Metarhizium rileyi SL Strain Liquid Fermentation Conditions: Optimization by Response Surface Methodology

doi:10.11924/j.issn.1000-6850.casb2022-0343

Abstract

Abstract:

Metarhizium rileyi is a widely distributed entomopathogenic fungi with high pathogenicity to noctuid pests. The present study aims to optimize the liquid fermentation conditions of Metarhizium rileyi, and further increase fermentation broth biomass. In this experiment, the single factor experiment was selected for evaluating the effect of three culture solutions on fermentation broth biomass of Metarhizium rileyi, and screened the main factors affecting bioyield by Plackett-Burman. The suitable range of key factors was determined by the steepest climbing test. The multiple regression equation of mycelial biomass in liquid fermentation of Metarhizium rileyi SL200714 was established by Response Surface Method. The results showed that the strain of Metarhizium rileyi SL200714 was subjected to liquid fermentation using SDY medium under the conditions of culture temperature of 25.08℃, rotating speed of 182.11 r/min, and liquid volume of 100.03/250 mL. In this optimal culture solution, 0.361 g mycelial biomass could be produced per 10 mL fermentation broth. Response surface methodology was used to optimize the liquid fermentation culture conditions of Metarhizium rileyi, and to improve the biological yield per milliliter of Metarhizium rileyi fermentation broth, and the study could lay a foundation for the subsequent industrial production of Metarhizium rileyi conidial powder.

Key words: Metarhizium rileyi, liquid fermentation, Plackett-Burman, response surface methodology, biomass

XUE Rui, SHEN Shaoyun, DENG Xizhou, CHEN Cong, PENG Yuejin, DU Guangzu, CHEN Bin. Metarhizium rileyi SL Strain Liquid Fermentation Conditions: Optimization by Response Surface Methodology[J]. Chinese Agricultural Science Bulletin, 2023, 39(6): 135-143.

