黄栌枯萎病菌拮抗细菌的分离与鉴定

doi:10.3969/j.issn.1000-2006.2015.06.004

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南京林业大学学报（自然科学版） ›› 2015, Vol. 58 ›› Issue (06): 17-23.doi: 10.3969/j.issn.1000-2006.2015.06.004

黄栌枯萎病菌拮抗细菌的分离与鉴定

孙妍,于地美,黄华毅,王永林,田呈明^*

省部共建森林培育与保护教育部重点实验室,北京林业大学林学院,北京 100083

出版日期:2015-11-30 发布日期:2015-11-30
基金资助:
收稿日期:2014-11-24 修回日期:2015-09-11
基金项目:中央高校基本科研业务费专项资金项目(xs201401)
第一作者:孙妍,本科生。*通信作者:田呈明,教授。E-mail: chengmt@bjfu.edu.cn。
引文格式:孙妍,于地美,黄华毅,等. 黄栌枯萎病菌拮抗细菌的分离与鉴定[J]. 南京林业大学学报:自然科学版,2015,39(6):17-23.

Isolation and identification of an antagonistic bacterium against smoke tree wilt fungus Verticillium dahliae

SUN Yan, YU Dimei, HUANG Huayi, WANG Yonglin, TIAN Chengming^*

Key Laboratory for Silviculture and Conservation of Ministry of Education, College of Forestry, Beijing Forestry University, Beijing 100083, China

Online:2015-11-30 Published:2015-11-30

摘要/Abstract

摘要： 为了获得能有效拮抗黄栌枯萎病的大丽轮枝菌(Verticillium dahliae)和尖孢镰刀菌(Fusarium oxysporum)的生防菌株,采用系列稀释分离法和平板对峙法从盐碱地土壤中筛选拮抗细菌,基于形态特征、生理生化特性和16S rDNA序列分析鉴定菌株,并测定其无菌滤液和挥发性气体的抑菌活性。筛选结果表明,菌株C-2-3-2是一株能有效抑制黄栌枯萎病菌和尖孢镰刀菌的生防菌株,经鉴定其为枯草芽孢杆菌(Bacillus subtilis)。该菌株的无菌滤液对黄栌枯萎病菌和尖孢镰刀菌均具有明显的体外抑菌作用,且能有效抑制黄栌枯萎病菌微菌核的形成。同时,其挥发性气体也能有效抑制黄栌枯萎病菌的生长。另外,研究还发现该菌株能产生铁载体和合成氨,并产生几丁质酶、纤维素酶、蛋白酶和β-1,3-葡聚糖酶。研究结果表明菌株C-2-3-2在黄栌枯萎病生物防治中具有较好的应用潜力和较高的研究价值。

Abstract: In order to isolate and characterize an antagonistic bacterial strain against Verticillium dahliae and Fusarium oxysporum, serial dilution and plate-confrontation methods were used for preliminary screening and selecting antagonistic bacteria from saline-alkali soil. Strain identification was based on morphological and physiological characteristics, and phylogenetic analysis of 16S rDNA sequence. The antagonistic effects of sterile culture filtrates and volatiles of antagonistic bacteria were determined. An antagonistic bacteria, named as C-2-3-2, was isolated and identified as Bacillus subtilis. The results indicated that sterile culture filtrate of the strain exhibited obvious in vitro inhibition against V. dahliae and F. oxysporum, and suppressed the production of microsclerotia of V. dahliae. The volatiles also effectively inhibited the mycelial growth of V. dahliae. In addition, the results indicated that C-2-3-2 could produce ammonia and siderophore, and also could produce cell wall degrading enzymes, such as chitinase, cellulose, protease and β-1,3-glucanase. It indicted that strain C-2-3-2 had high research value and potentials for biological control of Verticillium wilt of Cotinus coggygria.

中图分类号:

S763.1

孙妍,于地美,黄华毅,等. 黄栌枯萎病菌拮抗细菌的分离与鉴定[J]. 南京林业大学学报（自然科学版）, 2015, 58(06): 17-23.

SUN Yan, YU Dimei, HUANG Huayi, WANG Yonglin, TIAN Chengming. Isolation and identification of an antagonistic bacterium against smoke tree wilt fungus Verticillium dahliae[J].Journal of Nanjing Forestry University (Natural Science Edition), 2015, 58(06): 17-23.DOI: 10.3969/j.issn.1000-2006.2015.06.004.

