This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Schönfeld, J.
Right arrow Articles by Smalla, K.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Schönfeld, J.
Right arrow Articles by Smalla, K.
Agricola
Right arrow Articles by Schönfeld, J.
Right arrow Articles by Smalla, K.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, December 2003, p. 7248-7256, Vol. 69, No. 12
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.12.7248-7256.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Specific and Sensitive Detection of Ralstonia solanacearum in Soil on the Basis of PCR Amplification of fliC Fragments

J. Schönfeld,1 H. Heuer,2 J. D. van Elsas,3 and K. Smalla1,{dagger}*

Institute for Plant Virology, Microbiology, and Biosafety, Federal Biological Research Centre for Agriculture and Forestry, 38104 Braunschweig, Germany,1 Department of Biological Sciences, University of Idaho, Moscow, Idaho,2 Plant Research International, 6700 AA Wageningen, The Netherlands3

Received 9 March 2003/ Accepted 4 September 2003

Ralstonia solanacearum is the causative agent of bacterial wilt in many important crops. A specific and sensitive PCR detection method that uses primers targeting the gene coding for the flagella subunit, fliC, was established. Based on the first fliC gene sequence of R. solanacearum strain K60 available at GenBank, the Ral_fliC PCR primer system was designed; this system yielded a single 724-bp product with the DNAs of all of the R. solanacearum strains tested. However, R. pickettii and four environmental Ralstonia isolates also yielded amplicons. The Ral_fliC PCR products obtained with 12 strains (R. solanacearum, R. pickettii, and environmental isolates) were sequenced. By sequence alignment, Rsol_fliC primers specific for R. solanacearum were designed. With this primer system, a specific 400-bp PCR product was obtained from all 82 strains of R. solanacearum tested. Six strains of R. pickettii and several closely related environmental isolates yielded no PCR product; however, a product was obtained with one Pseudomonas syzygii strain. A GC-clamped 400-bp fliC product could be separated in denaturing gradient gels and allowed us to distinguish P. syzygii from R. solanacearum. The Rsol_fliC PCR system was applied to detect R. solanacearum in soil. PCR amplification, followed by Southern blot hybridization, allowed us to detect about one target DNA molecule per PCR, which is equivalent to 103 CFU g of bulk soil-1. The system was applied to survey soils from different geographic origins for the presence of R. solanacearum.

* Corresponding author. Mailing address: Institute for Plant Virology, Microbiology, and Biosafety, Federal Biological Research Centre for Agriculture and Forestry, Messeweg 11-12, 38104 Braunschweig, Germany. Phone: 49-531-2993814. Fax: 49-531-2993013. E-mail: K.Smalla{at}bba.de.

{dagger} Present address: Microbial Ecology, Groningen University Biological Center, 9751 Haren, The Netherlands.

Applied and Environmental Microbiology, December 2003, p. 7248-7256, Vol. 69, No. 12
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.12.7248-7256.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Kaestli, M., Mayo, M., Harrington, G., Watt, F., Hill, J., Gal, D., Currie, B. J. (2007). Sensitive and Specific Molecular Detection of Burkholderia pseudomallei, the Causative Agent of Melioidosis, in the Soil of Tropical Northern Australia. Appl. Environ. Microbiol. 73: 6891-6897 [Abstract] [Full Text]  
  • Castillo, J. A., Greenberg, J. T. (2007). Evolutionary Dynamics of Ralstonia solanacearum. Appl. Environ. Microbiol. 73: 1225-1238 [Abstract] [Full Text]  
  • Bergsma-Vlami, M., Prins, M. E., Staats, M., Raaijmakers, J. M. (2005). Assessment of Genotypic Diversity of Antibiotic-Producing Pseudomonas Species in the Rhizosphere by Denaturing Gradient Gel Electrophoresis. Appl. Environ. Microbiol. 71: 993-1003 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»