Detection of yellow fever virus: a comparison of quantitative real-time PCR and plaque assay

Yellow fever virus quantitation is performed routinely by cultivation of virus containing samples using susceptible cells. Counting of the resulting plaques provides a marker for the number of infectious particles present in the sample. This assay usually takes up to 5 days before results are obtained and must be carried out under L2 or U laboratory conditions, depending on the yellow fever virus strain used. For clinical diagnosis of yellow fever virus infections the cell culture-based approach takes too long and is of limited practical relevance. Recently, due to its considerable sensitivity, PCR has become a promising method for virus detection. However, whilst PCR can detect virus-specific nucleic acids, it does not allow conclusions to be drawn regarding the infectious potential of the virus detected. Nonetheless, for diagnostic purposes, a rapid, specific and sensitive virus PCR is preferable. Therefore, two independent yellow fever virus-specific real-time PCR assays were established and compared the viral RNA loads to the results of a traditional plaque assay. The estimated ratio of yellow fever virus genomes to infectious particles was between 1000: 1 and 5000: 1; both approaches displayed a comparable precision of < 45%. A significant correlation between genome number as determined by real-time PCR and the corresponding number of plaques in paired samples was found with a Pearson coefficient of correlation of r = 0.88 (P < 0.0001). (C) 2003 Elsevier Science B.V. All rights reserved.