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http://dx.doi.org/10.5423/RPD.2003.9.3.116

Nested PCR Detection of Pseudomonas syringae pv. actinidiae, the Causal Bacterium  

Jung, Jae-Sung (Department of Biology, Sunchon National University)
Han, Hyo-Shim (Department of Biology, Sunchon National University)
Jo, Youn-Seob (Subtropical Fruits Experiment Station)
Koh, Young-Jin (Department of Applied Biology and Sunchon National University)
Publication Information
Research in Plant Disease / v.9, no.3, 2003 , pp. 116-120 More about this Journal
Abstract
A PCR method that combines biological and enzymatic amplification of PCR targets was developed for the detection of Pseudomonas syringae pv. actinidiae on kiwifruit leaves. A nested PCR was performed with primers designes from the coding sequence of the cfl gene, which is involved in production of the phytotoxin coronatine. The first and second primer sets efficiently amplified expected 665 and 310-bp fragments, respectively. With two successive amplifications, as few as 20 CFU/ml of P. syringae pv. actinidiae could be detected on ethidium bromide-stained agarose gel. Leaf samples were collected from 4 kiwifruit trees showing yellow halo spots on leaves and incubated in pepton-sucrose broth for 12 h at $16^{\circ}$C before PCR amplification. Positive detection was obtained with one sample, which was proved as a diseased plant in the next spring.
Keywords
cfl gene; kiwifruit canker; nested PCR; Pseudomonas syringae pv. actinidiae;
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1 Scortichini, M. 1994. Occurrence of Pseudomonas syringae pv. actinidiae on kiwifruit in Italy. Plant Pathology 43: 1035-1038   DOI   ScienceOn
2 Tamura, K., Thkikawa, Y., Tsuyumu, S. and Goto, M. 1989. Characterization of the toxin produced by Pseudomonas syringae pv. actinidiae, the causal bacterium of kiwifruit canker Ann. Phytopathol. Soc. Japan 55: 512
3 Clerc, A., Manceau, A. and Nesme, X. 1998. Comparison of randomly amplified polymorphic DNA with amplified fragment length polymorphism to assess genetic diversity and genetic relatedness within genospecies III of Pseudomonas syringae. Appl. Environ. Microbiol. 64: 1180-1187
4 고영진, 차병진, 정희정, 이동현. 1994. 참다래 궤양병의 격발 및 확산. 한식병지 10: 68-72
5 Bender, C., Liyanage, H., Palmer, D., Ullrich, M., Young, S. and Mitchell, R. 1993. Characterization of the genes controlling biosynthesis of the polyketide phytotoxin coronatine including conjugation between coronafacic and coronamic acid. Gene 133: 31-38   DOI   ScienceOn
6 고영진, 박숙영, 이동현. 1996. 우리나라 참다래 궤양병 발생 특성 및 수간주입에 의한 방제. 한식병지 12: 324-330
7 Prosen, D., Hatziloukas, E., Schaad, N. W. and Panopoulos, N. J. 1993. Specific detection of Pseudomonas syringae pv. phaseolicola DNA bean seed by polymerase chain reactionbased amplification of a phaseolotoxin gene region. Phytopathology 83: 965-970   DOI   ScienceOn
8 Bultreys, A. and Gheysen, I. 1999. Biological and molecular detection of toxic lipodepsipeptide-producing Pseudomonas syringae strains and PCR identification in plants. Appl. Environ. Microbiol. 65: 1904-1909
9 Hildebrand, D. C., Schroth, M. N. and Sands, D. C., 1988. Pseudomonas, In : Laboratory Guide for Identification of Plant Pathogenic Bacteria, ed. by N. W. Schaad, pp.60-81. APS Press, St. Paul, Minn., USA
