棉花立枯病拮抗细菌的分离鉴定及抑菌活性

doi:10.13304/j.nykjdb.2022.0628

摘要/Abstract

摘要：

为获取对棉花立枯病具有生防作用的细菌，从棉田土壤中筛选出1株高效抑制立枯丝核菌的拮抗细菌W1，通过形态学特征结合16S rDNA序列分析鉴定其分类地位，并采用平板对峙法分析其抑菌活性。结果表明，拮抗细菌W1经鉴定为枯草芽孢杆菌（Bacillus subtilis），其对立枯丝核菌具有稳定的抑菌作用，发酵原液的抑菌率为82.12%，稀释5倍的无菌发酵液和菌体破碎液抑菌率分别为42.54%和46.15%。抑菌活性受培养基成分影响，BPA培养基的发酵液抑菌率最高，达98.34%，NA培养基抑菌率仅为17.12%。抑菌活性物质在80 ℃、中性和偏碱性条件下具有稳定性，对蛋白酶K处理不敏感。显微观察发现，菌株W1的主要拮抗机理是通过产生拮抗物质造成病原菌的菌丝消融、菌丝体断裂、扭曲和畸形。结果表明拮抗细菌W1是具有潜在应用前景的防治棉花立枯病及其他植物真菌病害的菌株资源。

关键词: 棉花立枯病, 分离鉴定, 枯草芽孢杆菌, 抑菌活性

Abstract:

In order to obtain the antagonistic bacteria against cotton damping-off， the bacteria W1 that effectively antagonize Rhizoctonia solani was isolated from cotton field soil. W1 was identified based on morphological characteristics and the phylogenetic analysis of partial 16S rDNA sequence， and its antifungal activity was analyzed by plate confrontation method. The results showed that strain W1 was identified as Bacillus subtilis. The inhibitory effect of fermentation liquid against Rhizoctonia solani was 82.12%， and the inhibition rate of sterile culture filtrates and cell fragmentation liquid diluted by 5-fold was 42.54% and 46.15%， respectively. The antagonistic activity were affected by the composition of culture medium， the inhibition rate of fermentation filtrate from BPA medium was highest with 98.34%， and that of NA medium was only 17.12%. The antagonistic substances were stable under 80 ℃， neutral and alkaline environment， and insensitive to proteinase K. The strain W1 resulted in mycelium ablation， mycelium fracture， distortion and malformation， which hindered the growth of Rhizoctonia solani. This study indicated that the antagonistic strain W1 was a potential strain resource for effective control of cotton damping-off and other plant fungal diseases.

Key words: Rhizoctonia solani, isolation and identification, Bacillus subtilis, antifungal activity

中图分类号:

S562

牛营超, 王星, 郭青云, 戴小华, 袁小勇, 陈琳. 棉花立枯病拮抗细菌的分离鉴定及抑菌活性[J]. 中国农业科技导报, 2023, 25(12): 138-144.

Yingchao NIU, Xing WANG, Qingyun GUO, Xiaohua DAI, Xiaoyong YUAN, Lin CHEN. Identification and Antifungal Activity of a Bacteria Strain Against Rhizoctonia solani[J]. Journal of Agricultural Science and Technology, 2023, 25(12): 138-144.

图/表 6

图1 菌株W1形态特征A：菌株W1菌落形态； B：菌落W1细胞形态

Fig. 1 Morphological characteristics of strain W1A： Colonial morphology of strain W1； B： Cell morphology of strain W1

图2 基于16S rDNA序列构建的菌株W1系统发育树

Fig. 2 Phylogenetic tree of strain W1 based on 16S rDNA sequence

图3 菌株W1对棉花立枯病菌生长的抑制作用A: W1 发酵液对棉花立枯病菌生长的抑制作用；B: W1无菌发酵液和菌体破碎液对棉花立枯病菌生长的抑制作用

Fig. 3 Inhibitory effect of strain W1 on the growth of R. solaniA；Inhibitory effect of W1 fermentation broth on the growth of R. solani； B: Inhibitory effect of W1 sterile culture filtrates and cell fragmentation liquid on the growth of R. solani

图4 不同培养基处理下W1无菌发酵液的抑菌率

Fig. 4 Inhibition rate of cell-free fermentation broth of strain W1 under different mediums

图5 不同条件下菌株W1无菌发酵液的稳定性

Fig. 5 Stability of antagonistic substances of strain W1 under different conditions

图6 菌株W1对棉花立枯菌丝生长的影响A：棉花立枯病菌正常菌丝； B~D：棉花立枯病菌受抑制的菌丝

Fig. 6 Effects of strain W1 on mycelial morphology of R. solaniA： Normal hyphae of R. solani； B~D： Inhibited hyphae of R. solani

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