参考文献

[1] 李俊文,张昆仑,马素双,等. 黄栌黄萎病病原菌的研究[J]. 林业科技通讯,1992(1): 21-25. Li J W, Zhang K L, Ma S S, et al. Research on the pathogens of Verticillium wilt fungus of smoke tree Cotinus coggygria[J]. Practical Forestry Technology, 1992(1):21-25.
[2] 雷增普. 北京地区黄栌黄萎病病原菌的研究[J]. 北京林业大学学报,1993,15(3): 88-93. Lei Z P. Study of Verticillium causing cotinus coggygria wilt in the Beijing area[J]. Journal of Beijing Forestry University, 1993,15(3):88-93.
[3] Wang Y, Xiao S, Xiong D, et al. Genetic transformation, infection process and qPCR quantification of Verticillium dahliae on smoke-tree Cotinus coggygria[J]. Australasian Plant Pathology, 2013, 42(1): 33-41.
[4] Klosterman S J, Atallah Z K, Vallad G E, et al. Diversity, pathogenicity, and management of Verticillium species[J]. Annual Review of Phytopathology. 2009, 47: 39-62.
[5] Agrios G. Plant Pathology[M]. Burlington: Elsevier Academic Press,1970.
[6] 田黎,王克荣,陆家云. 多菌灵、三环唑对大丽轮枝菌微菌核、黑色素形成及致病力的影响[J]. 植物病理学报,1998,28(3): 263-268. Tian L, Wang K R, Lu J Y. Effect of carbendazim and tricyclazole on microsclerotia and melanin formation of Verticillium dahliae[J]. Plant Protection, 1998, 28(3): 263-268.
[7] 王建美,才玉石,陶万强,等. 不同真菌菌株对黄栌的致病性研究[J]. 中国森林病虫,2008,27(3): 1-3. Wang J M, Cai Y S, Tao W Q, et al. Pathogenicity of different fungus strains on Cotinus coggygria[J]. Forest Pest and Disease, 2008, 27(3): 1-3.
[8] 王建美,田呈明,过颂新,等. 黄栌根内生真菌分离鉴定及拮抗真菌筛选[J]. 菌物研究,2008,6(1): 35-39. Wang J M, Tian C M, Guo S X, et al. Isolation and screening of endophytic antifungal fungi from Cotinus coggygria roots[J]. Journal of Fungal Research, 2008, 6(1): 35-39.
[9] 宋立洲,杜望光,李维维. 香山公园黄栌枯萎病防治技术研究[J]. 北京园林,2011,27(2): 51-56. Song L Z, Du W G, Li W W. Technology research of preventing and controlling toward Verticillium wilt disease in Xiangshan Park[J]. Journal of Beijing Garden, 2011, 27(2): 51-56.
[10] Huang H Y, Wu Z Q, Tian C M, et al. Identification and characterization of the endophytic bacterium Bacillus atrophaeus XW2, antagonistic towards Colletotrichum gloeosporioides[J]. Annals of Microbiology, 2015,65(3):1361-1371.
[11] Baysal Ö, Lai D, Xu H H, et al. A proteomic approach provides new insights into the control of soil-borne plant pathogens by Bacillus species[J]. PloS One, 2013, 8(1): e53182.
[12] 谢宗华,高健,王金宇,等. 一株水稻纹枯菌拮抗细菌的分离与鉴定[J]. 微生物学通报,2012,39(4): 477-485. Xie Z H, Gao J, Wang J Y, et al. Isolation and identification of an antagonistic bacterium against Rhizoctonia solani, the causing agent of rice sheath blight[J]. Microbiology China, 2012, 39(4): 477-485.
[13] 薛磊,王建涛,刘相春,等. 拮抗性链霉菌对大丽轮枝菌微菌核形成与萌发的影响[J]. 植物保护学报,2012,39(4): 289-296. Xue L, Wang J T, Liu X C, et al. Inhibition of antagonistic Streptomyces spp. on mircrosclerotia formation and germination of Verticillium dahlia[J]. Plant Protection, 2012, 39(4): 289-296.
[14] Zhang X Y, Li B Q, Wang Y G, et al. Lipopeptides, a novel protein, and volatile compounds contribute to the antifungal activity of the biocontrol agent Bacillus atrophaeus CAB-1[J]. Applied Microbiology and Biotechnology, 2013, 97(21):9525-9534.
[15] Nautiyal C S. An efficient microbiological growth medium for screening phosphate solubilizing microorganisms[J]. FEMS Microbiology Letters, 1999, 170(1): 265-270.
[16] Gordon S A, Weber R P. Colorimetric estimation of indoleacetic acid[J]. Plant Physiology, 1951, 26(1): 192-195.
[17] Jeffries P, Gianinazzi S, Perotto S, et al. The contribution of arbuscular mycorrhizal fungi in sustainable maintenance of plant health and soil fertility[J]. Biology and Fertility of Soils, 2003, 37(1): 1-16.
