三种昆虫病原线虫非生物胁迫耐受特性及其共生菌抑菌作用

doi:10.6048/j.issn.1001-4330.2023.12.024

新疆农业科学 ›› 2023, Vol. 60 ›› Issue (12): 3072-3079.DOI: 10.6048/j.issn.1001-4330.2023.12.024

• 畜牧兽医·微生物·农业装备工程与机械化 • 上一篇下一篇

三种昆虫病原线虫非生物胁迫耐受特性及其共生菌抑菌作用

陈澄¹(), 侯新强²(), 詹发强¹^,²(), 杨蓉², 包慧芳², 王宁², 史应武², 龙宣杞²

1.新疆大学生命科学与技术学院,乌鲁木齐 830046
2.新疆农业科学院微生物应用研究所/新疆特殊环境微生物重点实验室,乌鲁木齐 830091

收稿日期:2023-05-06 出版日期:2023-12-20 发布日期:2024-01-03
通信作者: 詹发强(1980-),男,新疆人,研究员,研究方向为农业微生物与生物防治,(E-mail)zfq_xj@126.com;侯新强(1976-),男,新疆人,副研究员,研究方向为农业微生物应用, (E-mail)996970105@qq.com
作者简介:陈澄(1998-),女,安徽人,硕士研究生,研究方向为生物与医药,(E-mail)330445212@qq.com
基金资助:
新疆维吾尔自治区公益性科研院所基本科研业务经费项目“新疆昆虫病原线虫资源收集鉴定与高活性抗逆品系筛选”(KY2021119);国家自然科学基金“塔河中游胡杨林昆虫病原线虫资源及其优势种适应性机制研究”(32160377);农业科技创新稳定支持项目“天敌资源挖掘和微生物农药创制及其应用”(xjnkywdzc-2022004-3);新疆维吾尔自治区“三农”骨干人才培养项目“新型杀虫防病微生物农药创制”(2022SNGGGCC028)

Study on abiotic stress tolerance of three entomopathogenic nematodes and its bacteriostasis of symbiotic bacteria

CHEN Cheng¹(), HOU Xinqiang²(), ZHAN Faqiang¹^,²(), YANG Rong², BAO Huifang², WANG Ning², SHI Yingwu², LONG Xuanqi²

1. College of Life Sciences and Technology, Xinjiang University, Urumqi 830046, China
2. Institute of Microbiology, Xinjiang Academy of Agricultural Sciences/ Xinjiang Key Laboratory of Special Environmental Microbiology, Urumqi 830091, China

Received:2023-05-06 Online:2023-12-20 Published:2024-01-03
Correspondence author: ZHAN faqiang (1980-), male, researcher, research direction: Application of agricultural microorganism and biocontrol, (E-mail)zfq_xj@126.com;HOU Xinqiang (1976-), male, associate researcher, research direction: Application of agricultural microorganisms, (E-mail)996970105@qq.com
Supported by:
Basic research business funding Project for public welfare research institutes of Xinjiang Uygur Autonomous Region "Identification of entomopathogenic nematode resources and screening of highly active and resistant strains in Xinjiang"(KY2021119);National Natural Science Foundation of China "Research on the entomopathogenic nematodes resources of Populus euphratica forest in midstream of Tarim River and the abiotic stress adaptation mechanism of dominant species"(32160377);Fund for Stable Support to Agricultural Sci-Tech Renovation "Mining of natural enemy resources and creation of microbial pesticides and their applications"(xjnkywdzc-2022004-3);Xinjiang Uygur Autonomous Region "Three dimensional rural" Backbone Talent Personnel Training Program "Creation of new microbial pesticides for insecticide and Suppressing phytopathogens"(2022SNGGGCC028)

