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Journal of Clinical Microbiology, September 2001, p. 3129-3134, Vol. 39, No. 9
0095-1137/01/$04.00+0 DOI: 10.1128/JCM.39.9.3129-3134.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Quantitative Detection of Streptococcus
pneumoniae in Nasopharyngeal Secretions by Real-Time
PCR
Oliver
Greiner,1
Philip J. R.
Day,1,2
Philipp
P.
Bosshard,3
Fatime
Imeri,3
Martin
Altwegg,3 and
David
Nadal1,*
Division of Infectious
Diseases1 and Division of
Oncology,2 University Children's Hospital of
Zurich, CH-8032 Zurich, and Department of Medical
Microbiology, University of Zurich, CH-8028
Zurich,3 Switzerland
Received 14 March 2001/Returned for modification 13 May
2001/Accepted 28 June 2001
Streptococcus pneumoniae is an important cause of
community-acquired pneumonia. However, in this setting the diagnostic
sensitivity of blood cultures is below 30%. Since during such
infections changes in the amounts of S. pneumoniae may
also occur in the upper respiratory tract, quantification of these
bacteria in nasopharnygeal secretions (NPSs) may offer a suitable
diagnostic approach. Real-time PCR offers a sensitive, efficient, and
routinely reproducible approach to quantification. Using primers and a
fluorescent probe specific for the pneumolysin gene, we were able to
detect DNA from serial dilutions of S. pneumoniae cells
in which the quantities of DNA ranged from the amounts extracted from 1 to 106 cells. No difference was noted when the same
DNA was mixed with DNA extracted from NPSs shown to be deficient of
S. pneumoniae following culture, suggesting that this
bacterium can be detected and accurately quantitated in clinical
samples. DNAs from Haemophilus influenzae,
Moraxella catarrhalis, or alpha-hemolytic streptococci other than S. pneumoniae were not amplified or were only
weakly amplified when there were
106 cells per reaction
mixture. When the assay was applied to NPSs from patients with
respiratory tract infections, the assay performed with a sensitivity of
100% and a specificity of up to 96% compared to the culture results.
The numbers of S. pneumoniae organisms detected by
real-time PCR correlated with the numbers detected by semiquantitative
cultures. A real-time PCR that targeted the pneumolysin gene provided a
sensitive and reliable means for routine rapid detection and
quantification of S. pneumoniae present in NPSs. This
assay may serve as a tool to study changes in the amounts of S.
pneumoniae during lower respiratory tract infections.
*
Corresponding author. Mailing address: Division of
Infectious Diseases, University Children`s Hospital of Zurich,
Steinwiesstrasse 75, CH-8032 Zurich, Switzerland. Phone: 41-1-266-7562. Fax: 41-1-266-7157. E-mail:
david.nadal{at}kispi.unizh.ch.
Journal of Clinical Microbiology, September 2001, p. 3129-3134, Vol. 39, No. 9
0095-1137/01/$04.00+0 DOI: 10.1128/JCM.39.9.3129-3134.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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