一株产表面活性素芽胞杆菌发酵优化及其防治应用

doi:10.13560/j.cnki.biotech.bull.1985.2025-0996

摘要/Abstract

摘要：

目的对一株来自沙漠土壤分离的产表面活性素的菌株Bacillus australimaris TRM82479进行发酵条件优化，并检测其对农业害虫的防治作用。方法通过Plackett-Burman实验筛选关键因素，结合Box-Behnken响应面实验优化发酵条件，采用排油圈法和HPLC-MS方法测定了碳源、氮源等多种营养物质以及培养时间、温度、摇床转速、初始pH、接种量、装液量对该菌株产表面活性素（surfactin）的产量影响；采用饲料染毒法和叶片浸渍法测定菌液的杀虫谱；采用五点取样法测定不同浓度菌液的大田防效。结果适合该菌表面活性素产生的最佳液体培养基组成及发酵参数为：麦芽糖8.912 g/L、酵母浸粉7.135 g/L、氯化钠7 g/L、硫酸镁0.2 g/L，发酵温度28 ℃、发酵时间75 h、转速160 r/min、装液量150 mL、接种量4 %；surfactin产量较优化前提高了约1.88倍，浓度约为1 355.78 mg/L。菌液对7种农业害虫都有一定的致死作用，杀虫谱较广，其中对蚜虫致死率最高，48 h校正死亡率达到了91.23%。田间试验中，浓度为10⁸、10⁷、10⁶ CFU/mL的上清液对蚜虫防治7 d后的防效分别为77.33%、70.48%和66.26%，均高于化学农药25%氟啶虫酰胺的56.95%。结论菌株TRM82479发酵工艺优化后，产量达到了1 355.78 mg/L，对多种农业害虫有良好的防治作用，对蚜虫7 d后的大田防效高达77.33%，显著高于化学农药25%氟啶虫酰胺，具有开发为微生物杀虫剂的巨大潜力。

关键词: 芽胞杆菌, 表面活性素, 排油圈法, 响应面实验, 广谱杀虫

Abstract:

Objective This study aimed to optimize the fermentation conditions and evaluate the insecticidal activity of a surfactin-producing strain, Bacillus australimaris TRM82479, isolated from desert soil. Method Key influencing factors were screened using the Plackett-Burman design, followed by optimization of fermentation conditions via the Box-Behnken response surface methodology. The effects of various nutrients (including carbon and nitrogen sources), cultivation time, temperature, shaking speed, initial pH, inoculation volume, and medium volume on surfactin production were determined using the oil displacement method and HPLC-MS. The insecticidal spectrum of the bacterial suspension was determined using the feed poisoning method and leaf dipping method; and the field control effects of the bacterial suspension at different concentrations were evaluated via the five-point sampling method. Result The optimal liquid medium composition and fermentation parameters for surfactin production were as follows: Maltose 8.912 g/L, yeast extract powder 7.135 g/L, sodium chloride 7 g/L, magnesium sulfate 0.2 g/L, fermentation temperature 28 ℃, fermentation time 75 h, shaking speed 160 r/min, medium volume 150 mL, and vaccination rate of 4%.Under these optimized conditions, surfactin yield increased approximately 1.88-fold, reaching a concentration of about 1 355.78 mg/L. The bacterial culture had insecticidal activity against seven agricultural pests, demonstrating a broad insecticidal spectrum. The highest mortality was observed against aphids, with a corrected mortality rate of 91.23% at 48 h. In field trials, the supernatants at concentrations of 10⁸, 10⁷, and 10⁶ CFU/mL provided control efficacies against aphids of 77.33%, 70.48%, and 66.26%, respectively, after seven days, all of which were higher than the 56.95% efficacy achieved by the chemical pesticide flonicamid (25%). Conclusion After optimizing the fermentation process of strain TRM82479, the yield reaches 1 355.78 mg/L, it exerts a good control effect on multiple agricultural pests, and its field control effect against aphids after 7 d is as high as 77.33%. This is significantly higher than that of the chemical pesticide 25% flonicamid, indicating great potential for development as a microbial insecticide.

Key words: Bacillus australimaris, surfactin, oil displacement assay, response surface methodology, broad-spectrum insecticide

苏畅, 文凤, 王业林, 邵瑞盈, 唐家杰, 夏占峰. 一株产表面活性素芽胞杆菌发酵优化及其防治应用[J]. 生物技术通报, 2026, 42(4): 332-344.

SU Chang, WEN Feng, WANG Ye-lin, SHAO Rui-ying, TANG Jia-jie, XIA Zhan-feng. Fermentation Optimization of a Surfactin-producing Bacillus Strain and Its Application in Prevention and Control[J]. Biotechnology Bulletin, 2026, 42(4): 332-344.

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因素 Factor	编码水平 Horizontal encoding
因素 Factor	-1	0	1
酵母浸粉 Yeast （g/L）	6	7	8
麦芽糖 Maltose （g/L）	10.5	9	7.5
摇床转速 Rotational speed （r/min）	150	160	170

因素 Factor	编码水平 Horizontal encoding
因素 Factor	-1	0	1
酵母浸粉 Yeast （g/L）	6	7	8
麦芽糖 Maltose （g/L）	10.5	9	7.5
摇床转速 Rotational speed （r/min）	150	160	170

组别 Group	碳源种类 Carbon source （g/L）	氮源种类 Nitrogen source （g/L）	排油圈直径 Diameter of oil ring drainage （mm）
1	淀粉	大豆蛋白胨	20.15
2	燕麦	酸水解酪蛋白	40.44
3	燕麦	大豆蛋白胨	18.13
4	麦芽糖	酸水解酪蛋白	29.83
5	燕麦	酵母浸粉	38.24
6	淀粉	酵母浸粉	30.78
7	麦芽糖	酵母浸粉	51.03
8	淀粉	酸水解酪蛋白	30.98
9	麦芽糖	大豆蛋白胨	39.11

组别 Group	碳源种类 Carbon source （g/L）	氮源种类 Nitrogen source （g/L）	排油圈直径 Diameter of oil ring drainage （mm）
1	淀粉	大豆蛋白胨	20.15
2	燕麦	酸水解酪蛋白	40.44
3	燕麦	大豆蛋白胨	18.13
4	麦芽糖	酸水解酪蛋白	29.83
5	燕麦	酵母浸粉	38.24
6	淀粉	酵母浸粉	30.78
7	麦芽糖	酵母浸粉	51.03
8	淀粉	酸水解酪蛋白	30.98
9	麦芽糖	大豆蛋白胨	39.11

来源 Source	平方和 Sum of squares	自由度 DF	均方 Mean square	F值 F-value	P值 P-value	显著性 Significance
模型 Model	3 699.19	8	462.40	18.73	0.017 4	*
A	265.27	1	265.27	10.74	0.046 5	*
B	2 278.11	1	2 278.11	92.25	0.002 4	**
C	262.95	1	262.95	10.65	0.047	*
D	68.23	1	68.23	2.76	0.195 1
E	39.07	1	39.07	1.58	0.297 4
F	70.7	1	70.7	2.86	0.189 2
G	651.02	1	651.02	26.36	0.014 3	*
H	63.85	1	63.85	2.59	0.206 2
残差 Residual	74.08	3	24.69
总和 Cor total	3 773.27	11
R²	0.980 4			Predicted R²	0.685 9
R²_adj	0.928 0			Adeq precision	11.991 4