Field-Collected Belgian Culex pipiens and Culex torrentium Mosquitoes are Competent for West Nile Virus Under Different Temperature Conditions.

Belgium detected West Nile virus (WNV) for the first time on its territory in August 2025 during a monitoring of wild birds. To support risk assessments, we determined the vector competence of field-collected Belgian Culex pipiens and Culex torrentium mosquitoes, which are considered important WNV vectors. Females fed on a WNV-spiked (lineage 1 Israel 98 strain) blood meal were kept for 14 days at (i) a constant 25°C, (ii) a 25/15°C day/night gradient reflecting current Belgian summers, or (iii) a 25/20°C gradient simulating a climate change scenario. After the 14-day incubation period, abdomens, legs/wings/head, and saliva were collected and tested in RT-qPCR to determine the infection, dissemination, and transmission efficiencies (TEs). Observed infection efficiencies (IEs) (25.5%, 32.7%, and 34.3%) did not significantly differ between temperature conditions, and mostly mosquitoes with the lowest viral loads were dissemination-positive. The dissemination efficiency (DE) at 25/15°C (28.6%) was higher than in both other conditions and statistically significant compared to 25/20°C (10.4%), indicating that WNV more easily passes the midgut barrier under current Belgian summer temperatures. The entry of WNV into the salivary glands did not seem to be altered by temperature, as no significant differences were observed between the obtained TEs (3.9%, 8.2%, and 1.5% at 25°C, 25/15°C, and 25/20°C, respectively). By performing virus isolations on saliva, we proved that Culex pipiens is a competent vector for WNV at 25 and 25/15°C, while this was not confirmed at 25/20°C. Notably, a small percentage of the field-collected mosquitoes in this study were identified as Culex torrentium (7%), which also showed high competence for WNV. To conclude, our results suggest that field-collected Belgian Culex pipiens mosquitoes are competent vectors for WNV at 25°C and under current Belgian temperatures, emphasizing their potential to contribute to local WNV circulation and the importance of ongoing surveillance in birds, horses, and mosquitoes.