10 Bereswill, S., Bugert, P., Volksch, B., Ullrich, M., Bender, C. L. and Geider, K. 1994. Identification and relatedness of coronatine-producing Pseudomonas syringae pathovars by PCR analysis and sequence determination of the amplification products. Appl. Environ. Microbiol. 60: 2924-2930
11 Takikawa, Y, Serizawa, S., Ichikawa, T., Tsuyumu, S. and Goto, M. 1989. Pseudomonas syringae pv. actinidiae pv. nov.: The causal bacterium of canker of kiwirfuit in Japan. Ann. Phytopathol. Soc. Japan 55: 437-444   DOI
12 Han, H. S., Koh, Y. J., Hur, J.-S. and Jung, J. S. 2003. Identification and characterization of coronatine-producing Pseudomonas syringae pv. actinidiae. J. Microbiol. Biotechnol. 13: 110-118
13 Lydon, J. and Patterson, C. D. 2001. Detection of tabtoxinproducing strains of Pseudomonas syringae by PCR. Lett. Appl. Microbiol. 32: 166-170   DOI   ScienceOn
14 Gardan, L., Shaff, H. and Grimont, P. A. 1997. DNA relatedness among pathovars of P. syringae and related bacteria. In : Pseudomonas syringae Pathovars and Related Pathogens, ed. by Rudolph, K., Burr, T. J., Mansfield, J. W., Stead, D., Vivian, A. and Von Kietzell, J., pp.445-448. Kluwer Academic Publishers, Dordrecht, The Netherlands
15 고영진. 1995. 참다래의 주요 병. 식물병과 농업 1: 3-13
16 Mazarei, M. and Mostofipour, P. 1994. First report of bacterial canker of kiwifruit in Iran. Plant Pathology 43: 1055-1056   DOI   ScienceOn
17 Audy, P., Braat, C. E., Saindon, G., Huang, H. C. and Laroche, A. 1996. A rapid and sensitive PCR-based assay for concurrent detection of bacteria causing common and halo blights in bean seed. Phytopathology 86: 361-366   DOI
18 Manceau, C. and Horvais, A. 1997. Assessment of genetic diversity among strains fo Pseudomonas syringae by PcR-restriction fragment length polymorphism analysis of rRNA operons with special emphasis on P. syringae pv. tomato. Appl. Environ. Microbiol. 63: 498-505
19 정재성, 한효심, 고영진. 2001. Pseudomonas syringae의 식물독소와 독소 생산 균주의 검출을 위한 PCR primer. 식물병연구 7: 123-133
20 Mitchell, R. E. 1976. Isolation and structure of a chlorosis inducing toxin of Pseudomonas phaseolicola. Phytochemistry 15: 1941-1947   DOI   ScienceOn
21 Schaad, N. W., Cheong, S. S., Tamaki, S., Hatziloukas, E. and Panopoulos, N. J. 1995. A combined biological and enzymatic amplification (BIO-PCR) technique to detect Pseudomonas syringae pv. phaseolicola in bean seed extracts. Phytopathology 85: 243-248   DOI   ScienceOn
22 Bender, C. L., Alarcon-Chaidez, F. and Gross, D. C. 1999. Pseudomonas syringae phytotoxins: Mode of action, regulation, and biosynthesis by peptide and polyketide synthetases. Microbiol. Mol. Rev. 63: 266-292
23 Louws, F. J., Fulbright, D. w., Stephens, C. T. and deBruijn, F. J. 1994. Specific genomic fingerprints of phytopathogenic Xanthomonas and Pseudomonas pathovars and strains generated with repetitive sequences and PCR. Appl. Environ. Microbiol. 60: 2286-2295
24 Sorensen, K. N., Kim, K. H. and Takemoto, J. Y 1998. PCR detection of cyclic lipodepsipeptide-producing Pseudomonas syringae pv. syringae and similarity of strains. Appl. Environ. Microbiol. 64: 226-230
25 고영진, 서정규, 이동현, 신종섭, 김승화. 1999. 참다래 궤양병의 약제 방제. 식물병과 농업 5: 95-99
26 Sawada, H., Takeuchi, T. and Matsuda, I. 1997. Comparative analysis of Pseudomonas syringae pv. actinidiae and pv. phaseolicola based on phaseolotoxin-resistant ornithine carbamoyltransferase gene (argK) and 16S-23S rRNA intergenic spacer sequences. Appl. Environ. Microbiol. 63: 282-288