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 Warsen, A. E.
Right arrow Articles by Call, D. R.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Warsen, A. E.
Right arrow Articles by Call, D. R.
Agricola
Right arrow Articles by Warsen, A. E.
Right arrow Articles by Call, D. R.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, July 2004, p. 4216-4221, Vol. 70, No. 7
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.7.4216-4221.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Simultaneous Discrimination between 15 Fish Pathogens by Using 16S Ribosomal DNA PCR and DNA Microarrays

Adelaide E. Warsen,1 Melissa J. Krug,1 Stacey LaFrentz,1 Danielle R. Stanek,2 Frank J. Loge,1,3 and Douglas R. Call1,4*

Department of Veterinary Microbiology and Pathology,1 Washington Animal Disease Diagnostic Laboratory,2 Department of Civil and Environmental Engineering,3 WSU & UI Center for Reproductive Biology, Washington State University, Pullman, Washington4

Received 15 December 2003/ Accepted 31 March 2004

We developed a DNA microarray suitable for simultaneous detection and discrimination between multiple bacterial species based on 16S ribosomal DNA (rDNA) polymorphisms using glass slides. Microarray probes (22- to 31-mer oligonucleotides) were spotted onto Teflon-masked, epoxy-silane-derivatized glass slides using a robotic arrayer. PCR products (ca. 199 bp) were generated using biotinylated, universal primer sequences, and these products were hybridized overnight (55°C) to the microarray. Targets that annealed to microarray probes were detected using a combination of Tyramide Signal Amplification and Alexa Fluor 546. This methodology permitted 100% specificity for detection of 18 microbes, 15 of which were fish pathogens. With universal 16S rDNA PCR (limited to 28 cycles), detection sensitivity for purified control DNA was equivalent to <150 genomes (675 fg), and this sensitivity was not adversely impacted either by the presence of competing bacterial DNA (1.1 x 106 genomes; 5 ng) or by the addition of up to 500 ng of fish DNA. Consequently, coupling 16S rDNA PCR with a microarray detector appears suitable for diagnostic detection and surveillance for commercially important fish pathogens.

* Corresponding author. Mailing address: Department of Veterinary Microbiology and Pathology, 490 Bustad Hall, Washington State University, Pullman, WA 99164-7040. Phone: (509) 335-6313. Fax: (509) 335-8529. E-mail: drcall{at}vetmed.wsu.edu.

Applied and Environmental Microbiology, July 2004, p. 4216-4221, Vol. 70, No. 7
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.7.4216-4221.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Miller, S. M., Tourlousse, D. M., Stedtfeld, R. D., Baushke, S. W., Herzog, A. B., Wick, L. M., Rouillard, J. M., Gulari, E., Tiedje, J. M., Hashsham, S. A. (2008). In Situ-Synthesized Virulence and Marker Gene Biochip for Detection of Bacterial Pathogens in Water. Appl. Environ. Microbiol. 74: 2200-2209 [Abstract] [Full Text]  
  • Han, W., Liu, B., Cao, B., Beutin, L., Kruger, U., Liu, H., Li, Y., Liu, Y., Feng, L., Wang, L. (2007). DNA Microarray-Based Identification of Serogroups and Virulence Gene Patterns of Escherichia coli Isolates Associated with Porcine Postweaning Diarrhea and Edema Disease. Appl. Environ. Microbiol. 73: 4082-4088 [Abstract] [Full Text]  
  • Brodie, E. L., DeSantis, T. Z., Joyner, D. C., Baek, S. M., Larsen, J. T., Andersen, G. L., Hazen, T. C., Richardson, P. M., Herman, D. J., Tokunaga, T. K., Wan, J. M., Firestone, M. K. (2006). Application of a High-Density Oligonucleotide Microarray Approach To Study Bacterial Population Dynamics during Uranium Reduction and Reoxidation. Appl. Environ. Microbiol. 72: 6288-6298 [Abstract] [Full Text]  
  • Soule, M., Kuhn, E., Loge, F., Gay, J., Call, D. R. (2006). Using DNA microarrays to identify library-independent markers for bacterial source tracking.. Appl. Environ. Microbiol. 72: 1843-1851 [Abstract] [Full Text]  
  • Giammarinaro, P., Leroy, S., Chacornac, J.-P., Delmas, J., Talon, R. (2005). Development of a New Oligonucleotide Array To Identify Staphylococcal Strains at Species Level. J. Clin. Microbiol. 43: 3673-3680 [Abstract] [Full Text]  
  • Soule, M., Cain, K., LaFrentz, S., Call, D. R. (2005). Combining Suppression Subtractive Hybridization and Microarrays To Map the Intraspecies Phylogeny of Flavobacterium psychrophilum. Infect. Immun. 73: 3799-3802 [Abstract] [Full Text]  
  • Panicker, G., Call, D. R., Krug, M. J., Bej, A. K. (2004). Detection of Pathogenic Vibrio spp. in Shellfish by Using Multiplex PCR and DNA Microarrays. Appl. Environ. Microbiol. 70: 7436-7444 [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»