摘要/Abstract

摘要：

【目的】研究三种6株系不同昆虫病原线虫对温度、盐浓度耐受特性,筛选出对非生物胁迫耐受性较好的株系,并测定其对大蜡螟幼虫的致病力,分析其共生细菌对金黄色葡萄球菌、沙门氏菌和大肠杆菌的抑制效果,为拮抗病原微生物提供更多新型拮抗菌资源。【方法】采用稀释法统计线虫总数量,分析不同线虫的繁殖能力;分别运用水浴加热法和盐浓度梯度法分析线虫的耐热性、耐盐性;利用梯度稀释法测定其不同剂量对大蜡螟幼虫的致病力;通过涂布法和打孔法测定不同时间段共生菌上清液对病原菌的抑制作用。【结果】从6种线虫中筛选得到一株繁殖和耐受性综合能力较强的菌株小卷蛾斯氏线虫(Steinernema - carpocapsae)XJ-94,每条大蜡螟体内产出的线虫数量为4.96×10⁴ IJs/larva,大蜡螟幼虫产出线虫数量平均为1.24×10⁴ IJs/g;40℃下,处理5 h后,线虫XJ-94存活率为33%,其余株系线虫均已死亡;在17%盐浓度下,处理3 h后,仍有68%的线虫存活。该线虫对大蜡螟幼虫具有较高的致病力,在48 h时,线虫剂量为160 IJs/larva和200 IJs/larva的校正死亡率达到100%。嗜线虫致病杆菌(Xenorhabdus nematophila)XJ-94共生菌发酵上清液在48 h时的抑菌效果最好,对大肠杆菌(E.coli)和金黄色葡萄球菌(S.aureus)的抑制作用显著高于沙门氏菌(Salmonella)。【结论】筛选得到一株繁殖能力和非生物胁迫耐受性较好的昆虫病原线虫株系,该线虫对大蜡螟幼虫具有较高的致病力,其共生菌对大肠杆菌和金黄色葡萄球菌的抑制作用较好。

关键词: 昆虫病原线虫; 致病力; 共生菌; 病原菌; 抑菌作用

Abstract:

【Objective】To study the tolerance characteristics of three species and 6 strains of entomopathogenic nematodes to temperature and salt concentration, in order to screen the strains with good tolerance to abiotic stress and determine their pathogenicity to Galleria mellonella larvae.to lay a related research foundation for the application of biological control of insect pests in the future.At the same time, the inhibitory effect of symbiotic bacteria on S.aureus, Salmonella and E.coli was studied in order to provide more new antagonistic bacteria resources for antagonistic pathogenic microorganisms.【Methods】The total number of nematodes was counted by dilution method, the reproductive ability of different nematodes was analyzed, and the heat tolerance and salt tolerance of nematodes were analyzed by water bath heating method and salt concentration gradient method, respectively.the pathogenicity of different doses to the Galleria mellonella larvae was determined by gradient dilution method, and the inhibitory effect of symbiotic supernatant on pathogens in different time periods was determined by coating method and drilling method.【Results】A strain of Steinernema-carpocapsae XJ-94 with strong comprehensive ability of reproduction and tolerance was screened from six species of nematodes.The number of nematodes produced by each large wax borer was 4.96 × 10⁴ IJs/larva, and the average number of nematodes per gram of wax borer larvae was 1.24 × 10⁴ IJs/g.After treatment at 40℃ for 5 hours, the survival rate of nematode XJ-94 was 33%, and the other strains were dead.At 17% salt concentration, 68% of the nematodes survived after treatment for 3 hours.The nematode had high pathogenicity to the Galleria mellonella larvae, and the corrected mortality of nematode doses of 160 IJs/larva and 200 IJs/larva reached 100% at 48 h.The fermentation supernatant of Xenorhabdus nematophila XJ-94 symbiotic bacteria had the best bacteriostatic effect at 48h, and its inhibitory effect on E.coli and S.aureus was significantly higher than that of Salmonella.【Conclusion】An entomopathogenic nematode strain with good reproductive ability and abiotic stress tolerance was obtained.The nematode had high pathogenicity to the Galleria mellonella larvae, and its symbiotic bacteria had good inhibitory effect on E.coli and S.aureus.

Key words: entomopathogenic nematode; pathogenicity; symbiotic bacteria; pathogenic bacteria; bacteriostasis

中图分类号:

S188

陈澄, 侯新强, 詹发强, 杨蓉, 包慧芳, 王宁, 史应武, 龙宣杞. 三种昆虫病原线虫非生物胁迫耐受特性及其共生菌抑菌作用[J]. 新疆农业科学, 2023, 60(12): 3072-3079.

