杨树湿心材致病细菌及其拮抗细菌的鉴定
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摘要
为分离并鉴定杨树湿心材病的主要致病细菌与其拮抗细菌,从不同杨树无性系发病组织中分离、纯化出优势菌株,分别回接实生苗进行致病性测定与病原菌鉴别;并进行形态学、生理生化特征、脂肪酸、16SrRNA基因序列以及多基因序列分析。结果表明:分离纯化得到了10株优势菌株,回接实验发现12-1菌株能导致杨树出现湿心材症状,鉴定其为杨柳欧文氏菌;从杨树心材中分离出了18个内生菌,通过拮抗菌筛选,发现J-4能有效抑制12-1菌株对杨树造成的湿心材现象,鉴定为类芽孢杆菌。因此,杨柳欧文氏菌是杨树湿心材病的主要致病菌,类芽孢杆菌是其拮抗菌。
In order to separate and purify the main pathogen bacteria to wet-heartwood disease in Populus and the correspondent antagonist bacteria,dominant strains were separated and purified from pathogenic tissues of different Populus varieties,then tested the pathogenicities by inoculation to sterile Populus seedlings. The pathogen and correspondent antagonist bacteria were classified and identified by morphology,physiological-biochemical characteristics,fatty acid composition analysis,16 SrRNA sequence and MLSA. The results show that 10 dominant strains are separated and purified,and pathogen 12-1 caused evident symptom after inoculation to sterile poplar seedlings,and it is classified and identified as Brenneria salicis; Eighteen endogenous strains are isolated from the stem of Populus trees,of which J-4 isolate is selected and verified as antagonist bacteria of the pathogen bacteria strain 12-1 by the confrontation culture and the inoculation in Populus sterile seedlings,and it is preliminary determined to be Paenibacillus taiwanensis. In conclusion,Brenneria salicis is the main pathogen to wet-heartwood disease in Populus,and Paenibacillus taiwanensis ias the correspondent antagonist bacteria.
In order to separate and purify the main pathogen bacteria to wet-heartwood disease in Populus and the correspondent antagonist bacteria,dominant strains were separated and purified from pathogenic tissues of different Populus varieties,then tested the pathogenicities by inoculation to sterile Populus seedlings. The pathogen and correspondent antagonist bacteria were classified and identified by morphology,physiological-biochemical characteristics,fatty acid composition analysis,16 SrRNA sequence and MLSA. The results show that 10 dominant strains are separated and purified,and pathogen 12-1 caused evident symptom after inoculation to sterile poplar seedlings,and it is classified and identified as Brenneria salicis; Eighteen endogenous strains are isolated from the stem of Populus trees,of which J-4 isolate is selected and verified as antagonist bacteria of the pathogen bacteria strain 12-1 by the confrontation culture and the inoculation in Populus sterile seedlings,and it is preliminary determined to be Paenibacillus taiwanensis. In conclusion,Brenneria salicis is the main pathogen to wet-heartwood disease in Populus,and Paenibacillus taiwanensis ias the correspondent antagonist bacteria.
引文
[1]徐红梅,胡兴宜,杜克兵,等.湖北省杨树湿心材的初步研究[J].湖北林业科技,2014,43(1):4-7.
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[3]刘杏娥,王小青,任海青,等.栽植密度对小黑杨边材、湿心材生长量的影响[J].林业科学研究,2007,20(4):490-494.
[4] Wang X,Jiang Z,Ren H. Distribution of wet heartwood in stems of Populus xiaohei from a spacing trial[J].Scandinavian Journal of Forest Research,2008,23(1):38-45.
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[8]赵桂华,赵楠.铁皮石斛叶斑病新病原:黑线炭疽菌的分离鉴定及致病性测定[J].西部林业科学,2016,45(3):20-25.
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[10]郑华英,戴雨生,解春霞,等.杨树红心病的形成原因[J].南京林业大学学报(自然科学版),2006,30(2):82-84.
[11]龙亚芹,段春芳,刘杰,等.小粒咖啡褐斑病病原菌鉴定及田间抗病性研究[J].西南林业大学学报,2017,37(2):152-157.
[12] 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.
[13]任欣正.植物病原细菌的分类和鉴定[M].北京:中国农业出版社,1999.
[14]李永.健康与染溃疡病107杨、中林46杨树皮真菌和细菌群落研究[D].北京:北京林业大学,2013.
[15] Maes M,Huvenne H,Messens E. Brenneria salicis,the bacterium causing watermark disease in willow,resides as an endophyte in wood[J]. Environmental Microbiology,2009,11(6):1453-1462.
[16] Huvenne H,Messens E,Maes M. Willow wood sap promotes the density-dependent pathogenesis of Brenneria salicis[J]. Environmental Microbiology,2009,11(6):1463-1472.
