一株灰茶尺蠖高毒力金龟子绿僵菌的筛选、鉴定与培养条件

李良德; 李金玉; 李慧玲; 张辉; 王定锋

doi:10.13305/j.cnki.jts.2025.03.007

茶叶科学 >

2025 , Vol. 45 >Issue 3: 427 - 438

DOI: https://doi.org/10.13305/j.cnki.jts.2025.03.007

研究报告

一株灰茶尺蠖高毒力金龟子绿僵菌的筛选、鉴定与培养条件

李良德 ,
李金玉 ,
李慧玲 ,
张辉 ,
王定锋

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福建省农业科学院茶叶研究所,福建福州 350013

李良德,男,助理研究员,主要从事茶树害虫分子生物学及防控技术研究。

收稿日期: 2024-12-06

修回日期: 2025-02-12

网络出版日期: 2025-06-18

基金资助

福建省属公益类科研院所基本科研专项（2021R1029009）、福建省农业科学院青年英才项目（YC2021008）、福建省农业科学院科技创新团队项目（CXTD2021001-3）

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Screening, Identification and Culture Conditions of a Highly Virulent Metarhizium anisopliae Strain against Ectropis grisescens Warre

LI Liangde ,
LI Jinyu ,
LI Huiling ,
ZHANG Hui ,
WANG Dingfeng

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Tea Research Institute, Fujian Academy of Agricultural Sciences, Fuzhou 350013, China

Received date: 2024-12-06

Revised date: 2025-02-12

Online published: 2025-06-18

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摘要

绿僵菌（Metarhizium）是一类重要的虫生真菌,在农林害虫生物防治中被广泛应用。为从7个保存的菌种中筛选出对灰茶尺蠖毒力最强的菌株,并验证该菌株在不同剂量下的毒力效果,通过形态学特征观察和多基因系统进化树分析方法确定其分类地位,并对菌株生长的最适培养基、温度、pH进行了研究。结果表明,供试的7个菌株对灰茶尺蠖3龄幼虫具有不同的防治效果,菌株Ma54的毒力最强,处理10 d后的校正死亡率达81.67%,半致死时间（LT₅₀）为4.93 d;灰茶尺蠖的死亡率与菌株Ma54的孢子浓度呈剂量效应,高浓度（1.0×10⁸个·mL^-1）处理7 d后,死亡率达98.33%,LT₅₀为4.09 d;菌株Ma54感染灰茶尺蠖7 d后,虫尸长满菌丝、呈黑绿色,在PDA培养基上生长14 d后,分生孢子向外扩散、呈深绿色;菌株Ma54的（Internal transcribed spacer,ITS）、β微管蛋白（β-tubulin）、RNA聚合酶Ⅱ第一大亚基RPB1基因的系统进化树都与金龟子绿僵菌（Metarhizium anisopliae）处在相同分支;菌株Ma54的菌落直径、单位面积产孢量、孢子萌发率在PPDA培养基、温度25 ℃、pH 7.0条件下最为理想。该研究获取了对灰茶尺蠖具有高毒力的菌株Ma54,鉴定为金龟子绿僵菌,初步明确了菌株的生长条件,为后续灰茶尺蠖的生物防治奠定了基础。

关键词： 灰茶尺蠖; 金龟子绿僵菌; 高毒力; 分类地位; 生长条件

本文引用格式

李良德 , 李金玉 , 李慧玲 , 张辉 , 王定锋 . 一株灰茶尺蠖高毒力金龟子绿僵菌的筛选、鉴定与培养条件[J]. 茶叶科学, 2025 , 45(3) : 427 -438 . DOI: 10.13305/j.cnki.jts.2025.03.007

