白菜黑腐病拮抗菌Lysobacter enzymogenes CX03的分离鉴定及生防效果研究

doi:10.16409/j.cnki.2095-039x.2021.06.015

摘要/Abstract

摘要： 从白菜根际土壤中分离到一株对白菜黑腐病菌具有显著拮抗效果的生防细菌CX03。根据生物学特征、生理生化分析、Biolog测定及多基因系统发育分析，鉴定菌株CX03为产酶溶杆菌Lysobacter enzymogenes。酶学试验结果表明，菌株CX03在代谢过程中产生纤维素酶、蛋白酶和几丁质酶。通过抑菌谱分析，证明菌株CX03具有广谱拮抗作用，对野油菜黄单胞野油菜致病变种、密执安棒杆菌密执安致病变种等5种病原细菌和尖孢镰孢菌、辣椒疫霉菌等4种病原真菌具有显著的拮抗效果。通过盆栽试验结果显示，菌株CX03处理组的白菜黑腐病症状明显减轻，防治效果达到90.4%，显著高于春雷霉素的防治效果84.1%。综上所述，本文首次报道了一株对植物病原细菌具有抗菌活性的产酶溶杆菌生防菌株CX03，其生防性状优良，在植物病害生物防治中具有一定的应用潜力，为国内生物防治研究提供了新的微生物资源。

关键词: 产酶溶杆菌, 黑腐病, 生物防治, 防效

Abstract: The biocontrol agent CX03 with strong antagonism to bacterial black rot of cabbage was isolated from the rhizosphere soil of pepper. According to biological characteristics, 16S rDNA sequence analysis, Biolog determination and physiological and biochemical analysis, the strain CX03 was identified as Lysobacter enzymogenes. The results of the enzymatic test showed that the strain CX03 produced cellulase, protease and chitinase in the metabolic process. The analysis of the antibacterial spectrum showed that the strain CX03 has a broad-spectrum antagonistic effect against five pathogenic bacteria including Xanthomonas campestris pv. campestris; Clavibacter michiganensis subsp. michiganensis, and four pathogenic fungi including Fusarium oxysporum and Phytophthora capsici. Treatment with strain CX03 significantly reduced the incidence of bacterial black rot of cabbage, and enhanced the control effect up to 90.4%, which was significantly higher than the control effect of kasugamycin. This study is the first report in China that L. enzymogenes has antibacterial activity against phytopathogenic bacteria, which provides new microbial resources for biological control research.

Key words: Lysobacter enzymogenes, black rot, biological control, control effect

中图分类号:

S476

张子玉, 谢学文, 石延霞, 柴阿丽, 李磊, 李宝聚. 白菜黑腐病拮抗菌Lysobacter enzymogenes CX03的分离鉴定及生防效果研究[J]. 中国生物防治学报, 2021, 37(6): 1221-1230.

ZHANG Ziyu, XIE Xuewen, SHI Yanxia, CHAI Ali, LI Lei, LI Baoju. Isolation, Identification and Biocontrol Effect of Lysobacter enzymogenes CX03[J]. Chinese Journal of Biological Control, 2021, 37(6): 1221-1230.