[17]池淼.杨属植物茎干内生菌多样性及拮抗细菌筛选[D].北京:中国林业科学研究院,2014.
[18] Kavino M,Harish S,Kumar N,et al. Rhizosphere and endophytic bacteria for induction of systemic resistance of banana plantlets against bunchy top virus[J]. Soil Biology&Biochemistry,2007,39(5):1087-1098.
[19] Brooks D S,Gonzalez C F,Appel D N,et al. Evaluation of endophytic bacteria as potential biological control agents for oak wilt[J]. Biological Control,1994,4(4):373-381.
[20]胥丽娜.杨树内生细菌的分离、多样性分析及解淀粉芽孢杆菌PEBA20的抑菌活性研究[D].泰安:山东农业大学,2009.
[21] Lelie D V D,Barac T,Taghavi S,et al. Response to Newman:New uses of endophytic bacteria to improve phytoremediation[J]. Trends in Biotechnology,2005,23(1):8-9.
[22] Taghavi S. Poplar and its bacterial endophytes:coexistence and harmony[J]. Critical Reviews in Plant Sciences,2009,28(5):346-358.
[2]祖勃荪.国外对杨树湿心材的研究[J].林业科学,2000,36(5):85-91.
[3]刘杏娥,王小青,任海青,等.栽植密度对小黑杨边材、湿心材生长量的影响[J].林业科学研究,2007,20(4):490-494.
[4] Wang X,Jiang Z,Ren H. Distribution of wet heartwood in stems of Populus xiaohei from a spacing trial[J].Scandinavian Journal of Forest Research,2008,23(1):38-45.
[5]晁龙军,曾大鹏,孙福在,等.引起杨树湿心材的一种病原真菌:Fusarium proliferatum(Matsushima)Nirenberg[J].林业科学,1998,34(5):69-73.
[6]赵桂华,孟庆庭,席刚俊.杨树红心病的研究[J].福建林学院学报,2010,30(3):207-211.
[7] Brady C L,Cleenwerck I,Denman S,et al. Proposal to reclassify Brenneria quercina(Hildebrand and Schroth1967)Hauben et al. 1999 into a new genus,Lonsdalea gen. nov. as Lonsdalea quercina comb. nov. descriptions of Lonsdalea quercina subsp. quercina comb. nov. Lonsdalea quercina subsp[J]. International Journal of Systematic&Evolutionary Microbiology,2011,62(7):1592.
[8]赵桂华,赵楠.铁皮石斛叶斑病新病原:黑线炭疽菌的分离鉴定及致病性测定[J].西部林业科学,2016,45(3):20-25.
[9]宋春草,王奎,梅莉,等.杨树湿心材病致病细菌的分离、鉴定和侵染途径[J].华中农业大学学报,2017,36(2):15-21.
[10]郑华英,戴雨生,解春霞,等.杨树红心病的形成原因[J].南京林业大学学报(自然科学版),2006,30(2):82-84.
[11]龙亚芹,段春芳,刘杰,等.小粒咖啡褐斑病病原菌鉴定及田间抗病性研究[J].西南林业大学学报,2017,37(2):152-157.
[12] 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.
[13]任欣正.植物病原细菌的分类和鉴定[M].北京:中国农业出版社,1999.
[14]李永.健康与染溃疡病107杨、中林46杨树皮真菌和细菌群落研究[D].北京:北京林业大学,2013.
[15] Maes M,Huvenne H,Messens E. Brenneria salicis,the bacterium causing watermark disease in willow,resides as an endophyte in wood[J]. Environmental Microbiology,2009,11(6):1453-1462.
[16] Huvenne H,Messens E,Maes M. Willow wood sap promotes the density-dependent pathogenesis of Brenneria salicis[J]. Environmental Microbiology,2009,11(6):1463-1472.
[17]池淼.杨属植物茎干内生菌多样性及拮抗细菌筛选[D].北京:中国林业科学研究院,2014.
[18] Kavino M,Harish S,Kumar N,et al. Rhizosphere and endophytic bacteria for induction of systemic resistance of banana plantlets against bunchy top virus[J]. Soil Biology&Biochemistry,2007,39(5):1087-1098.
[19] Brooks D S,Gonzalez C F,Appel D N,et al. Evaluation of endophytic bacteria as potential biological control agents for oak wilt[J]. Biological Control,1994,4(4):373-381.
[20]胥丽娜.杨树内生细菌的分离、多样性分析及解淀粉芽孢杆菌PEBA20的抑菌活性研究[D].泰安:山东农业大学,2009.
[21] Lelie D V D,Barac T,Taghavi S,et al. Response to Newman:New uses of endophytic bacteria to improve phytoremediation[J]. Trends in Biotechnology,2005,23(1):8-9.
[22] Taghavi S. Poplar and its bacterial endophytes:coexistence and harmony[J]. Critical Reviews in Plant Sciences,2009,28(5):346-358.