[18] De los Santos-Villalobos S, Bareera-Galicia G C, Miranda-Salcedo M A, et al. Burkholderia cepacia XXVI siderophore with biocontrol capacity against Colletotrichum gloeosporioides[J]. World Journal of Microbiology and Biotechnology, 2012, 28(8): 2615-2623.
[19] 高小宁,涂璇,黄丽丽,等. 产β-1,3-葡聚糖酶植物内生放线菌的筛选及抑菌活性研究[J]. 微生物学通报,2009,36(8): 1189-1194. Gao X N, Tu X, Huang L L, et al. The screen of plant endophytic actinomycetes producing β-1, 3-glucanase and antifungal activity of β-1,3-glucanase[J]. Microbiology China, 2009, 36(8): 1189-1194.
[20] Essghaier B, Fardeau M L, Cayol J L, et al. Biological control of grey mould in strawberry fruits by halophilic bacteria[J]. Journal of Applied Microbiology, 2009, 106(3): 833-846.
[21] Ren J H, Li H, Wang Y F, et al. Biocontrol potential of an endophytic Bacillus pumilus JK-SX001 against poplar canker[J]. Biological Control, 2013, 67(3): 421-430.
[22] 东秀株,蔡妙英. 常见细菌系统鉴定手册[M]. 北京: 科学出版社,2001. Dong X Z, Cai M Y. Identification methods of common bacteria[M]. Beijing: Science Press, 2001.
[23] 周德庆. 微生物学实验手册[M]. 上海:科学技术出版社,1986. Zhou D Q. Manual of microbiology laboratory[M]. Shanghai: Scientific and Technical Publishing House, 1986.
[24] 何红,邱思鑫,蔡学清,等. 辣椒内生细菌 BS-1和 BS-2在植物体内的定殖及鉴定[J]. 微生物学报, 2004,44(1): 13-18. He H, Qiu S Q, Cai X Q, et al. Colonization in plants and identification of endophytic bacteria BS-1 and BS-2 from Capsicum annuum[J]. Acta Microbiologica Sinica, 2004, 44(1): 13-18.
[25] Marchesi J R, Sato T, Weightman A J, et al. Design and evaluation of useful bacterium-specific PCR primers that amplify genes coding for bacterial 16S rRNA[J]. Applied and Environmental Microbiology, 1998, 64(2): 795-799.
[26] 葛瑾,颜容,宋立洲,等. 黄栌枯萎病的综合防治策略[J]. 中国城市林业,2007,5(3): 43-44. Ge J, Yan R, Song L Z, et al. Strategy for integrated control of Cotinus coggygria wilt[J]. Journal of Chinese Urban Forestry, 2007, 5(3): 43-44.
[27] 张进霞,袁洪水,王士英,等. 几种氨基酸铜对大丽轮枝菌微菌核形成的抑制作用[J]. 棉花学报,2006,18(1): 58-59. Zhang J X, Yuan H S, Wang S Y, et al. Inhibition of the cupric complexes of amino acid on microsclerotia formation of Verticillium dahliae[J]. Cotton Science, 2006, 18(1): 58-59.
[28] 田黎,王克荣,陆家云. 匐柄霉对大丽轮枝菌生长及微菌核形成的影响[J]. 中国生物防治,1998,14(1): 14-17. Tian L, Wang K R, Lu J Y. Inhibition of Stemphylium sp. on Verticillium dahliae mycelial growth and microsclerotium formation[J]. Chinese Journal of Biological Control, 1998, 14(1): 14-17.
[29] Debode J, de Maeyer K, Perneel M, et al. Biosurfactants are involved in the biological control of Verticillium microsclerotia by Pseudomonas spp.[J]. Journal of Applied Microbiology, 2007, 103(4): 1184-1196.
[30] Antonopoulos D F, Tjamos S E, Antoniou P P, et al. Effect of Paenibacillus alvei, strain K165, on the germination of Verticillium dahliae microsclerotia in planta[J]. Biological Control, 2008, 46(2): 166-170.
[31] 张艳群,来航线,韦小敏,等. 两株生防芽孢细菌筛选, 鉴定及拮抗研究[J]. 微生物学通报,2014,41(2): 281-289. Zhang Y Q, Lai H X, Wei X M, et al. Screening and identification and antagonism research of two biocontrol Bacillus strains[J]. Microbiology China, 2014, 41(2): 281-289.
[32] 黄海婵,裘娟萍. 枯草芽孢杆菌防治植物病害的研究进展[J]. 浙江农业科学,2005(3): 213-215. Huang H C, Qiu J P.Advances in research on control plant disease with Bacillus subtilis[J]. Journal of Zhejiang Agricultural Sciences, 2005,(3): 213-215.
[33] 赵新林,赵思峰. 枯草芽孢杆菌对植物病害生物防治的作用机理[J]. 湖北农业科学,2011,50(15): 3025-3028. Zhao X L, Zhao S F. Research advance in controlling plant diseases by Bacillus subtilis[J]. Hubei Agricultural Sciences, 2011, 50(15): 3025-3028.

黄栌枯萎病菌拮抗细菌的分离与鉴定

Isolation and identification of an antagonistic bacterium against smoke tree wilt fungus Verticillium dahliae

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