Abstract
A two step quantitative RT-PCR assay was validated to monitor the deformed wing virus (DWV) RNA loads in Apis mellifera L. and Varroa destructor. A pair of primers hybridising in a conserved domain of the putative DWV RNA polymerase gene region was designed. These primers amplified a 69-nucleotide fragment which was quantified using the SYBR-green chemistry. The experimental validation of the method showed that the RNA extraction and cDNA synthesis steps were responsible for the greatest variability in the results while assays repeated on different PCR plates were reproducible. Quantitative RTPCR analysis on drone bee prepupae showed that DWV RNA loads were higher in cells parasitized by several mother mites. In workers, DWV prevalence was directly correlated to mite infestation and DWV was detected in all the bee developmental stages except in eggs. Very important DWV RNA loads could be recorded even in absence of clinical sign; however bees emerging with deformed wings were predominantly infected by DWV. In mites collected on emerging bees, the DWV RNA yields varied from 10(4) to 10(6) copies per mite but might exceed 10(8) copies in some cases