CHEN Cheng, HOU Xinqiang, ZHAN Faqiang, YANG Rong, BAO Huifang, WANG Ning, SHI Yingwu, LONG Xuanqi. Study on abiotic stress tolerance of three entomopathogenic nematodes and its bacteriostasis of symbiotic bacteria[J]. Xinjiang Agricultural Sciences, 2023, 60(12): 3072-3079.

图/表 5

表1 大蜡螟幼虫体内分离出来的线虫

Tab.1 Nematodes isolated from larvae of Galleria mellonella

线虫编号 The nematode number	IJs/larva	IJs/g
All	5.24×10⁴	1.31×10⁴
XJ-94	4.96×10⁴	1.24×10⁴
40-4	2.91×10⁴	7.28×10³
XJ-39	3.30×10⁴	8.25×10³
XJ-62	2.68×10⁴	6.70×10³
B13	3.26×10⁴	8.15×10³

图1 线虫在35℃(A)、38℃(B)、40℃(C)温度下的耐受性

Fig.1 Tolerance of nematodes at temperatures of 35℃ (A), 38℃ (B), and 40℃ (C)

图2 线虫在5%(A)、9%(B)、13%(C)、17%(D)NaCl溶液中的耐受性

Fig.2 Tolerance of nematodes in 5% (A), 9% (B), 13% (C), 17% (D) NaCl solutions

图3 不同剂量小卷蛾斯氏线虫XJ-94对大蜡螟幼虫的致病力

Fig.3 Pathogenicity of different dosages of Steinernema - carpocapsae XJ-94 against Galleria mellonella larvae

图4 不同发酵时间对病原菌的抑菌活性注:数据为平均值±标准差,不同小字母表示0.05水平上差异显著

Fig.4 Activity activity against pathogenic bacteria at different fermentation times Note: Data were presented as mean±SD, data with different lowercase letters indicated significant difference at 0.05 level