Abstract

As an important entomogenous fungus, Metarhizium has been extensively utilized in the biological control of agricultural and forestry pests. This study aimed to screen the most infective strain against E. grisescens from 7 preserved strains and verify its infectivity at different doses. Subsequently, the taxonomic status of the strain was determined through observation of morphological characteristics and methods for multigene phylogenetic tree analysis. Finally, the optimal culture medium, temperature and pH value for strain growth were investigated. The results indicate that the seven tested strains exhibited different control effects on the 3rd instar larvae of E. grisescens, with strain Ma54 exhibiting the highest virulence, achieving a corrected mortality rate of 81.67% after 10 d of treatment and an LT₅₀ (lethal time for 50% mortality) of 4.93 d. The mortality rate of E. grisescens shows a dose-response relationship with the spore concentration of strain Ma54. After 7 d of treatment under a high concentration of Metarhizium (1.0×10⁸spores·mL^-1), the mortality rate reached 98.33%, and the LT₅₀value was 4.09 days. Seven days after strain Ma54 infected E. grisescens, the cadavers were covered with mycelia and turned dark green. After 14 days of growth on PDA medium, the conidia further spread and appeared dark green. The phylogenetic trees of the ribosomal transcription factor, β-tubulin, and the largest subunit of RNA polymerase Ⅱ of strain Ma54 all clustered with Metarhizium anisopliae. The colony diameter, spore production per unit area, and spore germination rate of strain Ma54 were optimal under conditions of a PPDA medium, a temperature of 25 ℃, and a pH value of 7.0. This study obtained the Ma54 strain with high virulence against E. grisescens, authenticated it as M. anisopliae, and preliminarily clarified its growth conditions of the strain, laying a foundation for its subsequent utilization in the biological control of E. grisescens.

Key words： Ectropis grisescens; Metarhizium anisopliae; high virulence; taxonomic status; growth conditions

