Screening and Identification of a Strain of Bacillus velezensis Against Botrytis cinerea

doi:10.13304/j.nykjdb.2024.0386

Abstract

Abstract:

In order to isolate a biocontrol bacteria with antagonistic effects on Botrytis cinerea，the antagonistic bacteria were screened from grape rhizosphere soils by plate confrontation method and liter paper method. A strain with strong antagonistic effect was selected and identified by morpholoigical， physiological and biochemical characteristics， 16S rDNA sequences analysis. The safety， growth-promoting and the control effect of the strain were tested by the blood plate method， seed germination experiments and detached fruit method. The results showed that 5 antagonistic bacteria were screened from 142 bacterial strains isolated from grape rhizosphere soils. These 5 antagonistic bacteria had different inhibitory effect on grape gray mold disease with the inhibition rates from 81.74% to 89.13%， and the inhibition zone were from 2.42 to 11.08 mm， among which the inhibitory effect of strain B1-4 was the highest. By morph-oloigical， physiological and biochemical characteristics， 16S rDNA sequences， strain B1-4 was identified as Bacillus velezensis，which had no hemolysis， could produce cellulase， protease and siderophores， and had nitrogen fixation effect. The germination rate and radicle length of cucumber and rapeseed seeds could be significantly improved by 100 times dilution of B1-4 fermentation solution. The protective effect of B1-4 fermentation liquid reached 70.31% and the rapeutic effect was 69.53% on postharvest grape fruit gray mold， which was no significant difference with 5 000 times dilution of 50% fludioxonil （71.88%）. Above results showed that B. velezensis B1-4 was a protential biocontrol strain for control B.cinerea.

Key words: Bacillus velezensis, Botrytis cinerea, isolation and identification, growth promotion, antagonistic effect

摘要：

为筛选对葡萄灰霉病菌具有拮抗作用的生防细菌，采用平板对峙法和滤纸片法从葡萄根际土壤中分离具有拮抗作用的细菌，选择拮抗作用较强菌株，通过形态特征、生理生化特性及16S rDNA序列分析对其进行鉴定，并通过血平板法、种子萌发试验和离体果实法对其安全性、促生性和防治效果进行检测。结果显示，从葡萄根际土壤中分离的142株细菌中筛选出5株具有拮抗作用的细菌，这5株拮抗细菌对葡萄灰霉病菌具有不同程度的抑制作用，抑菌率为81.74%~89.13%，抑菌带宽为2.42~11.08 mm，其中菌株B1-4对葡萄灰霉病菌的拮抗作用最强；经形态特征、生理生化特性及16S rDNA序列分析鉴定，B1-4为贝莱斯芽孢杆菌（Bacillus velezensis）。该菌株不具备溶血性，能够分泌纤维素酶和蛋白酶，并具有固氮与产铁载体能力；其发酵液的100倍稀释液可显著提高黄瓜、小油菜种子的萌发率和胚根长度。B1-4发酵原液对采摘后葡萄灰霉病的保护作用防效为70.31%，治疗作用防效为69.53%，与50%咯菌腈5 000倍稀释液防效（71.88%）差异不显著。因此，贝莱斯芽孢杆菌B1-4是一株具有防控葡萄灰霉病潜力的生防菌株。

关键词: 贝莱斯芽孢杆菌, 葡萄灰霉病菌, 分离鉴定, 促生作用, 拮抗作用

CLC Number:

S182

Haiyan LI, Ting ZHANG, Xinchang LI, Ling ZHANG, Pei WANG, Minhu BAO. Screening and Identification of a Strain of Bacillus velezensis Against Botrytis cinerea[J]. Journal of Agricultural Science and Technology, 2025, 27(8): 110-118.

李海燕, 张婷, 李新畅, 张玲, 王培, 鲍民胡. 一株拮抗葡萄灰霉病菌的贝莱斯芽孢杆菌筛选及鉴定[J]. 中国农业科技导报, 2025, 27(8): 110-118.

Figures/Tables 9

References 33

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分级Grade	分级标准 Grading standard
0级Level 0	无病斑 No spots
1级Level 1	病斑面积占5%以下 Lesion area<5%
3级Level 3	病斑面积占6%~15% 6%< lesion area<15%
5级Level 5	病斑面积占16%~30% 16%< lesion area<30%
7级Level 7	病斑面积占31%~50% 31%< lesion area<50%
9级Level 9	病斑面积占50%以上 Lesion area>50%

分级Grade	分级标准 Grading standard
0级Level 0	无病斑 No spots
1级Level 1	病斑面积占5%以下 Lesion area<5%
3级Level 3	病斑面积占6%~15% 6%< lesion area<15%
5级Level 5	病斑面积占16%~30% 16%< lesion area<30%
7级Level 7	病斑面积占31%~50% 31%< lesion area<50%
9级Level 9	病斑面积占50%以上 Lesion area>50%

菌株Strain	抑菌率Inhibition rate/%	抑菌带Inhibitory zone/mm
B1-4	89.13±5.43 a	11.08±0.90 a
0-1	88.04±2.94 a	8.33±2.39 b
0-0	84.13±0.38 b	3.58±1.78 c
1-4-1	81.96±0.75 b	2.42±1.38 c
C₁	81.74±0.65 b	8.25±0.97 b

菌株Strain	抑菌率Inhibition rate/%	抑菌带Inhibitory zone/mm
B1-4	89.13±5.43 a	11.08±0.90 a
0-1	88.04±2.94 a	8.33±2.39 b
0-0	84.13±0.38 b	3.58±1.78 c
1-4-1	81.96±0.75 b	2.42±1.38 c
C₁	81.74±0.65 b	8.25±0.97 b

处理 Treatment	黄瓜Cucumber		小油菜Rape
处理 Treatment	萌发率 Germination rate/%	胚根长度 Root length/cm	萌发率 Germination rate/%	胚根长度 Root length/cm
CK1	92.67±3.01 b	4.60±0.16 b	86.00±2.00 b	3.38±0.22 b
B1-4	0.00±0.00	—	0.00±0.00	—
B1-4₅₀	88.00±2.00 c	2.67±0.24 c	72.67±3.06 c	2.41±0.28 c
B1-4₁₀₀	95.33±1.15 a	4.88±0.29 a	93.33±2.31 a	3.76±0.33 a
B1-4₂₀₀	92.00±2.00 b	4.49±0.14 b	84.67±1.15 b	3.22±0.19 b
NB	91.33±1.15 b	4.49±0.16 b	84.67±4.16 b	3.20±0.23 b