Figures/Tables 10

References 32

[1]	杜广祖, 郑亚强, 陈斌, 等. 几种植物油对莱氏野村菌分生孢子萌发的影响[J]. 微生物学通报, 2015, 42(4):690-698.
[2]	KISH L P, SAMSON R A, ALLEN G E. The genus Nomuraea Maublance[J]. Journal of invertebrate pathology, 1974, 24(2):154-158. doi: 10.1016/0022-2011(74)90004-4 URL
[3]	KISH L P, ALLEN G E. The biology and ecology of Nomuraea rileyi and a program for predicting its incidence on Anticarsia gemmatalis in soybean[J]. Florida agricultural experiment station, 1978, 48(2):47-48.
[4]	KEPLER R M, HUMBER R A, BISCHOFF J F, et al. Clarification of generic and species boundaries for Metarhizium and related fungi through multigene phylogenetics[J]. Mycologia, 2014, 106(4):811-829. doi: 10.3852/13-319 URL
[5]	VISALAKSHI M, VARMA P, KISHORE, S V, et al. Studies on mycosis of Metarhizium (Nomuraea) rileyi on Spodoptera frugiperda infesting maize in Andhra Pradesh, India[J]. Egyptian journal of biological pest control, 2020, 30(1):135-145. doi: 10.1186/s41938-020-00335-9 URL
[6]	龙秀珍, 高旭渊, 曾宪儒, 等. 一株莱氏绿僵菌的筛选及其对草地贪夜蛾的毒力[J]. 中国生物防治学报, 2021, 37(6):1111-1119.
[7]	EDEGAR F, ALEXANDRE S, HORACIO H, et al. Metarhizium (Nomuraea) rileyi as biological control agent[J]. Biocontrol science and technology, 2017, 27(11):1-22. doi: 10.1080/09583157.2016.1258776 URL
[8]	丁雪玲, 姚凤銮, 郑宇, 等. 莱氏绿僵菌FJMR2对草地贪夜蛾不同虫态的致病性研究[J]. 应用昆虫学报, 2022, 59(3):551-558.
[9]	杜广祖, 张小娇, 罗艳, 等. 莱氏绿僵菌Nr0815分生孢子在化学农药中的相容性[J]. 河南农业科学, 2020, 49(5):104-110.
[10]	刘守柱, 张蕾蕾, 胥佳睿, 等. 亚致死剂量高效氯氰菊酯对莱氏绿僵菌的增效作用[J]. 河南农业科学, 2021, 50(5):74-79.
[11]	陶新娉, 王彬, 管清真, 等. 莱氏绿僵菌对斜纹夜蛾的致病力及生理效应[J]. 微生物学通报, 2017, 44(6):1349-1357.
[12]	KIM S, KIM J C, LEE S J, et al. Beauveria bassiana ERL836 and JEF-007 with similar virulence show different gene expression when interacting with cuticles of western flower thrips, Frankniella occidentalis[J]. BMC genomics, 2020, 21(1):836-836. doi: 10.1186/s12864-020-07253-y pmid: 33246406
[13]	ROSANA A R R, POKORNY S, KLUTSCH J G, et al. Selection of entomopathogenic fungus Beauveria bassiana (Deuteromycotina:Hyphomycetes) for the biocontrol of Dendroctonus ponderosae (Coleoptera:Curculionidae, Scolytinae) in Western Canada[J]. Applied microbiology and biotechnology, 2021, 105(6): 1-17. doi: 10.1007/s00253-020-10971-8 URL
[14]	FENG M G, POPRAWSKI T J, KHACHATOURIANS G G. Production, formulation and application of the entomopathogenic fungus Beauveria bassi and for insect control:current status[J]. Biocontrol science and technology, 1994, 4(1):3-34. doi: 10.1080/09583159409355309 URL
[15]	杜广祖, 郑亚强, 陈斌, 等. 莱氏野村菌Nr120815菌株在不同稻谷基质上的产孢特性研究[J]. 云南农业大学学报:自然科学, 2016, 31(5):791-798.
[16]	林健文, 罗志兵, 骆星丹, 等. 莱氏野村菌固体发酵培养基筛选与发酵条件优化[J]. 植物保护学报, 2009, 36(5):437-442.
[17]	DEVI P S V. Conidia production of the entomopathogenic fungus Nomuraea rileyi and its evaluation for control of Spodoptera litura (Fab) on ricinus communis[J]. Academic press, 1994, 63(2):145-150.
[18]	唐维媛, 邢丛丛, 董永刚, 等. 对斜纹夜蛾高致病性莱氏野村菌Nr19固体培养条件的优化[J]. 中国生物防治学报, 2016, 32(2):180-188.
[19]	HEGEDUS D D, BIDOCHKA M J, KHACHATOURIANS G G. Beauveria bassiana submerged conidia production in a defined medium containing chitin, two hexosamines or glucose[J]. Applied microbiology biotechnology, 1990, 33:641-647. doi: 10.1007/BF00604930 URL
[20]	ROMBACH M C. Production of Beauveria bassiana (Deuteromycotina: Hyphomycetes) sympoduloconidia in submerged culture[J]. Entomophaga, 1989, 34(1):45-52. doi: 10.1007/BF02372586 URL
[21]	PHAM T A, KIM J J, KIM S G, et al. Production of blastospore of entomopathogenic Beauveria bassianain a submerged batch culture[J]. Mycobiology, 2009, 37(3):218-224. doi: 10.4489/MYCO.2009.37.3.218 URL
[22]	张功营, 杨华, 董丽红, 等. 响应面法优化绿僵菌产绿僵菌素A的培养条件[J]. 华南农业大学学报, 2018, 39(1):76-82.
[23]	李姝江, 彭娇洋, 朱涵明月, 等. 莱氏野村菌(Nomuraea rileyi)固液双相发酵配方研究[J]. 四川林业科技, 2017, 38(5):33-37.
[24]	董海龙. 疣孢漆斑菌(Myrothecium verrucaria)对蔬菜根结线虫寄生致死作用及应用研究[D]. 晋中: 山西农业大学, 2015:47.
[25]	刘朋肖, 常煦, 成柳洁, 等. 酿酒酵母Y3401产己酸乙酯发酵条件的优化[J]. 中国食品学报, 2022, 22(2):178-189.
[26]	尹乐斌, 李乐乐, 何平, 等. 枯草芽孢杆菌发酵豆渣制备多肽及其活性研究[J]. 中国酿造, 2022, 41(1):75-79.
[27]	张朝正, 李意, 赵华. 响应面法优化壳聚糖酶发酵培养基[J]. 中国酿造, 2022, 41(1):197-203.
[28]	李义华, 刁红亮, 周稳稳, 等. 碳源对玫烟色虫草菌丝生长、芽生孢子产生及其耐热性的影响[J]. 菌物学报, 2022, 41(1):51-58.
[29]	吴正铠. 中国虫生真菌研究与应用:第2卷[M]. 北京: 中国农业科技出版社, 1991:5-11.
[30]	程红艳, 郭伟, 常明昌, 等. 蛹虫草菌丝体液体发酵培养条件的优化研究[J]. 山西农业大学学报:自然科学版, 2011, 31(1):66-72.
[31]	郭霄飞, 李艳婷, 王志伟, 等. 雁北嗜蓝孢孔菌液体发酵条件的优化[J]. 山西农业科学, 2020, 48(12):1950-1954.
[32]	杨小龙, 石朝慧, 乔燚, 等. 基于聚氨酯海绵载体的棘孢木霉固体发酵培养基筛选[J]. 中国生物防治学报, 2022, 38(1):88-96.