参考文献

[1] 李良德, 王定锋, 李慧玲, 等. 茶尺蠖蜕皮激素受体基因Eo-EcR的克隆、生物信息学分析和表达检测[J]. 昆虫学报, 2015, 58(10): 1063-1071.
Li L D, Wang D F, Li H L, et al.Molecular cloning, bioinformatics analysis and expression profiling of ecdysteroid receptor gene Eo-EcR in Ectropis obliqua (Lepidoptera: Geometridae)[J]. Acta Entomologica Sinica, 2015, 58(10): 1063-1071.
[2] 王志博, 刘永健, 白家赫, 等. 沃尔巴克氏菌调控小茶尺蠖和灰茶尺蠖杂交卵孵化[J]. 昆虫学报, 2022, 65(8): 999-1009.
Wang Z B, Liu Y J, Bai J H, et al.Wolbachia regulates the egg hatching of hybrids of Ectropis obliqua and E. grisescens (Lepidoptera: Geometridae)[J]. Acta Entomologica Sinica, 2022, 65(8): 999-1009.
[3] 郭世保, 陈俊华, 张龙, 等. 蠋蝽对灰茶尺蠖幼虫的捕食能力及种内干扰反应[J]. 茶叶科学, 2024, 44(4): 609-617.
Guo S B, Chen J H, Zhang L, et al.The predatory ability and intraspecific interference response of Arma chinensis to the larvae of Ectropis grisescens[J]. Journal of Tea Science, 2024, 44(4): 609-617.
[4] 税良勇, 赵忠祎, 冯印, 等. 茶尺蠖羧酸酯酶EoCarE592的重组表达及其对农药的降解能力测定[J]. 昆虫学报, 2024, 67(1): 48-57.
Shui L Y, Zhao Z Y, Feng Y, et al.Recombinant expression of carboxylesterase EoCarE592 of Ectropis obliqua (Lepidoptera: Geometridae) and determination of its ability to degrade pesticides[J]. Acta Entomologica Sinica, 2024, 67(1): 48-57.
[5] 刘蓉, 蔡霓, 农向群, 等. 金龟子绿僵菌对苜蓿生长的影响及对苜蓿抗蚜性的诱导作用[J]. 植物保护学报, 2024, 51(2): 442-449.
Liu R, Cai N, Nong X Q, et al.Effects of entomopathogenic fungus Metarhizium anisopliae on alfalfa growth promotion and anti-aphid induction[J]. Journal of Plant Protection, 2024, 51(2): 442-449.
[6] 万依依, 张华峰, 倪德芳, 等. 金龟子绿僵菌对入侵性害虫椰子织蛾的致病力[J]. 山东农业大学学报(自然科学版), 2023, 54(5): 663-669.
Wan Y Y, Zhang H F, Ni D F, et al.Pathogenicity of Metarhizium anisopliae to invasive insect Opisina arenosella walker[J]. Journal of Shandong Agricultural University (Natural Science Edition), 2023, 54(5): 663-669.
[7] 郑丽莎. 金龟子绿僵菌和球孢白僵菌可湿性菌剂的制备及其对玉米螟的杀虫效果[D]. 泰安: 山东农业大学, 2024.
Zheng L S.Preparation of wettable bactericides of Metarhizium Anisopliae and Beauveria bassiana and their insecticidal effect on corn borer [D]. Taian: Shandong Agricultural University, 2024.
[8] 陈峰, 王长方, 王俊, 等. 一株金龟子绿僵菌对茶尺蠖的致病力和胞外酶活性[J]. 中国生物防治学报, 2021, 37(1): 124-129.
Chen F, Wang C F, Wang J, et al.Pathogenicity and extracellular enzyme activities of Metarhizium anisopliae to Ectropis obliqua hypulina Wehrli[J]. Chinese Journal of Biological Control, 2021, 37(1): 124-129.
[9] 谭格, 李天铭, 周志成, 等. 金龟子绿僵菌对斜纹夜蛾幼虫的生防效果、抗氧化酶活性和肠道细菌群落的影响[J]. 微生物学报, 2021, 61(5): 1171-1183.
Tan G, Li T M, Zhou Z C, et al.Biocontrol of Spodoptera litura larvae by Metarhizium anisopliae[J]. Acta Microbiologica Sinica, 2021, 61(5): 1171-1183.
[10] 邓璇璇, 匡炜, 方宝华, 等. 金龟子绿僵菌制剂对二化螟的防效及体内相关酶活性的影响[J]. 湖南农业大学学报(自然科学版), 2024, 50(4): 65-69.
Deng X X, Kuang W, Fang B H, et al.Effects of Metarhizium anisopliae preparation against Chilo suppressalis and on related enzyme activities in vivo[J]. Journal of Hunan Agricultural University (Natural Sciences), 2024, 50(4): 65-69.
[11] 康奎, 蔡尤俊, 龚俊, 等. 金龟子绿僵菌对褐飞虱繁殖力的影响[J]. 中国生物防治学报, 2022, 38(1): 180-187.
Kang K, Cai Y J, Gong J, et al.Effects of Metarhizium anisopliae on the fecundity of Nilaparvata lugens[J]. Chinese Journal of Biological Control, 2022, 38(1): 180-187.
[12] 张谦, 王燕, 董明, 等. 金龟子绿僵菌与化学农药减量联用对棉花苗蚜的防控效果[J]. 中国生物防治学报, 2023, 39(4): 797-803.
Zhang Q, Wang Y, Dong M, et al.Control of cotton seedling aphids using Metarhizium anisopliae combined with reduced-dosages chemical pesticides[J]. Chinese Journal of Biological Control, 2023, 39(4): 797-803.
[13] 裴松松, 吴轩, 李瑞军, 等. 对西花蓟马高效金龟子绿僵菌菌株筛选及在花生田间的应用效果[J]. 中国生物防治学报, 2021, 37(4): 732-739.
