Measurement of the infection and dissemination of bluetongue virus in culicoides biting midges using a semi-quantitative rt-PCR assay and isolation of infectious virus.
Authors: Veronesi Eva, Antony Frank, Gubbins Simon, Golding Nick, Blackwell Alison, Mertens Peter Pc, Brownlie Joe, Darpel Karin E, Mellor Philip S, Carpenter Simon
Journal: PloS one
Summary
# Editorial Summary Between 2006 and 2009, bluetongue virus outbreaks across northern Europe prompted investigation into whether local Culicoides species could act as competent vectors, yet the semi-quantitative RT-PCR methods used to detect viral RNA in midges had never been formally validated for this application, making interpretation of results unreliable. Veronesi and colleagues addressed this critical gap by establishing a time-series study in two laboratory-reared Culicoides colonies, directly comparing semi-quantitative RT-PCR detection of BTV RNA against the gold standard of infectious virus isolation in cell culture. The validation demonstrated that RT-PCR successfully tracked BTV dissemination through the midges' bodies over time, with detectable viral RNA appearing before infectious virus could be cultured and persisting after infectious titres declined, suggesting the assay captures the full infection trajectory rather than only viable virus. For equine and livestock professionals, this validation provides confidence that field studies using RT-PCR to identify BTV-infected Culicoides populations are generating interpretable data, strengthening the epidemiological evidence needed to assess vector competence and disease transmission risk in previously affected regions. The work establishes a diagnostic framework that has become essential for surveillance programmes monitoring both bluetongue and other Culicoides-borne viruses including African horse sickness virus and Schmallenberg virus across Europe.
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Practical Takeaways
- •sqPCR assays are now validated for reliably detecting BTV in Culicoides vectors, improving diagnostic accuracy in outbreak investigations
- •Results from European vector studies using sqPCR can be interpreted with greater confidence for understanding BTV epidemiology
- •Understanding viral dissemination in Culicoides helps inform control strategies during future bluetongue, AHSV, or Schmallenberg outbreaks affecting livestock and horses
Key Findings
- •Semi-quantitative RT-PCR assays were validated for detecting BTV RNA in Culicoides species through time-series analysis
- •BTV RNA detection by sqPCR was compared with traditional infectious virus isolation on cell culture
- •Two colony species of Culicoides were used to establish viral dissemination patterns
- •The study provided methodological validation for interpreting previous European BTV outbreak investigations