 1 (ToxoDB #9) is a dominant genotype prevalent in China. Among the isolates of Chinese 1, the Wh3 and Wh6 have different virulence and pathogenicity in mice. However, little has been known about their difference at the genomic level. Thus the next-generation sequencing (NGS) approach was used to discover the association of the phenotypical variations with the genome sequencing data and the expression and polymorphisms of the key effectors.
RESULTS
We successfully sequenced the genome of Chinese 1 strains of Wh3 and Wh6. The average sequencing depths were 63.91 and 63.61 for Wh3 and Wh6, respectively. The variations of both isolates were identified in comparison with reference genome of type I GT1 strain. There were 505,645 and 505,856 SNPs, 30,658 and 30,004 indels, 4661 and 2320 SVs, and 1942 and 3080 CNVs for Wh3 and Wh6, respectively. In target search variations of particular factors of T. gondii, the dense granule protein 3 (GRA3) and rhoptry neck protein 3 (RON3) were found to have 35 SNPs, 2 indels and 89 SNPs, 6 indels, respectively. GRA3 and RON3 were both found to have higher expression levels in less virulent Wh6 than in virulent Wh3. Both strains of type Chinese 1 share polymorphic GRA15II and ROPI/III with type I, II, and III strains.
CONCLUSIONS
Sequencing of the two strains revealed that genome structure of Chinese 1 and type I strains has considerable genomic variations. Sequencing and qRT-PCR analyses of 26 effectors displayed a remarkable variation that may be associated with phenotype and pathogenic differences.
背景
刚地弓形虫是一种细胞内原生动物，可感染包括人类在内的大多数恒温动物，感染率很高。刚地弓形虫的垂直传播会导致流产，对人类构成严重威胁，并给畜牧业生产造成巨大损失。与北美和欧洲的刚地弓形虫种群结构不同，中国1型（ToxoDB #9）是在中国流行的优势基因型。在中国1型分离株中，Wh3和Wh6在小鼠体内具有不同的毒力和致病性。然而，它们在基因组水平上的差异鲜为人知。因此，采用下一代测序（NGS）方法来发现表型变异与基因组测序数据以及关键效应因子的表达和多态性之间的关联。
结果
我们成功对中国1型Wh3和Wh6菌株的基因组进行了测序。Wh3和Wh6的平均测序深度分别为63.91和63.61。与I型GT1菌株的参考基因组相比，鉴定了两种分离株的变异。Wh3和Wh6分别有505,645和505,856个单核苷酸多态性（SNP）、30,658和30,004个插入缺失（indel）、4661和2320个结构变异（SV）以及1942和3080个拷贝数变异（CNV）。在刚地弓形虫特定因子的靶向搜索变异中，致密颗粒蛋白3（GRA3）和棒状体颈部蛋白3（RON3）分别有35个SNP、2个indel和89个SNP、6个indel。发现GRA3和RON3在毒力较低的Wh6中的表达水平均高于毒力较强的Wh3。中国1型的两种菌株与I型、II型和III型菌株共享多态性GRA15II和ROPI/III。
结论
对这两种菌株的测序表明，中国1型和I型菌株的基因组结构存在相当大的基因组变异。对26个效应因子的测序和qRT-PCR分析显示出显著变异，这可能与表型和致病性差异有关。
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