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Journal of Clinical Microbiology, September 2003, p. 4366-4371, Vol. 41, No. 9
0095-1137/03/$08.00+0     DOI: 10.1128/JCM.41.9.4366-4371.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Comparison and Evaluation of Real-Time PCR, Real-Time Nucleic Acid Sequence-Based Amplification, Conventional PCR, and Serology for Diagnosis of Mycoplasma pneumoniae

Kate E. Templeton,1 Sitha A. Scheltinga,1 A. Willy Graffelman,2 Jolanda M. van Schie,1 Jantine W. Crielaard,1 Peter Sillekens,3 Peterhans J. van den Broek,4 Herman Goossens,1,5 Matthias F. C. Beersma,1 and Eric C. J. Claas1*

Departments of Medical Microbiology,1 Infectious Diseases, Center of Infectious Diseases,2 Department of General Practice and Nursing Home Medicine, Leiden University Medical Center, Leiden,4 BioMerieux, Boxtel, The Netherlands,3 Department of Microbiology, University of Antwerp, Antwerp, Belgium5

Received 17 February 2003/ Returned for modification 26 May 2003/ Accepted 3 July 2003

Mycoplasma pneumoniae is a common cause of community-acquired pneumonia and lower-respiratory-tract infections. Diagnosis has traditionally been obtained by serological diagnosis, but increasingly, molecular techniques have been applied. However, the number of studies actually comparing these assays is limited. The development of a novel duplex real-time PCR assay for detection of M. pneumoniae in the presence of an internal control real-time PCR is described. In addition, real-time nucleic acid sequence-based amplification (NASBA) on an iCycler apparatus is evaluated. These assays were compared to serology and a conventional PCR assay for 106 clinical samples from patients with lower-respiratory-tract infection. Of the 106 samples, 12 (11.3%) were positive by all the molecular methods whereas serology with acute sample and convalescent samples detected 6 (5.6%) and 9 (8.5%), respectively. Clinical symptoms of the patients with Mycoplasma-positive results were compared to those of the other patients with lower-respiratory-tract infections, and it was found that the results for mean lower age numbers as well as the presence of chills, increased erythrocyte sedimentation rate, and raised C-reactive protein levels showed significant differences. Molecular methods are superior for diagnosis of M. pneumoniae, providing more timely diagnosis. In addition, using real-time methods involves less hands-on time and affords the ability to monitor the reaction in the same tube.

* Corresponding author. Mailing address: Department of Medical Microbiology, Center of Infectious Diseases, Leiden University Medical Center, P.O. Box 9600, 2300 RC Leiden, The Netherlands. Phone: 31 71 526 3650. Fax: 31 71 524 8148. E-mail: E.Claas{at}LUMC.NL.

Journal of Clinical Microbiology, September 2003, p. 4366-4371, Vol. 41, No. 9
0095-1137/03/$08.00+0     DOI: 10.1128/JCM.41.9.4366-4371.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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