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中国农学通报

中国农学通报 ›› 2012, Vol. 28 ›› Issue (24): 183-187.

• 生物技术科学 • 上一篇    下一篇

香蕉内生拮抗细菌KKWB-10的分离鉴定及其内生性证明

王宇光 顾文亮 卢雪花 张欣 夏启玉   

  • 收稿日期:2011-11-21 修回日期:2012-02-22 出版日期:2012-08-25 发布日期:2012-08-25

Isolation and Identification of An Endophytic Bacterium KKWB-10 Against Banana Fusarium Wilt and Its Verification of Endophytic Activity

  • Received:2011-11-21 Revised:2012-02-22 Online:2012-08-25 Published:2012-08-25

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摘要：

从香蕉体内分离拮抗香蕉枯萎病菌的内生细菌，旨在应用于香蕉枯萎病的生物防治。从健康香蕉植株的球茎分离到多株细菌，对其进行了拮抗香蕉枯萎病菌4号小种的实验，选择其中1株拮抗性较强的菌株KKWB-10进行了形态特征、生理生化测定、16S rDNA序列分析，并通过浸根法将标记了绿色荧光蛋白GFP的该菌（K-pUCK7-1′GT菌）回接无菌的香蕉组培苗，培养10天后，制备香蕉苗的根和茎的手工切片于激光共聚焦显微镜下观察。抑菌实验结果表明，该菌株对香蕉枯萎病菌4号生理小种有较强的抑制作用。通过形态学观察、生理生化指标测定、16S rDNA序列同源性分析，将该菌株初步鉴定为阴沟肠杆菌(Enterobacter cloacae)。激光共聚焦显微镜观察发现经K-pUCK7-1′GT菌液处理的香蕉组培苗的根和茎的切片中均发现绿色荧光，而未处理的对照组的香蕉苗却没有发现绿色的荧光，表明该菌株能定殖于香蕉的根和茎内。该菌株可直接作为生防菌剂，也可用作外源抗病基因在香蕉植株体内表达的载体，在香蕉枯萎病菌的生物防治中具有一定的潜在价值。

关键词: CMV, CMV

Abstract:

The endophytic bacteria against banana fusarium wilt from banana were isolated in order to study the biocontrol of banana fusarium wilt. Bacteria were isolated from the corm of healthy banana, and their antagonistic abilities to Fusarium oxysporum f.sp cubense race 4 (Foc 4) were respectively determined. KKWB-10 strain exhibiting excellent resistance against Foc 4 was identified by morphological, physiological and 16S rDNA gene sequence. A GFP-labeling KKWB-10 strain K-pUCK7-1′GT was inoculated into banana tissue culture plantlets using the root dipping method. After 10 days of growth, freehand sections of the stems and roots of these banana plantlets were observed by laser confocal scanning microscope. The results of antagonistic experiments showed that KKWB-10 strain was an antagonistic bacterium that exhibited excellent resistance against Foc 4. The strain was primarily identified as Enterobacter cloacae based on the results of its 16S rDNA gene sequence homology, the physiological and morphological characteristics. Green fluorescence was observed in the roots and stems of banana plantlets inoculated with GFP-labeling KKWB-10 strain, but not in uninoculated controls. The result suggested that the KKWB-10 strain could colonize the roots and stems of banana plants. The KKWB-10 strain can be used as not only biocontrol agents but also vectors for the introduction of heterologous resistance genes into banana plants. Thus the KKWB-10 strain would be a potential natural resource for the biological control of banana fusarium wilt.