参考文献 32

[1]	杨秀芬, 杨怀文. 昆虫病原线虫的致病机理[J]. 中国生物防治, 1998, 14(4):38-42.
	YANG Xiufen, YANG Huaiwen. The pathogenic mechanism of entomopathogenic nematodes[J]. Institute of Biological Control, 1998, 14(4):38-42.
[2]	Toepfer S, Kurtz B, Kuhlmann U. Influence of soil on the efficacy of entomopathogenic nematodes in reducing Diabrotica virgifera in maize[J]. Journal of Pest Science, 2010, 83:257-264. PMID
[3]	Bal H K, Grewal P S. Lateral dispersal and foraging behavior of entomopathogenic nematodes in the absence and presence of mobile and non-mobile hosts[J]. PLoS ONE, 2015, 10(6):e0129887. DOI URL
[4]	Poinar Jr., G.O.. Nematodes for Biological Control of Insects[J]. CRC, Boca Raton, FL, 1979,277.
[5]	Kaya H.K., Gaugler R.. Entomopathogenic nematodes[J]. Annual Rev.Entomol, 1993,38,181-206.
[6]	Laznik Ž., Tóth T., Lakatos T., et al. Control of the Colorado potatobeetle (Leptinotarsa decemlineata [Say]) on potato under field conditions: a comparison of the efficacy of foliar application of two strains of Steinernema feltiae (Filipjev) and spraying with thiametoxam[J]. Plant Disease Protect, 2010,117,129-135.
[7]	Akhurst R.J., Boemare N.E.. Biology and taxonomy of Xenorhabdus.In: Gaugler, R., Kaya, H.K.(Eds.), Entomopathogenic Nematodes in Biological Control[J].CRC Press, Boca Raton, FL, USA, 1990:75-90.
[8]	Liu Q Z, Glazer I. Desiccation survival of entomopathogenic nematodes of the genus Heterorhabditis[J]. Phytoparasitica, 2000, 28(4):331-340. DOI URL
[9]	Liu Q Z, Piggott S J, Solomon A, et al. Physiological and biochemical changes in nematodes of the genus Heterorhabditis following desiccation[J]. Phytoparasitica, 2002, 30(3):253-261.
[10]	Chung H J, Lee D W, Yoon H S, et al. Temperature and dose effects on the pathogenicity and reproduction of two Korean isolates of Heterorhabditis bacteriophora (Nematoda: Heterorhabditidae)[J]. Journal of Asia-Pacific Entomology, 2010, 13(4):277-282.
[11]	Dutky S R, Thompson J V, Cantwell G E. A techique for the mass propagation of the DD-136 nematode[J]. Journal of the Insect Pathology, 1964, 6(4):417-422
[12]	杨秀芬, 简恒, 张善稿. 嗜线虫杆菌代谢物对病原菌的抑制作用及稳定性研究[A].程登发.植物保护21世纪展望[C].北京:中国科学技术出版社, 1998:689-692.
	YANG Xinfen, JIAN Heng, ZHANG Shuangao. The hibition and stability of Xenorhabdus nematophila on pathogenic bacteria[A].Cheng DF.The 21st Century Outlook for plant conservation[C].Beijing: Science and Technology Press of China, 1998:689-692.
[13]	Gaugler R., Kaya H.K.. Entomopathogenic Nematodes in Biological Control[J]. CRC Press, Boca Raton, FL, 1990.
[14]	张彩铃, 王志敏, 王思雨, 等. 昆虫病原线虫对暗黑鳃金龟的致病力[J/OL]. 中国生物防治学报:1-10[2023-02-14].
	Zhang Cailing, Wang Zhimin, Wang Siyu, et al. Virulence of Entomopathogenic Nematodes to Holotrichia parallela Larvae[J/OL]. Chinese Journal of Biological Control:1-10[2023-02-14].
[15]	余向阳, 王冬兰, 刘济宁, 等. 昆虫病原线虫的室内感染活性及其所受温湿度的影响[J]. 江苏农业学报, 2003,(1):13-17.
	YU Xiangyang, WANG Donglan, LIU Jining, et al. Infectivity of Steinernema carpocapsae Ohio to Some Insect Pests and Influence of Temperature and Humidity under Laboratory Condition[J]. Jiangsu Journal of Agriculture Sciences, 2003,(1)13-17.
[16]	Dunphy GB, Webster JM. Temperature effects on the growth and virulence of Steinernema feltiae strains and Heterorhabditis heliothidis[J]. Journal of Nematology, 1986, 18(2):270-272. PMID
[17]	White G F. A method for obtaining infective nematode larvae from cultures[J]. Science, 1927, 66(1709):302-303. PMID
[18]	Harlan D P, Dutky S R, Padgett G R, et al. Parasitism of neoaplactana dutkyi in white-fringed beetle larvae[J]. Journal of Nematology, 1971, 3(3):280-283. PMID
