The Phylogeny Of Six Species Sea Cucumbers In South China Sea And Studies On Embryonic And Larval Development Of Sea Cucumber,Holothuria Scabra And Stichopus Variegates

Sea cucumber belong to phylum Echinodermata, class Holothuroidea and is a very important group of economic animal. Since ancient times, sea cucumber is one kinds of very slap-up nutrient and medicinal food. With the development of science and technology, there has been a great deal of researches showing that many kinds of ingredient in sea cucumber have a good effect in preventing and curing different diseases, having a huge potential for development of marine drug. Because the sea cucumber is hard to be identified with morphological characteristic and the genetic relationship of sea cucumber could not be affirmed by morphological taxonomy, the Genbank of DNA code should be perfected and the taxonomic status of sea cucumber should be explored by molecular systematology. At the same time, there are not many researches in edible sea cucumbers in the South China Sea, so it is necessary to explore and cultivate these kinds of sea cucumbers to meet market demand.The16S rDNA and the18S rDNA of sea cucumber are good molecular markers of phylogeny. The rate of evolution of16S rDNA is faster, so it is applied to classification on level of genus even species. But the rate of evolution of18S rDNA is slower, so it is applied to classification on level of family or higher taxonomic category. The technologies of PCR and gel electrophoresis were used in this research. The DNA samples of Holothuria scabra, Stichopus variegates, H. leucospilota, H. aff impatiens, H. cinerascens and H. fuscocinerea were extracted and sequenced. And the16S rDNA and18S rDNA sequences of some species of Echinodermata were downloaded from Genbank to be compared as external reference samples. And then, the software of ClustalX and Mega4.1were used to structure phylogenetic trees. Conclusions were as follows:1> The complete length of16S rDNA was576bp, and the average content of composition T, C, A, G within16S rDNA of13species sea cucumbers were25.1%,23.7%,29.6%,21.7%respectively, and the transition/transversion(R) and average genetic distance were1.468and0.196respectively. The complete length of18S rDNA was1413bp, and the average content of composition T, C, A, G within18S rDNA of14species of Echinodermata were22.3%.24.6%,23.9%,29.2%respectively, and the transition/transversion(R) and average genetic distance were1.585and0.082respectively. The R of two DNA were low, so the mutagenesis of16S rDNA and18S rDNA were saturated. At the same time, the contents of C+G were all higher than that of A+G in two DNA.2、The phylogenetic tree of16S rDNA was constituted by two big branches:One included species of Holothuria, Holothuriidae, another one included species of Stichopus, Stichopodidae; and it was accord with traditional taxonomy. The phylogenetic tree of18S rDNA was constituted by two big branches too:one included species of Holothuriidae, Stichopodidae and Thyonella gemmata of Dendrochirota, another one included species of Metacrinus, sea chestnuts and sea cucumbers of Apoda. But it was not accord with traditional taxonomy.At the same time, the larva of H. scabra and S. variegates were observed in this research. The bloodstocks were induced spawning artificially by using the method of streaming and drying in the shade jointly. Until the embryos developed into late gastrula stage, the Chlorella sp., Isochrysis galbana and Chaetoceros muelleri were took into pool as bait. When30%of larva developed into doliolaria stages, the plastic films adhered with diatom were threw into pool as adherences. Larva was observed and measured with microscope and were photographed with camera. Some results as follows:1、There were "caudateprocess" within the bodies of H. atra and H. scabra of Holothuriidae, and the ossicle was in the tail end of "caudateprocess". But S. variegates and A. japonicus of Stichopodidae had no this organ and the ossicle were in the cusp of posterolateral arms. Thus, the larval forms of Holothuriidae and Stichopodidae may be different.2., The developmental times were vary in different sea cucumbers. The larval development of H. scabra was faster than that of S. variegates under the same temperature. Through comparison, spacing interval of cleavage of S. variegates and A. japonicus was about40min, and20min approximately was within H. scabra.3、Stomach ulcer disease not only happened in auricularia of A japonicus but also in H. scabra and S. variegates, because of quality of bait was not good. But the disease could be controlled by changing amount of water and feeding bait with good quality.4、Larval arms and the number of hyaline spheres were important marks of larval health. The number of hyaline spheres is5pairs. The more hyaline spheres developed and the stronger the larval arm was, the healther larva was.