Tatonova Yulia V, Chelomina Galina N, Besprosvannykh Vladimir V
Institute of Biology and Soil Science, Far Eastern Branch, Russian Academy of Sciences, 100-letiya Street, 159, Vladivostok 690022, Russia.
Parasitol Int. 2012 Dec;61(4):664-74. doi: 10.1016/j.parint.2012.07.005. Epub 2012 Jul 21.
The present study examined the molecular organisation and sequence variation in the nuclear ribosomal DNA (rDNA) region, including the two internal transcribed spacers (ITS1 and ITS2) and the 5.8S gene of the Clonorchis sinensis from the Russian Far East. The relevant sequences from other parts of this species' area were downloaded from GenBank. The results showed 100% identity for all investigated 5.8S-ITS2 rDNA sequences. In contrast, two levels of intraspecific variations were revealed in the complete ITS1 sequences. The intra-genomic variation resulted from a C/T polymorphism in a single position. The inter-individual differences between the ITS1 sequences were both due to nucleotide and size polymorphisms resulting from a varying number of five-nucleotide repeats and followed by two ITS1 length variants. These variant frequencies correlate with the clonorchiasis level in some geographical localities. ITS1 differences, both in the mutation profile and mutation localisation, were revealed between northern and southern geographical samples. The presence of GC boxes that are identical to known regulatory motifs in eukaryotes was detected within the ITS1 sub-repeats. The predicted secondary structures for ITS1 consist of two large branches, one of which was invariable, while another depended on ITS1 length. The predicted secondary structure for ITS2 includes four helices around the core. The main differences between C. sinensis and other opisthorchids were localised on the tops of helices 2, 3, and 4. A phylogenetic MST reconstruction subdivided all ITS1 sequences into two well differentiated clusters, each with the major widespread ribotype, and showed that ribotype diversity in both Russia and Korea is much lower than in China. The results obtained demonstrate the feasibility of complete ITS1 sequences in C. sinensis population genetics and can be considered as a basis for further studies of the parasite infection because they may help to elucidate the molecular mechanisms of pathogen evolution and adaptation.
本研究检测了来自俄罗斯远东地区华支睾吸虫核糖体DNA（rDNA）区域的分子组织和序列变异，包括两个内部转录间隔区（ITS1和ITS2）以及5.8S基因。从GenBank下载了该物种分布区其他地区的相关序列。结果显示，所有检测的5.8S-ITS2 rDNA序列具有100%的同一性。相比之下，完整的ITS1序列显示出两种水平的种内变异。基因组内变异源于单个位置的C/T多态性。ITS1序列之间的个体差异既源于核苷酸多态性，也源于由不同数量的五核苷酸重复序列导致的大小多态性，并产生了两种ITS1长度变体。这些变体频率与某些地理区域的华支睾吸虫病水平相关。在北方和南方地理样本之间，ITS1在突变谱和突变定位方面均存在差异。在ITS1亚重复序列中检测到与真核生物中已知调控基序相同的GC框。预测的ITS1二级结构由两个大分支组成，其中一个是不变的，而另一个则取决于ITS1长度。预测的ITS2二级结构包括围绕核心的四个螺旋。华支睾吸虫与其他后睾吸虫之间的主要差异位于螺旋2、3和4的顶部。系统发育MST重建将所有ITS1序列细分为两个高度分化的簇，每个簇都有主要广泛分布的核糖型，并表明俄罗斯和韩国的核糖型多样性远低于中国。所得结果证明了完整ITS1序列在华支睾吸虫群体遗传学中的可行性，可作为进一步研究寄生虫感染的基础，因为它们可能有助于阐明病原体进化和适应的分子机制。