[19]	刘红兵, 刘奇志. 来自新疆的昆虫病原线虫的鉴定及生物学特性的研究[J]. 中国生物防治学报, 2016, 32(1):106-111. DOI
	LIU Hongbin, LIU Qizhi. Identification and Biological Characteristics of Entomopathogenic Nematode Isolated from Xinjiang[J]. Chinese Journal of Biological Control, 2016, 32(1):106-111. DOI
[20]	田成丽, 刘金文, 张金花, 等. 小杆线虫 Oscheius tipulae JL1的鉴定及生物学特性[J]. 中国生物防治学报, 2019, 35(6):900-907. DOI
	TIAN Chengli, LIU Jinwen, ZHANG Jinhua, et al. Identification and Biological Characteristics of Entomopathogenic Nematode JLSY 003[J]. Chinese Journal of Biological Control, 2019, 35(6):900-907. DOI
[21]	Glazer I, Liran N, Poinar G O, et al. Identification and biological activity of newly isolated Heterorhabditidae populations from Israel[J]. Fundamental of Applied Nematology, 1993, 16(5):467-472.
[22]	颜珣, 谷星慧, 韩日畴. 昆虫病原线虫防治草地贪夜蛾的研究进展[J]. 环境昆虫学报, 2019, 41(4):695-700.
	YAN Xun, GUO Xinghui, HAN Richou. Research advances in using entomopathogenic nematodes to control the fall armyworm, Spodoptera frugiperda[J]. Journal of Environmental Entomology, 2019, 41(4):695-700.
[23]	Mazurkiewicz A, Tumialis D, Jakubowska M. Biological control potential of native entomopathogenic nematodes(Steinernematidae and Heterorhabditidae)against Mamestra brassicae L.(Lepidoptera:Noctuidae)[J]. Agriculture, 2020, 10(9):1-9. DOI URL
[24]	Ramalingam Karthik Raja, Alagarsamy Arun, Mustapha Touray, et al. Antagonists and defense mechanisms of entomopathogenic nematodes and their mutualistic bacteria[J]. Biological Control, 2021, 152(prepublish).
[25]	钱秀娟. 甘肃省昆虫病原线虫资源及其对非生物胁迫的适应性研究[D]. 兰州: 甘肃农业大学, 2014.
	QIAN Xiujuan. Resources of entomopathogenic nematodes and their adaptability to abiotic stress in Gansu Province[D]. Lanzhou: Gansu Agricultural University, 2014.
[26]	刘奇志, 王玉柱. 斯氏线虫cB-2y品系对李实蜂的侵染试验[J] .昆虫天敌, 1993, 15(2):84-87.
	LIU Qizhi, WANG Yuzhu. Infectivity of entomopathogenic nematode Steinernema spp.CB-2y strain to sawfly hoplocampa spp[J]. Insect natural enemy, 1993, 15(2):84-87.
[27]	刘树森, 李克斌, 刘春琴, 等. 嗜菌异小杆线虫沧州品系对暗黑鳃金龟幼虫的致病力[J]. 植物保护, 2010, 36(5):96-100.
	LIU Shusen, LI Kebin, LIU Chunqin, et al. Virulence of Heterorhabditis bacteriophora strain Cangzhou to larvae of Holotrichia parallela[J]. PlantProtection, 2010, 36(5):96-100.
[28]	马娟, 郭笑笑, 李秀花, 等. 不同品系昆虫病原线虫抗逆性分析[J]. 中国生物防治学报, 2022, 38(1):188-195. DOI
	MA Juan, GUO Xiaoxiao, LI Xiuhua, et al. Tolerance of Different Entomopathogenic Nematodes to Environmental Stresses[J]. Chinese Journal of Biological Control, 2022, 38(1):188-195. DOI
[29]	张安邦, 南宫自艳, 宋萍, 等. 昆虫病原线虫对黄地老虎致病力的研究[J]. 环境昆虫学报, 2015, 37(3):591-597.
	ZHANG Anbang, NANGONG Ziyan, SONG Ping, et al. Efficacy of entomopathogenic nematodes against Agrotis segetum larvae[J]. Journal of Environmental Entomology, 2015, 37(3):591-597.
[30]	张丽娟, 刘奇志, 王玉柱, 等. 小杆线虫 Rhabditis (Oscheius) sp.生物学特性测定[J]. 安徽农业科学, 2010, 38(1):212-213.
	ZHANG Lijuan, LIU Qizhi, WANG Yuzhu, et al. Determination on Biological Characteristics of Nematode Rhabditis (Oscheius) sp.[J]. Journal of Anhui Agricultural Science, 2010, 38(1):212-213.
[31]	刘奇志, 张丽娟, 王玉柱, 等. 昆虫病原线虫(Steinernema longicaudum BPS品系)生物学测定[J]. 中国农学通报, 2009, 25(2):236-239.
	LIU Qizhi, ZHANG Lijuan, WANG Yuzhu, et al. Bio-Analysis on EPNs of Steinernema longicaudum BPS Strain[J]. Chinese Agricultural Science Bulletin, 2009, 25(2):236-239.
[32]	肖尧, 杨秀芬, 杨怀文. 致病杆菌和发光杆菌抗菌代谢产物研究进展[J]. 中国生物防治学报, 2011, 27(4):553-558.
	XIAO Rao, YANG Xiufen, YANG Huaiwen. Advances in Antimicrobial Metabolites of the Symbiotic Bacteria Associated with Entomopathogenic Nematodes[J]. Chinese Journal of Biologi-cal Control, 2011, 27(4):553-558.