Pei S S, Wu X, Li R J, et al.Screening of efficient Metarhizium anisopliae strains for Frankliniella occidentalis and its application in peanut field[J]. Chinese Journal of Biological Control, 2021, 37(4): 732-739.
[14] 陈祝安, 黄基荣. 不同来源绿僵菌对云斑金龟蛴螬致病力评价[J]. 微生物学通报, 1997, 24(2): 81-83.
Chen Z A, Huang J R.Assessment of virulence on strains from different origin in Metarhizium anisopliae for the white grub[J]. Microbiology China, 1997, 24(2): 81-83.
[15] 张亚倩, 谷静秀, 苑士涛, 等. 2亿活孢子/g金龟子绿僵菌CQMa421颗粒剂防治花生地老虎田间药效试验[J]. 农药, 2019, 58(9): 684-686.
Zhang Y Q, Gu J X, Yuan S T, et al.Field efficacy test of 200 million live spores/gram Metarhizium metarhiziae CQMa421 granules in controlling peanut cutworm[J]. Agrochemicals, 2019, 58(9): 684-686.
[16] 谭荣荣, 陈勋, 黄丹娟, 等. 灰茶尺蠖核型多角体病毒对两种尺蠖的致病性[J]. 中国生物防治学报, 2023, 39(3): 684-689.
Tan R R, Chen X, Huang D J, et al.Pathogenicity of Ectropis grisescens Nucleopolyhedrovirus on Ectropis grisescens Warren and Ectropis obliqua Prout[J]. Chinese Journal of Biological Control, 2023, 39(3): 684-689.
[17] 王峰, 刘斌, 农向群, 等. 绿僵菌属4个生防潜力菌株的多基因鉴定[J]. 中国生物防治学报, 2017, 33(6): 780-787.
Wang F, Liu B, Nong X Q, et al.Multigene identification of four potential biocontrol strains in Metarhizium genus[J]. Chinese Journal of Biological Control, 2017, 33(6): 780-787.
[18] 王定锋, 李良德, 李慧玲, 等. 一株对茶丽纹象甲高毒力白僵菌菌株的筛选、鉴定与培养研究[J]. 茶叶科学, 2021, 41(1): 101-112.
Wang D F, Li L D, Li H L, et al.Screening, identification and culture conditions of a high-virulent isolate of Beauveria against Myllocerinus Aurolineatus[J]. Journal of Tea Science, 2021, 41(1): 101-112.
[19] 王定锋, 刘丰静, 李慧玲, 等. 球孢白僵菌XJBb3005对茶丽纹象甲致病力的时间-剂量-死亡率模型分析[J]. 福建农业学报, 2013, 28(8): 807-811.
Wang D F, Liu F J, Li H L, et al.Time-dose-mortality model analysis of Beauveria bassiana XJBb3005 against Myllocerinus aurolineatus[J]. Fujian Journal of Agricultural Sciences, 2013, 28(8): 807-811.
[20] 侯明明, 刘义豪, 王滨. 昆虫病原真菌对梨网蝽防治潜力的室内评测[J]. 中国生物防治学报, 2015, 31(6): 853-859.
Hou M M, Liu Y H, Wang B.Laboratory assessment on virulence of entomogenous fungi against Stephanitis nashi[J]. Chinese Journal of Biological Control, 2015, 31(6): 853-859.
[21] 王萌, 殷幼平, 王中康, 等. 10株绿僵菌菌株分类地位的多基因系统进化分析[J]. 植物保护, 2014, 40(5): 14-21.
Wang M, Yin Y P, Wang Z K, et al.Multilocus phylogenetic analysis of the taxonomic status of 10 strains of Metarhizium[J]. Plant Protection, 2014, 40(5): 14-21.
[22] Driver F, Milner R J, Trueman J W H. A taxonomic revision of Metarhizium based on a phylogenetic analysis of rDNA sequence data[J]. Mycological Research, 2000, 104(2): 134-150.
[23] Bischoff J F, Rchncr S A, Humber R A.Metarhizium frigidum spp, nov.: a cryptic species of M. anisopliae and a member of the M. flavoviride complex[J]. Mycologia, 2006, 98(5): 737-745.
[24] Bischoff J F, Rehner S A, Humber R A.A multilocus phylogeny of the Metarhizium anisopliae lineage[J]. Mycologia, 2009, 101(4): 512-530.
[25] Fernández-Bravo M, Garrido-Jurado I, Valverde-García P, et al.Responses to abiotic environmental stresses among phylloplane and soil isolates of Beauveria bassiana from two holm oak ecosystems[J]. Journal of Invertebrate Pathology, 2016, 141(9): 6-17.
[26] 林华峰, 王萍莉, 张磊, 等. 布氏白僵菌和金龟子绿僵菌两变种的生长性状及其对蛴螬的毒力测定[J]. 中国生物防治学报, 2006, 22(2): 123-127.
Lin H F, Wang P L, Zhang L, et al.Growth of Beauveria brongniartii and two varieties of Metarhizium anisopliae and their virulence against white grubs[J]. Chinese Journal of Biological Control, 2006, 22(2): 123-127.

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