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Application of a Real-Time Polymerase Chain Reaction with Internal Positive Control for Detection and Quantification of Enterovirus in Cerebrospinal Fluid

  •  S. Monpoeho
  •  M. Coste-Burel
  •  M. Costa-Mattioli
  •  B. Besse
  •  J. Chomel
  •  S. Billaudel
  •  V. Ferré
Article

Abstract.

A quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) method based on TaqMan technology was developed to determine the presence and amount of enterovirus RNA. In order to prevent false-negative results, a one-step multiplex RT-PCR was optimized. It contains two dual-labelled fluorogenic probes to quantify the 5′ noncoding region of enterovirus and detect an internal positive control. In the present study, 104 cerebrospinal fluid samples collected during an outbreak of enteroviral meningitis were analyzed using this method. Amplification of the internal positive control was effective in all but two specimens, confirming the absence of PCR inhibitors and allowing the results of amplification to be validated. The sensitivity of the RT-PCR was 96.8%, while that of cell culture was 34.9%. Genomic viral loads found ranged between 3.3 and 5.9 log10 copies per milliliter of cerebrospinal fluid (mean, 4.8 log10 copies/ml). This fluorogenic enterovirus RT-PCR allows large numbers of samples to be screened rapidly. Moreover, its sensitivity and reproducibility make it highly reliable. With these characteristics, the enterovirus RT-PCR can be a useful tool that may offer considerable benefit in the clinical management of patients with enteroviral infections.

Keywords

Meningitis Noncoding Region Internal Positive Control Enteroviral Infection Fluorogenic Probe 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Springer-Verlag 2002

Authors and Affiliations

  •  S. Monpoeho
    • 1
  •  M. Coste-Burel
    • 1
  •  M. Costa-Mattioli
    • 1
  •  B. Besse
    • 1
  •  J. Chomel
    • 2
  •  S. Billaudel
    • 1
  •  V. Ferré
    • 1
  1. 1.Laboratoire de Virologie, Institut de Biologie, CHRU Hotel Dieu, 9 Quai Moncousu, 44093 Nantes Cedex 01, France
  2. 2.Centre National de Référence des Entérovirus, Teaching Hospital, Lyon, France

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