Bluetongue serotype 4 (BTV4) has been detected for the first time in tissue samples from 2 mouflons (Ovis aries musimon) from the South of Spain, in a retrospective study. The samples included in this study had been fixed and paraffin-embedded for over a year prior to their analysis using a BTV group-specific and a BTV4-specific RT-PCR test. Lung and lymphatic nodes were found positive in both specimens. The amplified DNA was confirmed to be BTV4 by sequencing the RT-PCR products and comparing them with other sequences from GenBank. The combination of RNA extraction from paraffin-embedded samples and serotype-specific real-time RT-PCR assays provides the tools for the detection of BTV from samples stored for a long time. The results shown in this study set out the basis for a greater survey with fixed samples from different species of wild ruminants that the veterinary services have been collecting for years.
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