Quantification of Marek's disease virus in chicken lymphocytes using the polymerase chain reaction with fluorescence detection.
A quantitative assay was developed for Marek's disease virus (MDV). The assay determines the numbers of viral genomes present in samples by polymerase chain reaction (PCR) amplification of a portion of the viral genome for a restricted number of cycles. Fluorescent-tagged primers are used for the PCR amplification which allows quantification of the fluorescent product. Previously, quantitation of Marek's disease virus has required plaque assays, which are laborious and potentially error-prone, and this had limited quantitative comparisons. The PCR assay is rapid, less laborious and can be applied to high levels of accuracy, since replicate assays can be carried out relatively easily. The PCR-based assay assesses the number of viral genomes present in the sample, rather than the numbers of infected cells measured in the plaque assay, however correlation between the two assays is high, suggesting viral copy number per cell may be rather uniform. In crosses between genetically resistant and susceptible animals the PCR-based assay was correlated significantly with subsequent development of disease, and was a better predictor than the plaque assay of the likelihood of development of pathological disease in the birds studied.
Bumstead
N
N
Institute for Animal Health, Compton Laboratory, Berkshire, UK. Bumstead@bbsrc.ac.uk
Sillibourne
J
J
Rennie
M
M
Ross
N
N
Davison
F
F
D000818:Animals; D002645:Chickens / Q000821:virology*; D004247:DNA / Q000032:analysis; D005453:Fluorescence; D006566:Herpesviridae Infections / Q000235:genetics / Q000378:metabolism; D008381:Herpesvirus 2, Gallid / Q000378:metabolism*; D008214:Lymphocytes / Q000821:virology*; D008380:Marek Disease / Q000401:mortality; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D013997:Time Factors; D010948:Viral Plaque Assay
D004247:DNA