4种大型水母类ITS-5.8S rDNA序列分析及其在钵水母类系统分析中的应用

ITS-5.8S rDNA sequences of four Scyphomedusae jellyfish species and their applicability as a phylogenetic analysis marker

由于钵水母类生物地理学研究的缺乏以及不同时期形态变异较大等原因,对其分类鉴定比较混乱和困难。为弥补形态学分类的缺陷,采用通用引物PCR扩增法,测定了分布于黄海北部和辽东湾海域海蜇（Rhopilema esculentum）、沙蜇（Nemopilema nomurai）、海月水母（Aurelia sp.）、白色霞水母（Cyanea nozakii）4种大型水母的ITS-5.8S rDNA序列,同时利用Gen Bank数据库中已有的钵水母纲（Scyphomedusae）ITS1（the Ribosomal First Internal Transcribed Spacer）同源序列对其进行序列分析并构建系统树,分析ITS1序列片段在大型水母种类鉴定方面的可行性及其在钵水母类系统及演化中的应用。结果显示,4种水母的ITS-5.8S rDNA序列变异较大且具有明显的序列长度多态性,序列长度范围675~833 bp。钵水母纲很多种类的ITS1序列具有种内长度多态性现象,这种长度多态性主要是由于微卫星重复次数不同所造成的。钵水母纲科间遗传距离为0.295~0.491,种间遗传距离为0.024~0.812;除白色霞水母和海蜇外,种内个体间遗传距离为0.000~0.099。采用ML法（maximum likelihood）和贝叶斯法（Bayesian）构建的分子系统树拓扑结构不完全相同且与形态分类学的观点不太一致。研究表明,ITS基因序列在钵水母纲不同阶元间变异较大,适合于钵水母纲种类鉴定和属内种间水平的系统进化研究。

The class Scyphomedusae currently comprises 5 orders, 19 families, and about 202 species. These jellyfish are important pelagic predators at the center of concern about the ＂rise in gelatinous zooplankton＂ and degradation of coastal ecosystems. Previous phylogenetic relationship studies among Scyphomedusae species relied primarily on morphological data. However, morphology-based phylogeny and classification of this group are difficult due to the high diversity of morphological characters, and the topic remains controversial. Large molecular level advancements have yielded interesting results among the Scyphomedusae taxa. Nemopilema nomurai, Rhopilema esculentum, Cyanea nozakii, and Aurelia sp. are four common macro-jellyfish species in the Bohai and Yellow seas. In this study, the nuclear ITS-5.8S rDNA gene fragments from polyps, ephyrae, and adults of these species were amplified using polymerase chain reaction techniques. Homologous sequences of other Scyphomedusae jellyfish from Gen Bank were also included in this study. The genetic information indices, including base composition, sequence variation, and Kimura two-parameter genetic distance were examined. Phylogenetic trees were constructed based on the maximum-likelihood and Bayesian inference methods to determine the phylogenetic relationships among Scyphomedusae jellyfish. Significant length variations in the ITS-5.8S rDNA region were observed（length, 675–833 bp）. A total of 886 aligned base pairs, with 507 variable sites and 197 parsimony informative sites, were detected in the four jellyfish species. Inter-individual internal transcribed spacer region（ITS）1 polymorphisms were observed in most of the Scyphomedusae jellyfish species（four families and nine species）. Repeats in microsatellite ITS loci play an important role in length heteroplasmy. The genetic distances among families, species, and individuals were 0.295–0.491, 0.024–0.812, and 0–0.099, respectively. The phylogenic analysis showed that the topologies of the maximum-likelihood and Bayesian inference trees were somewhat different and different from morphological taxonomy. In conclusion, ITS is an appropriate barcoding tool to identify jellyfish species and classify young jellyfish. Furthermore, ITS1 was highly divergent among different Scyphomedusae jellyfish and could be an appropriate marker for molecular systematic studies at the species level. Additional in-depth studies with more samples are needed to further clarify the position of Scyphomedusae jellyfish in jellyfish taxonomy.