昆虫病原线虫共生细菌的分子鉴定及其培养特性的初步研究  

Molecular identification and cultivation characteristics of entomopathogenic bacteria

作　　者：方香玲[1,2] 郭琪[2] 易晓华[2] 王永宏[2] 张兴[2] 

机构地区：[1]西北农林科技大学生命科学学院 [2]西北农林科技大学无公害农药研究服务中心,陕西杨凌712100

出　　处：《西北农林科技大学学报（自然科学版）》2008年第12期199-204,211,共7页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家"863"计划项目(2003AA241140);西北农林科技大学青年学术骨干支持项目(52211241)

摘　　要：【目的】确定昆虫病原线虫共生细菌YL001和YL002菌株的分类及其系统进化地位，并为该类生物资源的后续研究及开发应用奠定基础。【方法】对YL001和YL002菌株进行分子鉴定，PCR扩增并克隆其16srRNA全序列，对其全序列进行了序列测定及系统进化分析，并采用参数测定法对其培养特性进行了初步研究。【结果】YL001和YL002菌株与嗜线虫致病杆菌（Xenorhabdus nematophila）种内菌株形成一个类群，序列同源性大于99％，与发光杆菌（Photorhabdus）属内菌株的序列同源性大于94％。2菌株的延缓期、对数生长期、稳定期和衰亡期分别为0～6，6～18，18～66和66h；培养12h后，YL001和YL002菌株发酵液的pH值分别降低至5．70和5．56，此后逐渐上升，至发酵结束时其pH值分别为7．74和8．07t培养0～18h时2菌株发酵液中还原糖含量迅速降低，此后保持稳定；12h时氨基氮含量达到最低，此后开始缓慢上升；YL001菌株培养54h后及YL002菌株培养42和66h后，其对番茄灰霉病菌和枯草芽孢杆菌的抑制作用最强。【结论】YL001和YL002菌株属于嗜线虫致病杆菌种内的菌株；YL001和YL002菌株的培养特性符合细菌生长和代谢的一般规律，其适宜的发酵周期分别为54和66h。[Objective] The research determined the classification and systemic evolution status of ento mopathogenic bacteria YL001 and YL002,and laid the foundation for further research, development and application of such biological resource. [Method] Molecular identification was done for YL001 and YL002 strain,the complete 16S rRNA sequence was amplified and cloned; Sequence and phylogenetical analysis were studied with parameters determination. [Result] Strairis YL001 and YL002 formed a monophyletic clade with strains of X. nematophilus with sequence homology outweighing 99% and the sequence homology to genus Photorhabdus outweighing 94%. The lag, logarithmic, stationary and contabescence phase of the two bacteria were 0--6,6--18,18--66 and 66 h respectively;pH of strains YL001 and YL002 reduced to 5.70 and 5.56 at 12 h,respectively,and then rose gradually to 7.74 and 8.07 respectively when culture finished. Glucose content reduced quickly during 0--18 h, and then kept stable. Amido nitrogen content reached the lowest at 12 h and then rose slowly. YL001 exhibited the highest inhibitory effects at 54 h and YL002 exhibited highest inhibitory effects at 42 and 66 h respectively on B. cirerea and B. subtilis. [Conclusion] Strains YL001 and YL002 belonged to X. nematophilus. Cultivation characteristics of strains YL001 and YL002 were in accordance with the general disciplines of bacteria growth and metabolism,and the optimum fermentation period was 54 and 66 h respectively.

关 键 词：昆虫病原线虫 共生细菌 16S rRNA全序列 系统发育树 培养特性 

分 类 号：S476.15[农业科学—农业昆虫与害虫防治]