三种昆虫病原线虫非生物胁迫耐受特性及其共生菌抑菌作用

Study on abiotic stress tolerance of three entomopathogenic nematodes and its bacteriostasis of symbiotic bacteria

在线阅读

PDF下载

可视化

摘要/Abstract

引用本文

使用本文

图/表 5

参考文献 32

相关文章 15

编辑推荐

Metrics

[1]	徐李娟, 陈勇, 王则玉, 王博, 艾尼江·尔斯满, 郭瑞, 李克梅, 宋素琴. 新疆拜城县马铃薯疮痂病病原菌的分离鉴定及生物学特性分析[J]. 新疆农业科学, 2023, 60(9): 2258-2265.
[2]	高宇洁, 詹发强, 陈澄, 包慧芳, 杨蓉, 王宁, 侯新强, 侯敏, 史应武, 龙宣杞. 黑曲霉拮抗菌Xenorhabdus bovienii445筛选、鉴定及发酵优化[J]. 新疆农业科学, 2022, 59(5): 1203-1215.
[3]	高勇刚, 高海峰, 吕卓, 徐李娟, 古丽尼沙·沙依木, 唐琦勇, 顾美英, 张丽娟, 朱锦泉, 宋素琴. 新疆北部沿天山主要种植区马铃薯病害分析[J]. 新疆农业科学, 2022, 59(1): 170-178.
[4]	吕卓, 高勇刚, 唐琦勇, 楚敏, 顾美英, 朱静, 程思琦, 马晓雪, 宋素琴, 王静. 奇台窖藏马铃薯病害病原鉴定[J]. 新疆农业科学, 2020, 57(5): 902-909.
[5]	詹发强, 侯敏, 杨蓉, 王宁, 包慧芳, 侯新强, 崔卫东, 龙宣杞. 1株新疆胡杨林昆虫病原线虫鉴定及生物学特性[J]. 新疆农业科学, 2020, 57(3): 536-544.
[6]	申慧, 魏冬梅, 段魏魏, 梁永增, 王咏星. 一株鱼类水霉病原拮抗菌JL50分离鉴定及其拮抗特性分析[J]. 新疆农业科学, 2019, 56(6): 1127-1135.
[7]	楚敏, 顾美英, 唐琦勇, 朱静, 郝秀英, 张志东. 达坂城蚕豆根腐病病原菌鉴定及生防菌拮抗效果分析[J]. 新疆农业科学, 2019, 56(10): 1904-1911.
[8]	俞燕,蔡梦杭,徐灿,黄家风. 不同碳氮比对棉花黄萎病菌主要生物学性状及致病力的影响[J]. 新疆农业科学, 2019, 56(1): 84-92.
[9]	蔡梦杭,徐灿,刘启,俞燕,黄家风. 北疆棉区棉花黄萎病菌培养性状及致病力分化[J]. 新疆农业科学, 2019, 56(1): 93-103.
[10]	牛春林,叶晓芸,艾尼,王振华,热依汗,姜国勇,高雁,宋素琴,楚敏. 克拉玛依果树腐烂病病原菌鉴定及其拮抗菌的筛选[J]. 新疆农业科学, 2018, 55(2): 304-311.
[11]	吴玉鹏, 赵晓梅. 不同产地库尔勒香梨萼端黑斑病病原菌的分离鉴定[J]. 新疆农业科学, 2018, 55(1): 116-122.
[12]	马盾;王奉斌;吴强;张燕红;袁杰;海里且木;赵志强;曹伟. 新疆水稻穗腐病病原菌种类鉴定及防控研究[J]. , 2017, 54(6): 1093-1098.
[13]	刘凡;李敏;顾爱星;李克梅;范钧星;玛依古丽·安那提. 薰衣草精油对几种植物病原菌的抑菌作用[J]. , 2017, 54(5): 938-944.
[14]	李先强;赵丽;刘俊峰;郭雪峰;江春雨;刘永宏. 氟苯尼考对三种猪呼吸道病原菌标准株的体外抗菌类型研究[J]. , 2017, 54(3): 557-563.
[15]	周扬;吴琼;刘梦丽;白剑宇;李进;雷斌;郭庆元. 南疆棉区棉花立枯病菌和红腐病菌种间及种内菌株间的致病力比较[J]. , 2017, 54(3): 489-496.