甘薯茎腐病症状及其病原鉴定
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摘要
甘薯茎腐病是危害甘薯的一种严重病害,甘薯茎、叶柄、叶片和块根均可被害。该病典型症状是茎基部发黑和变软腐烂,叶发黄,茎和块根维管束黑褐色,引起块根腐烂、有臭味。通过对甘薯茎腐病典型症状样本的采集、病原菌的分离和纯化以及致病性测定,明确该病害是一种细菌病害。通过对甘薯茎腐病菌的菌体形态和培养特性观察发现,病原菌是革兰氏阴性细菌,菌体短杆状,大小约为2.36μm×0.4μm,周生鞭毛,可在烟草上激发过敏性反应(HR)。Biolog测定、脂肪酸分析、16S rDNA序列分析、MALDI-TOF质谱鉴定和8个看家基因(dnaX、rplB、fusA、gapA、gyrA、purA、recA和rpoS)的序列系统发育分析,发现该病原菌与达旦提狄克氏菌Dickeya dadantii高度一致。这些结果说明,浙江省发生的小番薯病害是甘薯茎腐病,病原为D.dadantii。
Bacterial stem and root rot of sweet potato is a severe disease which can infect its stems,petioles,leaves and tuberous roots. The typical symptoms of the disease include black and rot basal stems,yellow leaves,dark brown vascular stems and smelly tuberous roots. On the basis of typical symptoms,pathogen isolation,purification and pathogenicity test,it was confirmed that it was a bacterial disease. Through the observation of the bacterial morphological and cultural characteristics,it was found that the pathogen is a short rod-shaped,gram-negative bacterium with peritrichous flagella,2.36 μm × 0.4 μm in size,and can elicit the hypersensitive response( HR) in tobacco. According to Biolog test,analysis of fatty acid methyl esters and 16 S rDNA sequence,MALDI-TOF mass spectrometry test and phylogenetic sequence analysis of 8 house-keeping genes,dnaX,rplB,fusA,gapA,gyrA,purA,recA and rpoS,it was found that the pathogen shares high similarity with Dickeya dadantii. These results indicated that the stem rot disease of sweet potato in Zhejiang Province was caused by D. dadantii.
Bacterial stem and root rot of sweet potato is a severe disease which can infect its stems,petioles,leaves and tuberous roots. The typical symptoms of the disease include black and rot basal stems,yellow leaves,dark brown vascular stems and smelly tuberous roots. On the basis of typical symptoms,pathogen isolation,purification and pathogenicity test,it was confirmed that it was a bacterial disease. Through the observation of the bacterial morphological and cultural characteristics,it was found that the pathogen is a short rod-shaped,gram-negative bacterium with peritrichous flagella,2.36 μm × 0.4 μm in size,and can elicit the hypersensitive response( HR) in tobacco. According to Biolog test,analysis of fatty acid methyl esters and 16 S rDNA sequence,MALDI-TOF mass spectrometry test and phylogenetic sequence analysis of 8 house-keeping genes,dnaX,rplB,fusA,gapA,gyrA,purA,recA and rpoS,it was found that the pathogen shares high similarity with Dickeya dadantii. These results indicated that the stem rot disease of sweet potato in Zhejiang Province was caused by D. dadantii.
引文
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[8]Samson R,Legendre J B,Christen R,et al.Transfer of Pectobacterium chrysanthemi(Burkholder et al.1953)Brenner et al.1973 and Brenneria paradisiaca to the genus Dickeya gen.nov.as Dickeya chrysanthemi comb.nov.and Dickeya paradisiaca comb.nov.and delineation of four novel species,Dickeya dadantii sp.nov.,Dickeya dianthicola sp.nov.,Dickeya diefenbachiae sp.nov.and Dickeya zeae sp.nov.[J].International Journal of Systematic and Evolutionary Microbiology,2005,55(4):1415-1427.
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[11]Tian Y L,Zhao Y Q,Yuan X L,et al.Dickeya fangzhongdai sp.nov.,a plant-pathogenic bacterium isolated from pear tree(Pyrus pyrifolia)[J].International Journal of Systematic and Evolutionary Microbiology,2016,66(8):2831-2835.
