草莓胶孢炭疽菌拮抗细菌贝莱斯芽孢杆菌JK3的鉴定及其抗菌活性

利用平板对峙法从堆肥中分离筛选到1株对草莓胶孢炭疽菌具有较强拮抗作用的细菌株系JK3。通过生理生化特征、16S rDNA和rpoB序列分析,将JK3菌株鉴定为贝莱斯芽孢杆菌。利用离体草莓叶片测定了JK3发酵上清液对草莓炭疽病的防治效果。结果显示:JK3发酵上清液对草莓胶孢炭疽菌引起的草莓炭疽病有明显预防效果,但治疗作用较差;提前24 h或接种胶孢炭疽菌孢子的同时喷施JK3发酵上清液,防治效果超过95%;接种胶孢炭疽菌孢子后24 h喷施发酵上清液,防治效果为11.5%。对JK3发酵上清液进行不同的温度、pH和蛋白酶处理,结果显示,发酵上清中的抑菌物质对pH和蛋白酶处理不敏感,但高温可降低其抑菌活性,推测主要抗菌活性成分不是蛋白质。由于芽孢杆菌多产生脂肽类抗菌物质,利用相关合成基因检测引物进行PCR扩增并测序,结果显示,JK3菌株含有脂肽类抗菌物质生物合成相关基因srfAA、bymB、fenD、bacA和ituC。以上结果表明,贝莱斯芽孢杆菌JK3有开发为防治胶孢炭疽菌引起的草莓炭疽病的生防制剂潜力。

Identification of antagonistic bacteria Bacillus velezensis JK3 against anthracnose of strawberry and its antipathogenic activity

A bacterial isolate JK3, which was isolated from compost, showed stronger antagonistic activity against Colletotrichum gloeosporioides causing strawberry anthracnose in dual culture assay. The isolate JK3 was identified as Bacillus velezensis by physiological and biochemical characterization, 16S rDNA and rpoB gene sequence analysis. The inhibitory effects of JK3 fermented liquid on C. gloeosporioides were evaluated with the detached strawberry leaves. JK3 fermented liquid showed significant protection on strawberry anthracnose but poor therapeutic effect. More than 95% control efficiency was achieved by spraying JK3 fermented liquid 24 h before inoculation with the spore suspension of C. gloeosporioides or simultaneous inoculation. Only 11.5% control efficiency was resulted by spraying JK3 fermented liquid 24 h after inoculation with the pathogen. The antagonistic activity of the fermented liquid was stable to different pH values and proteases, but sensitive to high temperature. Thus implied that active component was not related to proteins. It has been reported that most isolates of Bacillus spp. produce active lipopeptides. The existences of lipopeptide biosynthetic genes in JK3 were examined by PCR amplification with proper primer pairs and DNA sequencing. Results showed that the JK3 contained srfAA, bymB, fenD, bacA and ituC genes. The above data indicated that B. velezensis JK3 had a potential to be developed as biocontrol agent for controlling strawberry anthracnose caused by C. gloeosporioides.