编号	因素	水平
编号	因素	-1	1
A	温度	20℃	30℃
B	转速	150 r/min	210 r/min
C	装液量	50 mL	150 mL
D	pH	5	7
E	接种量	10⁵孢子/mL	10⁷孢子/mL
F	无机盐含量	0%	5%
G	维生素含量	0%	5%
H	空项
I	空项

编号	因素	水平
编号	因素	-1	1
A	温度	20℃	30℃
B	转速	150 r/min	210 r/min
C	装液量	50 mL	150 mL
D	pH	5	7
E	接种量	10⁵孢子/mL	10⁷孢子/mL
F	无机盐含量	0%	5%
G	维生素含量	0%	5%
H	空项
I	空项

Source	Sum of Squares	df	Mean Square	F-value	P-value
Model	0.1538	7	0.022	11.87	0.0154
A-温度	0.0865	1	0.0865	46.77	0.0024
B-转速	0.0294	1	0.0294	15.87	0.0163
C-装液量	0.0146	1	0.0146	7.89	0.0483
D-pH	0.0001	1	0.0001	0.0645	0.8121
E-接种量	0.0136	1	0.0136	7.32	0.0537
F-无机盐含量	0.0003	1	0.0003	0.1689	0.7021
G-维生素含量	0.0093	1	0.0093	5.03	0.0884
Residual	0.0074	4	0.0019
Cor Total	0.1612	11

Source	Sum of Squares	df	Mean Square	F-value	P-value
Model	0.1538	7	0.022	11.87	0.0154
A-温度	0.0865	1	0.0865	46.77	0.0024
B-转速	0.0294	1	0.0294	15.87	0.0163
C-装液量	0.0146	1	0.0146	7.89	0.0483
D-pH	0.0001	1	0.0001	0.0645	0.8121
E-接种量	0.0136	1	0.0136	7.32	0.0537
F-无机盐含量	0.0003	1	0.0003	0.1689	0.7021
G-维生素含量	0.0093	1	0.0093	5.03	0.0884
Residual	0.0074	4	0.0019
Cor Total	0.1612	11

序号	温度/℃	转速/(r/min)	装液量/mL	生物量/(g/10mL)
1	21	120	60	0.1134
2	23	140	80	0.2735
3	25	160	100	0.3224
4	27	180	120	0.3021
5	29	200	140	0.1423