Abstract:

Objective To isolate Gordonia from the gut of Periplaneta americana, and to carry on classification, identification and phylogenetic analysis on the Gordonia. Methods Taking the gut was acquired from captured P. americana by opening the abnormal with scalpel from the side after surface disinfection. After grinding, the homogenate was be diluted to the required concentration (10, 0.005, 0.0025 μg/ml). An aliquot of 0.1 ml was drawn on the tablet, and cultured at 28℃ for at least one week, to isolate the Gordonia. Finally, morphological observation and 16S rDNA gene sequences analysis were utilized to study their classification and identification. The sequence data were analyzed using Mega 5.1 software, maximum likelihood method, and phylogenetic tree. Results Six Gordonia were isolated and identified from the gut of wild P. americana. The gram stain was positive, the colonies were small, orange red, granular, the slightly wetsurface, with production of orange red pigment. The thalli was short rod. The mycelium was straight rod or spiral, without transverse septum, consistent with the basic characteristics of Gordon's bacteria. After the amplification of 16S rDNA gene, 1% agarose gel electrophoresis, six samples all showed target bands (1.5 kb), the sequencing results showed that the isolates were Gordonia. Phylogenetic tree analysis showed that the isolates obtained were close to the Gordon's genetic relationship in soil, plant and marine. Conclusion There is existing Gordonia in the gut of P. americana. The Gordonia isolated from soil, plants and marine have the function of degrading the biological macromolecules and absorbing heavy metals, which can lay foundation for further study of symbiont flora in P. americana.

Key words: Periplaneta americana, Gordonia, 16S rDNA, Phylogenetic

摘要：

目的 从美洲大蠊肠道内分离戈登菌,对其进行分类鉴定和系统发育学分析。方法 室外捕获美洲大蠊样品,体表消毒后,用手术刀从侧面将其腹部切开,取出肠道,匀浆至所需浓度(10、0.005、0.0025μg/ml),吸取悬浮液0.1 ml于平板上,28℃培养至少1周,分离美洲大蠊肠道内的戈登菌。通过分离菌株形态观察和16S rDNA序列分析分类鉴定,运用Mega 5.1软件,利用最大似然值法,构建系统发育树。结果 从美洲大蠊肠道分离出戈登菌6株。菌株革兰染色呈阳性,菌落较小,呈橘红色,颗粒状,表面稍微湿润,产橘红色色素。菌体呈短棒状,菌丝呈直杆状或螺旋状,无横隔,符合戈登菌的基本特征。16S rDNA基因扩增后,测序结果表明,所分离菌株均为戈登菌。系统发育树分析表明,所分离菌株与分离自土壤、植物和海洋中的戈登菌亲缘关系较近。结论 首次从美洲大蠊肠道内分离到戈登菌,为进一步研究美洲大蠊内生戈登菌的作用奠定基础。

关键词: 美洲大蠊, 戈登菌, 16S rDNA, 系统发育学