海黄瓜共附生细菌多样性及其对甘蔗鞭黑粉菌的抑制作用

Diversity of symbiotic bacteria isolated from Pseudocnus echinatus and its antifungal activity against Sporisorium scitamineum

【目的】研究海黄瓜(Pseudocnus echinatus)共附生细菌多样性及其发酵产物对甘蔗鞭黑粉菌的抑制作用,为甘蔗黑穗病的防治及新型生物农药的研发提供新途径。【方法】利用微生物纯培养技术分离纯化海黄瓜的表皮、肠、胃和肠内容物等组织的共附生细菌,并通过16S r DNA序列分析共附生细菌种类多样性及其在海黄瓜不同部位组织的分布特征;采用牛津杯法测定共附生细菌发酵产物对甘蔗鞭黑粉菌的抑制作用。【结果】从海黄瓜各部位组织共分离得到79株共附生细菌,归属于4门26科35属46种;优势菌属为:芽孢杆菌属(Bacillus,16.5%)、鞘氨醇盒菌属(Sphingopyxis,8.9%)、小单孢菌属(Micromonospora,6.3%)和柠檬酸杆菌属(Citrobacter,6.3%)。分别以甘蔗鞭黑粉菌的"+"型担孢子、"-"型担孢子和菌丝为靶标,采用牛津杯法筛选得到19株活性菌,其中以芽孢杆菌属的BGMRC03005对甘蔗鞭黑粉菌菌丝、"+"型担孢子和"-"型担孢子的抑制作用最强,抑菌圈直径分别为25.40、19.90和14.25 mm。【结论】海黄瓜中可培养细菌种类丰富,且富含对甘蔗鞭黑粉菌有抑制作用的菌株。筛选获得的芽孢杆菌属细菌BGMRC03005对3种鞭黑粉菌形态均有抑制作用,具有开发成为微生物农药的潜力。

【Objective】The purpose of this study was to provide new approach for prevention and control of sugarcane smut and development of new biological pesticide by investigating the diversity of symbiotic bacteria isolated from Pseudocnus echinatus and testing the bacteriostatic activities of their fermentations against Sporisorium scitamineum.【Method】Pure culture technology was used to isolate and purify the symbiotic bacteria from various parts of P.echinatus including its epidermis,intestine,stomach and intestinal contents.Diversity of symbiotic bacteria and its distribution characteristics in different tissues of P.echinatus were analyzed through 16S rDNA.Inhibition effects of the fermentations from symbiotic bacteria against S.scitamineum were detected by Oxford cup method.【Result】Seventy-nine symbiotic bacteria belonging to 46 species,35 genera,26 families,4 phyla were isolated from various parts of P.echinatus,and the dominant genera were Bacillus(16.5%),Sphingopyxis(8.9%),Micromonospora(6.3%)and Citrobacter(6.3%).Taking“+”shaped,sporidum,“-”shaped sporidum and hypha of S.scitamineum as targets,19 live strains were screened using Oxford cup method.Among theses strains,the inhitition effects of Bacillus siamensis starin BGMRC03005 were the strongest against hypha(diameter of inhibition zone:25.40 mm),“+”shaped sporidum(diameter of inhibition zone:19.90 mm)and“-”shaped sporidum(diameter of inhibition zone:14.25 mm)of S.scitamineum.【Conclusion】The species of cultivated bacteria in P.echinatus are diversified and most of them have strong antifungal activities against S.scitamineum.The selected B.siamensis strain BGMRC03005 has inhibition effects on the three forms of S.scitamineum,which could be developed into microbial pesticide.