TY - JOUR T1 - Bluetongue virus detection by two real-time RT-qPCRs targeting two different genomic segments. JF - J Virol Methods Y1 - 2007 A1 - Toussaint, J F A1 - Sailleau, C A1 - Bréard, E A1 - Zientara, S A1 - Kris De Clercq KW - Animals KW - Bluetongue KW - Bluetongue virus KW - Cell Line KW - Cricetinae KW - DNA Primers KW - DNA Probes KW - Genome, Viral KW - Nucleic Acid Amplification Techniques KW - Reproducibility of Results KW - Reverse Transcriptase Polymerase Chain Reaction KW - RNA, Messenger KW - RNA, Viral KW - Serotyping KW - Sheep Diseases KW - Sheep, Domestic KW - Time Factors AB -

The detection of the bluetongue virus (BTV) by conventional methods is especially difficult and labour-intensive. Molecular diagnosis is also complex because of the high genetic diversity between and within the 24 serotypes of BTV. In the present study, two laboratories joined forces to develop and validate two new RT-qPCRs detecting and amplifying BTV segments 1 and 5. The 2 assays detect strains from all 24 serotypes. They both have a detection limit of 0.01 ECE50 and all 114 samples from BTV-free goats, sheep and cattle were negative. The two assays resulted in similar C(t) values when testing biological samples collected in sheep infected experimentally with a field strain of BTV from the Mediterranean basin. On average, the C(t) values obtained with the 2 methods applied to the 24 serotypes were not significantly different from each other, but some moderate to high differences were seen with a few strains. Therefore these two methods are complementary and could be used in parallel to confirm the diagnosis of a possible new introduction of BTV. An RT-qPCR amplifying a fragment of the beta-actin mRNA was also developed and validated as internal control for the bluetongue specific assays. The three assays described allow a reliable and rapid detection of BTV.

VL - 140 CP - 1-2 U1 - http://www.ncbi.nlm.nih.gov/pubmed/17196266?dopt=Abstract M3 - 10.1016/j.jviromet.2006.11.007 ER -