[12]Brady C L,Cleenwerck I,Denman S,et al.Proposal to reclassify Brenneria quercina(Hildebrand and Schroth 1967)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.iberica subsp.nov.and Lonsdalea quercina subsp.britannica subsp.nov.,emendation of the description of the genus Brenneria,reclassification of Dickeya dieffenbachiae as Dickeya dadantii subsp.dieffenbachiae comb.nov.,and emendation of the description of Dickeya dadantii[J].International Journal of Systematic and Evolutionary Microbiology,2012,62(Pt7):1592-1602.
[13]John M,Stephane G,Shimpei M,et al.Top 10 plant pathogenic bacteria in molecular plant pathology[J].Molecular Plant Pathology,2012,13(6):614-629.
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[16]Waleron M,Waleron K,Podhajska A J,et al.Genotyping of bacteria belonging to the former Erwinia genus by PCR-RFLP analysis of a recA gene fragment[J].Microbiology,2002,148(2):583-595.
[17]Waleron M,Waleron K,Lojkowska E.Genotypic characterisation of the Erwinia genus by PCR-RFLP analysis of rpoS gene[J].Plant Protection Science,2002,38(S2):288-290.
[18]Sawiak M,van Beckhoven J R C M,Speksnijder A GC L,et al.Biochemical and genetical analysis reveal a new clade of biovar 3 Dickeya spp.strains isolated from potato in Europe[J].European Journal of Plant Pathology,2009,125(2):245-261.
[19]Liu P,Huang G M,Liu Y,et al.Molecular detection of Erwinia chrysanthemi(in Chinese)[J].Acta Phytopathologica Sinica(植物病理学报),2007,37(6):584-587.
[20]Smid E J,Jansen A J,Gorris L M.Detection of Erwinia carotovora subsp.atroseptica and Erwinia chrysanthemi in potato tubers using polymerase chain reaction[J].Plant Pathology,1995,44(6):1058-1069.
[21]Huang L F,Fang B P,Luo Z X,et a1.First report of bacterial stem and root rot of sweet potato caused by a Dickeya sp.(Erwinia chrysanthemi)in China[J].Plant Disease,2010,94(12):1503.
[22]Glorimar M,Kevin L S,Daniel M J,et al.Phylogeny and classification of Dickeya based on multilocus sequence analysis[J].International Journal of Systematic and Evolutionary Microbiology,2013,63(Pt9):3524-3539.
[2]Schaad N W,Brenner D A.Bacterial wilt and root rot of sweet potato caused by Erwinia chrysanthemi[J].Phytopathology,1977,67(3):302-308.
[3]Tamura I,Azegami K,Miura T,et al.Bacterial stem and root rot of sweet potato caused by Erwinia chrysanthemi[J].Annals Phytopathological Society of Japan,1998,64(4):376.
[4]Varela G,Hernández Y,Trujillo G.Bacterial stem rot of sweet potato(Ipomoea batatas(L.)Lam.)caused by Erwinia chrysanthemi in Maracay,Aragua state,Venezuela[J].Revista De La Facultad De Agronomía Universidad Central De Venezuela,1999,25(1):29-39.
[5]Huang L F,Luo Z X,Fang B P,et al.A new bacterial stem and root rot disease of sweet potato in Guangdong,China(in Chinese)[J].Acta Phytopathologica Sinica(植物病理学报),2011,41(1):18-23.
[6]Qin S Y,Huang L F,Ge C B,et al.Molecular identification of a new bacterial stem rot disease of sweet potato in Henan(in Chinese)[J].Crops(作物杂志),2013,(6):52-55.
[7]Gao B,Wang R Y,Ma J,et al.Preliminary study on bacterial stem and root rot disease of sweet potato in Hebei Province(in Chinese)[J].Plant Protection(植物保护),2015,41(3):119-122.
