大量鞭毛虫栖息在低等白蚁肠道内,是白蚁赖以生存的共生微生物。不同种类的鞭毛虫共同作用形成了一套降解食物的系统,为宿主提供营养和能量。研究鞭毛虫群落结构是揭示其各组成种类生理功能的基础。利用形态特征进行物种鉴定受鞭毛虫生长发育阶段、样品制备方法等多种因素的影响,而基于分子标记的分子生物学方法能不受这些因素的制约来研究复杂的微生物群落。本研究结合形态特征鉴定和分子生物学方法研究台湾乳白蚁Coptotermes formosanus肠道鞭毛虫群落结构,并对这些方法进行了比较。通过光学显微镜和扫描电子显微镜进行形态观察鉴定,确定了台湾乳白蚁肠道内的3种鞭毛虫,分别为伪披发虫Pseudotrichonympha grassii、全鞭毛虫Holomastigotoides mirabile和旋披发虫Spirotrichonympha leidyi。18S rDNA文库限制性片段长度多态性分析较形态鉴定能够反映群落更复杂的物种多样性。利用光学显微镜进行细胞计数较18S rDNA文库克隆数能更准确地反映各种鞭毛虫数量,每头工蚁肠道内平均含伪披发虫780±179头,全鞭毛虫1630±391头,旋披发虫2950±1003头。本研究建立了光学显微镜形态鉴定和18S rDNA分子标记相结合调查鞭毛虫多样性和数量的方法,为进一步研究白蚁肠道共生生物的功能奠定了基础。
A large number of flagellates live in the intestine of lower termites, which is vital for the survival of host termites. Different species of flagellates form a food digestion system to provide nutrients and energy for the host. Studies on the community structure of flagellates will provide the basis for elucidating the physiological functions of each member. Species identification based on morphological characteristics is influenced by the flagellate growth and developmental stages, sample preparation methods and other factors, but molecular biological methods based on molecular markers can avoid these drawbacks in studying complex microbial communities. In this study, the community structure of flagellates in the gut of Coptotermes formosanus was investigated with molecular biological methods combining with morphological characteristics. Three methods were also compared in studying community structure. Three species of flagellates (Pseudotrichonympha grassii, Holomastigotoides mirabile, and Spirotrichonympha leidyi) were identified by optical and scanning electron microscopic observation. Compared with morphological identification, 18S rDNA restriction fragment length polymorphism analysis can reveal more species diversity. Cell counting under optical microscopy is more accurate than 18S rDNA analysis in investigating the amount of each flagellate. The average number of flagellates per termite worker was 780±179 for P. grassii, 1 630±391 for H. mirabile, and 2 950±1 003 for Spirotrichonympha leidyi. This study established a method of combining optical microscopy observation with 18S rDNA analysis to investigate intestinal flagellate community, which is the foundation to clarify the biological functions of the gut symbionts.