This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Supplemental material
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 Pingle, M. R.
Right arrow Articles by Barany, F.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Pingle, M. R.
Right arrow Articles by Barany, F.

 Previous Article  |  Next Article 

Journal of Clinical Microbiology, June 2007, p. 1927-1935, Vol. 45, No. 6
0095-1137/07/$08.00+0     doi:10.1128/JCM.00226-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Multiplexed Identification of Blood-Borne Bacterial Pathogens by Use of a Novel 16S rRNA Gene PCR-Ligase Detection Reaction-Capillary Electrophoresis Assay{triangledown} ,{dagger}

Maneesh R. Pingle,1 Kathleen Granger,1 Philip Feinberg,1 Rebecca Shatsky,2 Bram Sterling,1 Mark Rundell,1 Eric Spitzer,3 Davise Larone,1,2 Linnie Golightly,1,4 and Francis Barany1*

Department of Microbiology,1 Department of Pathology and Laboratory Medicine,2 Division of Infectious Disease and International Medicine, Weill Medical College of Cornell University, New York, New York,4 Department of Pathology, Stony Brook University Medical Center, Stony Brook, New York3

Received 29 January 2007/ Returned for modification 19 March 2007/ Accepted 2 April 2007

We have developed a novel high-throughput PCR-ligase detection reaction-capillary electrophoresis (PCR-LDR-CE) assay for the multiplexed identification of 20 blood-borne pathogens (Staphylococcus epidermidis, Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, Enterococcus faecium, Listeria monocytogenes, Streptococcus pneumoniae, Streptococcus pyogenes, Streptococcus agalactiae, Escherichia coli, Klebsiella pneumoniae, Haemophilus influenzae, Pseudomonas aeruginosa, Acinetobacter baumannii, Neisseria meningitidis, Bacteroides fragilis, Bacillus anthracis, Yersinia pestis, Francisella tularensis, and Brucella abortus), the last four of which are biothreat agents. The method relies on the amplification of two regions within the bacterial 16S rRNA gene, using universal PCR primers and querying the identity of specific single-nucleotide polymorphisms within the amplified regions in a subsequent LDR. The ligation products vary in color and size and are separated by CE. Each organism generates a specific pattern of ligation products, which can be used to distinguish the pathogens using an automated software program we developed for that purpose. The assay has been verified on 315 clinical isolates and demonstrated a detection sensitivity of 98%. Additionally, 484 seeded blood cultures were tested, with a detection sensitivity of 97.7%. The ability to identify geographically variant strains of the organisms was determined by testing 132 isolates obtained from across the United States. In summary, the PCR-LDR-CE assay can successfully identify, in a multiplexed fashion, a panel of 20 blood-borne pathogens with high sensitivity and specificity.

* Corresponding author. Mailing address: Department of Microbiology, Weill Medical College of Cornell University, 1300 York Avenue, Box 62, New York, NY 10021. Phone: (212) 746-6509. Fax: (212) 746-8104. E-mail: barany{at}med.cornell.edu

{triangledown} Published ahead of print on 11 April 2007.

{dagger} Supplemental material for this article may be found at http://jcm.asm.org.

Journal of Clinical Microbiology, June 2007, p. 1927-1935, Vol. 45, No. 6
0095-1137/07/$08.00+0     doi:10.1128/JCM.00226-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Severgnini, M., Cremonesi, P., Consolandi, C., Caredda, G., De Bellis, G., Castiglioni, B. (2009). ORMA: a tool for identification of species-specific variations in 16S rRNA gene and oligonucleotides design. Nucleic Acids Res 37: e109-e109 [Abstract] [Full Text]  
  • Cremonesi, P., Pisoni, G., Severgnini, M., Consolandi, C., Moroni, P., Raschetti, M., Castiglioni, B. (2009). Pathogen detection in milk samples by ligation detection reaction-mediated universal array method. J DAIRY SCI 92: 3027-3039 [Abstract] [Full Text]  
  • Das, S., Pingle, M. R., Munoz-Jordan, J., Rundell, M. S., Rondini, S., Granger, K., Chang, G.-J. J., Kelly, E., Spier, E. G., Larone, D., Spitzer, E., Barany, F., Golightly, L. M. (2008). Detection and Serotyping of Dengue Virus in Serum Samples by Multiplex Reverse Transcriptase PCR-Ligase Detection Reaction Assay. J. Clin. Microbiol. 46: 3276-3284 [Abstract] [Full Text]  
  • Rondini, S., Pingle, M. R., Das, S., Tesh, R., Rundell, M. S., Hom, J., Stramer, S., Turner, K., Rossmann, S. N., Lanciotti, R., Spier, E. G., Munoz-Jordan, J., Larone, D., Spitzer, E., Barany, F., Golightly, L. M. (2008). Development of Multiplex PCR-Ligase Detection Reaction Assay for Detection of West Nile Virus. J. Clin. Microbiol. 46: 2269-2279 [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»