[8]Samson R,Legendre J B,Christen R,et al.Transfer of Pectobacterium chrysanthemi(Burkholder et al.1953)Brenner et al.1973 and Brenneria paradisiaca to the genus Dickeya gen.nov.as Dickeya chrysanthemi comb.nov.and Dickeya paradisiaca comb.nov.and delineation of four novel species,Dickeya dadantii sp.nov.,Dickeya dianthicola sp.nov.,Dickeya diefenbachiae sp.nov.and Dickeya zeae sp.nov.[J].International Journal of Systematic and Evolutionary Microbiology,2005,55(4):1415-1427.
[9]Parkinson N,DeVos P,Pirhonen M.et al.Dickeya aquatica sp.nov.,isolated from waterways[J].International Journal of Systematic and Evolutionary Microbiology,2014,64(7):2264-2266.
[10]Van der Wolf J M,Nijhuis E H,Kowalewska,et al.Dickeya solani sp.nov.,a pectinolytic plant-pathogenic bacterium isolated from potato(Solanum tuberosum)[J].International Journal of Systematic and Evolutionary Microbiology,2014,64(Pt3):768-774.
[11]Tian Y L,Zhao Y Q,Yuan X L,et al.Dickeya fangzhongdai sp.nov.,a plant-pathogenic bacterium isolated from pear tree(Pyrus pyrifolia)[J].International Journal of Systematic and Evolutionary Microbiology,2016,66(8):2831-2835.
[12]Brady C L,Cleenwerck I,Denman S,et al.Proposal to reclassify Brenneria quercina(Hildebrand and Schroth 1967)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.iberica subsp.nov.and Lonsdalea quercina subsp.britannica subsp.nov.,emendation of the description of the genus Brenneria,reclassification of Dickeya dieffenbachiae as Dickeya dadantii subsp.dieffenbachiae comb.nov.,and emendation of the description of Dickeya dadantii[J].International Journal of Systematic and Evolutionary Microbiology,2012,62(Pt7):1592-1602.
[13]John M,Stephane G,Shimpei M,et al.Top 10 plant pathogenic bacteria in molecular plant pathology[J].Molecular Plant Pathology,2012,13(6):614-629.
[14]Schaad N W,Jones J B,Chun W.Laboratory guide for identification of plant pathogenic bacteria[M].3rd Edition.Minn:American Phytopathological Socity,2001:5-6.
[15]Weisburg W G,Barns S M,Pelletier D A,et al.16Sribosomal DNA amplification for phylogenetic study[J].Journal of Bacteriology,1991,173(2):697-703.
[16]Waleron M,Waleron K,Podhajska A J,et al.Genotyping of bacteria belonging to the former Erwinia genus by PCR-RFLP analysis of a recA gene fragment[J].Microbiology,2002,148(2):583-595.
[17]Waleron M,Waleron K,Lojkowska E.Genotypic characterisation of the Erwinia genus by PCR-RFLP analysis of rpoS gene[J].Plant Protection Science,2002,38(S2):288-290.
[18]Sawiak M,van Beckhoven J R C M,Speksnijder A GC L,et al.Biochemical and genetical analysis reveal a new clade of biovar 3 Dickeya spp.strains isolated from potato in Europe[J].European Journal of Plant Pathology,2009,125(2):245-261.
[19]Liu P,Huang G M,Liu Y,et al.Molecular detection of Erwinia chrysanthemi(in Chinese)[J].Acta Phytopathologica Sinica(植物病理学报),2007,37(6):584-587.
[20]Smid E J,Jansen A J,Gorris L M.Detection of Erwinia carotovora subsp.atroseptica and Erwinia chrysanthemi in potato tubers using polymerase chain reaction[J].Plant Pathology,1995,44(6):1058-1069.
[21]Huang L F,Fang B P,Luo Z X,et a1.First report of bacterial stem and root rot of sweet potato caused by a Dickeya sp.(Erwinia chrysanthemi)in China[J].Plant Disease,2010,94(12):1503.
[22]Glorimar M,Kevin L S,Daniel M J,et al.Phylogeny and classification of Dickeya based on multilocus sequence analysis[J].International Journal of Systematic and Evolutionary Microbiology,2013,63(Pt9):3524-3539.