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Eur Respir J 2003; 21:944-951
Copyright ©ERS Journals Ltd 2003

Evaluation of a quantitative real-time PCR for the detection of respiratory syncytial virus in pulmonary diseases

I. Borg1,2, G. Rohde2, S. Löseke1, J. Bittscheidt3, G. Schultze-Werninghaus2, V. Stephan3 and A. Bufe1

1 Dept of Experimental Pneumology, Ruhr-University Bochum, 2 Dept of Internal Medicine, Division of Pneumology, Allergology andSleep Medicine, University Hospital Bergmannsheil and 3 University Children's Hospital, Bochum, Germany

CORRESPONDENCE: I. Borg, Experimental Pneumology, BGFA XU 17, University Hospital Bergmannsheil, Bürkle-de-la-Camp-Platz 1, D-44789 Bochum, Germany. Fax: 49 2343074682. E-mail: Irmgard.Borg@ruhr-uni-bochum.de

Keywords: bronchiolitis, chronic obstructive pulmonary disease, pulmonary diseases, real-time polymerase chain reaction, respiratory syncytial virus, viral load

Received: September 24, 2002
Accepted January 29, 2003

This study was supported by the Bundesministerium für Bildung und Forschung (BMBF) grant 01GC-9802/8 and 01GC-0101.

Respiratory syncytial virus (RSV) is known to cause acute lower respiratory tract infections (ARI) in young children and is involved in exacerbation of chronic obstructive pulmonary disease (COPD) in adults. The role of RSV in stable COPD and the viral load in different respiratory diseases has not been investigated to date.

The present authors established and evaluated a quantitative TaqMan® real-time polymerase chain reaction assay specific for RSV subgroup A. Absolute quantification for the determination of viral load input was achieved using a control plasmid. The assay allowed for a quantification over a >6-log range and a detection limit of <10 RSV copies per reaction mixture.

The assay was 30 times more sensitive than conventional nested polymerase chain reaction assays. Interassay sd was 1.3 and coefficient of variation 4.7% on average. Clinical specimens from infants with ARI (n=62) and elderly adults with COPD (n=125) were compared for viral loads. A total of 47% RSV-positive samples were found in the ARI study group and 28% in the COPD study group. The viral load of both study groups was found to differ significantly. In the ARI study group the viral load was increased almost 2000-fold, suggesting acute infection in this group and former or latent infection in the COPD group.

Respiratory syncytial virus-A specific TaqMan® real-time polymerase chain reaction assay is a sensitive and rapid method for the determination of viral load in clinical samples. It enables differential statements concerning the involvement of respiratory syncytial virus in acute lower respiratory tract infections and chronic obstructive pulmonary disease to be achieved.




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