利用PCR-DGGE法分析暗纹东方鲀的弧菌菌落组成

Analysis on Vibrio bacterial flora in Takifugu obscurus by PCR-DGGE

采用16S rDNA特征序列PCR-DGGE法,分析了不同饵料饲养的暗纹东方的表皮、性腺和肠道中,以弧菌为主的细菌群落的组成。结果表明:(1)共鉴定出暗纹东方体内18种微生物,均归属于变形细菌的gamma亚群,其中13种为弧菌科细菌,3种为肠杆菌科细菌,2种为气单胞菌属细菌;(2)投喂鲜活鱼虾的与投喂人工配合饲料的暗纹东方肠道和性腺中的细菌组成是不同的,在性腺中分离到的所有12种细菌中,只有2种细菌是相同的,约占17%;在肠道中分离到的所有13种细菌中,有4种是相同的,约占31%。而在表皮上所分离到的11种细菌中,有7种细菌是相同的,约占64%;(3)DGGE特征条带V1、V13、V18、V14和V17所代表的细菌只在表皮组织被分离到,V6所代表的细菌仅出现在以鲜活鱼虾作为饵料的暗纹东方性腺中,V5所代表的细菌仅出现在以鲜活鱼虾作为饵料的暗纹东方肠道中;(4)DGGE特征条带V5、V8、V13、V18、V6、V14和V17所代表的细菌不能在TCBS培养基上培养,占18种被鉴定的细菌总数的39%。

The bacterial flora in different tissues of puffer fish Fugu obscurus, which were fed with two different of kinds of food, were under research using PCR-DGGE technology. Total 18 species were identified as γ- Proteobacteria, including 13 kinds of Vibrionaceae, three kinds of Enterobacteriac and two kinds of Aeromonas. There existed difference in bacteria flora of intestine and gonad between two groups of puffer fish, which were fed with two different kinds of food, one （N group） consisted of fresh fish and shrimp, another （C group） is artificial fodder. In gonads, only two same kinds of bacteria , 17 percent of total 12 species, were found in the two groups. Four kinds of bacteria, 31 percent of total 11 species in intestine, were determined as same distribution between two groups of fish. However, the members of bacteria flora on different skins were most same, sharing same as 64 percent of total 11 species. Specific bacteria represented by specific DGGE bands V1,V13, V18,V14 and V17, only appeared on the fish skin. The bacteria of V6 band only emerged in the fish gonad tissue of N group, which were fed with fresh fish and shrimp. Likewise, only in the fish intestine of N group, the bacteria of V5 band were found. 39 percent of total identified bacteria in this research could not be cultured on TCBS plate.