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 Skow, A.
Right arrow Articles by Kaul, K. L.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Skow, A.
Right arrow Articles by Kaul, K. L.

 Previous Article  |  Next Article 

Journal of Clinical Microbiology, June 2005, p. 2876-2880, Vol. 43, No. 6
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.6.2876-2880.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Species-Level Identification of Staphylococcal Isolates by Real-Time PCR and Melt Curve Analysis

Áine Skow, Kathy A. Mangold, Mohammed Tajuddin, Anne Huntington, Brett Fritz, Richard B. Thomson Jr., and Karen L. Kaul*

Department of Pathology and Laboratory Medicine, Evanston Northwestern Healthcare and Northwestern University Feinberg School of Medicine, Evanston, Illinois

Received 13 September 2004/ Returned for modification 18 October 2004/ Accepted 7 January 2005

A real-time PCR assay was developed to identify common staphylococcal species. A single set of consensus primers was designed to amplify a portion of the 16S rRNA gene, and a pair of fluorescence resonance energy transfer probes was used to identify species based on the unique melt properties of the probes resulting from sequence variations in the amplicons from each species. Nine common staphylococcal strains (S. aureus, S. capitis, S. epidermidis, S. haemolyticus, S. hominis, S. lugdunensis, S. schleiferi, S. simulans, and S. warneri) were used for assay development. The species-specific melting profiles were validated by correctly identifying 36 of 37 coagulase-negative staphylococcal (CoNS) isolates identified by ribotyping. In a study of clinical isolates, the PCR/melt curve approach correctly identified 56/56 S. aureus isolates identified by coagulase/protein A latex agglutination. Fifty-four CoNS clinical isolates characterized using the API Staph assay were studied, with the PCR/melt curve approach yielding matching identifications for 32/54 (59%). The API Staph assay was unable to identify 18 CoNS isolates, and differing results were obtained for 4 isolates. Sequencing of the 22 discrepant or unidentified CoNS samples revealed that the PCR/melt curve results were correct for all but one isolate. Thus, PCR/melt curve analysis achieved a nearly 100% accuracy and performed better than biochemical testing. Performance of the PCR/melt curve approach requires less than 2 h after colony selection. This method thus provides a rapid and accurate approach to the identification of staphylococcal species in the clinical laboratory.

* Corresponding author. Mailing address: Department of Pathology, Evanston Northwestern Healthcare, 2650 Ridge Avenue, Evanston, IL 60201. Phone: (847) 570-2052. Fax: (847) 733-5012. E-mail: k-kaul{at}northwestern.edu.

Journal of Clinical Microbiology, June 2005, p. 2876-2880, Vol. 43, No. 6
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.6.2876-2880.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Pinsky, B. A., Samson, D., Ghafghaichi, L., Baron, E. J., Banaei, N. (2009). Comparison of Real-Time PCR and Conventional Biochemical Methods for Identification of Staphylococcus lugdunensis. J. Clin. Microbiol. 47: 3472-3477 [Abstract] [Full Text]  
  • Nuryastuti, T., van der Mei, H. C., Busscher, H. J., Iravati, S., Aman, A. T., Krom, B. P. (2009). Effect of Cinnamon Oil on icaA Expression and Biofilm Formation by Staphylococcus epidermidis. Appl. Environ. Microbiol. 75: 6850-6855 [Abstract] [Full Text]  
  • Zouache, K., Voronin, D., Tran-Van, V., Mavingui, P. (2009). Composition of Bacterial Communities Associated with Natural and Laboratory Populations of Asobara tabida Infected with Wolbachia. Appl. Environ. Microbiol. 75: 3755-3764 [Abstract] [Full Text]  
  • Sastalla, I., Chim, K., Cheung, G. Y. C., Pomerantsev, A. P., Leppla, S. H. (2009). Codon-Optimized Fluorescent Proteins Designed for Expression in Low-GC Gram-Positive Bacteria. Appl. Environ. Microbiol. 75: 2099-2110 [Abstract] [Full Text]  
  • Mehta, M. S., Paule, S. M., Thomson, R. B., Kaul, K. L., Peterson, L. R. (2009). Identification of Staphylococcus Species Directly from Positive Blood Culture Broth by Use of Molecular and Conventional Methods. J. Clin. Microbiol. 47: 1082-1086 [Abstract] [Full Text]  
  • Frank, K. L., del Pozo, J. L., Patel, R. (2008). From Clinical Microbiology to Infection Pathogenesis: How Daring To Be Different Works for Staphylococcus lugdunensis. Clin. Microbiol. Rev. 21: 111-133 [Abstract] [Full Text]  
  • Taponen, S., Koort, J., Bjorkroth, J., Saloniemi, H., Pyorala, S. (2007). Bovine Intramammary Infections Caused by Coagulase-Negative Staphylococci May Persist Throughout Lactation According to Amplified Fragment Length Polymorphism-Based Analysis. J DAIRY SCI 90: 3301-3307 [Abstract] [Full Text]  
  • Shah, M. M., Iihara, H., Noda, M., Song, S. X., Nhung, P. H., Ohkusu, K., Kawamura, Y., Ezaki, T. (2007). dnaJ gene sequence-based assay for species identification and phylogenetic grouping in the genus Staphylococcus. Int. J. Syst. Evol. Microbiol. 57: 25-30 [Abstract] [Full Text]  
  • Shittu, A., Lin, J., Morrison, D., Kolawole, D. (2006). Identification and molecular characterization of mannitol salt positive, coagulase-negative staphylococci from nasal samples of medical personnel and students.. J Med Microbiol 55: 317-324 [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»