参考文献

[1] 李明远.十字花科蔬菜黑腐病的发生与防治[J].当代蔬菜, 2004(11):36-37.
[2] 喻子牛,邵宗泽,孙明.中国微生物基因组研究[J].遗传, 2012(11):1408-1408.
[3] 彭炜.植物细菌性病害和病原细菌分类研究进展[C]//公共植保与绿色防控, 2010.
[4] 姜英华,胡白石,刘凤权.植物土传病原菌拮抗细菌的筛选与鉴定[J].中国生物防治学报, 2005, 21(4):260-264.
[5] Lou L, Chen H, Cerny R L, et al. Unusual activities of the thioesterase domain for the biosynthesis of the polycyclic tetramate macrolactam HSAF in Lysobacter enzymogenes C3[J]. Biochemistry, 2012, 51(1):4.
[6] 姬广海,吴亚鹏,白学慧,等.抗生素溶杆菌对魔芋软腐病和根际微生物多样性的影响[J].江西农业大学学报, 2009(3):125-129, 170.
[7] Puopolo G, Palmieri M C, Giovannini O, et al. Impact of temperature on the survival and the biocontrol efficacy of Lysobacter capsici AZ78 against Phytophthora infestans[J]. Biocontrol, 2015, 60(5):681-689.
[8] 钱国良.产酶溶杆菌生防相关基因的克隆、功能分析及工程菌构建[D].南京:南京农业大学, 2009.
[9] Folman L B, Postma J. Characterization of Lysobacter enzymogenes (Christensen and Cook 1978) strain 3.1T8, a powerful antagonist of fungal diseases of cucumber[J]. Microbiology Reviews, 2003, 158(2):107-115.
[10] Chen J, Moore W H, Yuen G Y, et al. Influence of Lysobacter enzymogenes strain C3 on nematodes[J]. Journal of Nematology, 2006, 38(2):233-239.
[11] Kobayashi D Y, Reedy R M, Palumbo J D, et al. A clp gene homologue belonging to the crp gene family globally regulates lytic enzyme production, antimicrobial activity, and biological control activity expressed by Lysobacter enzymogenes strain C3[J]. Applied and Environmental Microbiology, 2005, 71(1):261-269.
[12] Kobayashi D Y, El-Barrad E H. Selection of bacterial antagonists using enrichment cultures for the control of summer patch disease in kentucky bluegrass[J]. Current Microbiology, 1996, 32(2):106-110.
[13] Liu Y, Qiao J, Liu Y, et al. Characterization of Lysobacter capsici strain NF87-2 and its biocontrol activities against phytopathogens[J]. European Journal of Plant Pathology, 2019, 155(3):859-869.
[14] 刘朝霞.辣椒溶杆菌X2-3抗菌物质的初步分离及其特性分析[D].泰安:山东农业大学, 2016.
[15] 赵杨扬.抗生素溶杆菌中吩嗪类物质的鉴定及其生物合成机制研究[D].南京:南京农业大学, 2016.
[16] Qian G L, Hu B S, Jiang Y H, et al. Identification and characterization of Lysobacter enzymogenes as a biological control agent against some fungal pathogens[J]. Agricultural Sciences in China, 2009, 8(1):68-75.
[17] 程丽娟,薛泉宏.微生物学实验技术教程[M].北京:科学出版社, 2012.
[18] Chatterjee A, Cui Y, Liu Y, et al. Inactivation of rsmA leads to overproduction of extracellular pectinases, cellulases, and proteases in Erwinia carotovora subsp. carotovora in the absence of the starvation/cell density-sensing signal, N-(3-oxohexanoyl)-L-homoserine lactone[J]. Applied and Environmental Microbiology, 1995, 61(5):1959-1967.
[19] Weinert N, Meincke R, Gottwald C, et al. Bacterial diversity on the surface of potato tubers in soil and the influence of the plant genotype[J]. FEMS Microbiology Ecology, 2010, 74(1):114-123.
[20] Stonier T. Agrobacterium tumefaciens conn.II. production of an antibiotic substance[J]. Journal of Bacteriology, 1960, 79(6):889-898.
[21] Reid J D, Ogrydziak D M. Chitinase-overproducing mutant of Serratia marcescens[J]. Applied and Environmental Microbiology, 1981, 41(3):664-669.
[22] 东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京:科学出版社, 2001.
[23] Dunbar J, Ticknor L O, Kuske C R. Assessment of microbial diversity in four south western United States soils by 16S rRNA geneterminal restriction fragment analysis[J]. Applied and Environmental Microbiology, 2000, 66(7):2943-2950.
[24] 付业勤,蔡吉苗,刘先宝,黄贵修.香蕉内生细菌分离、活性评价及数量分布[J].热带作物学报, 2007(4):78-83.
[25] 康锋,刘明昊.不同处理防治白菜黑腐病效果试验[J].现代农村科技, 2016,(7):46.
[26] 谢朝生,徐小明,殷玉明,等. 9种农药防治柑桔溃疡病田间药效评价报告[J].现代园艺, 2013(9):8-9.
[27] 姬广海.溶杆菌属及其在植物病害防治中的研究进展[J].云南农业大学学报(自然科学版), 2011, 26(1):124-130.
[28] Christensen P, Cook F D. Lysobacter, a new genus of non-fruiting, gliding bacteria with s high base ratio[J]. International Journal of Systematic and Bacteriology, 1978, 28(3):367-393.
[29] Yu F G, Rivera K Z, Zhu X C, et al. Structure and biosynthesis of heat-stable antifungal factor (HSAF), a broad-spectrum antimycotic with a novel mode of action[J]. Antimierobial Agents and Chemotherpy, 2007, 51(1):67-72.
[30] Zhao Y, Qian G, Ye Y, et al. Heterocyclic aromatic N-oxidation in the biosynthesis of phenazine antibiotics from Lysobacter antibioticus[J]. Organic Letters, 2016, 18(10):2495-2498.
[31] Wei L F, Zhou L H, Ji G H, et al. Control of rice bacterial leaf blight by antibacterial substances from Lysobacter antibioticus strain 13-1(in Chinese)[J]. Microbiology China, 2014, 41(2):274-280.
[32] Harad K I, Suzuki M, Kato A, et al. Separation of WAP-8294A components, a novel anti-methicillin-resistant Staphylococcus aureus antibiotic, using high-speed counter-current chromatography[J]. Journal of Chromatography A, 2001, 932(1):75-81.
[33] Hashizume H, Hirosawa S, Sawa R, et al. Tripropeptins, novel antimicrobial agents produced by Lysobacter sp.[J]. Cheminform, 2010, 35(1):52-58.