Establishing the risk of West Nile virus transmission through mosquito bites using the digital PCR and real-time PCR methods

Abstract

West Nile virus (WNV) is a re-emerging zoonotic pathogen which is a threat to both human and animal health. In Europe, there has been a marked expansion of WNV outbreaks in recent decades, causing more than 2000 symptomatic cases in 2018 alone. Winter temperatures between 2°C and 6°C were one of the strongest predictors of annual West Nile virus infections; a possible explanation for this result is that successful overwintering of infected adult mosquitoes (probably Culex pipiens) is the key to the intensity of outbreaks in the following year. The aim of the study was to compare the two diagnostic methods Real-Time PCR and dPCR, used in the detection of West NILE virus in mosquito vectors. Between April 2023 and June 2024, mosquitoes were captured from the Danube Delta area and from the northeastern part of Romania in the city of Iasi, using the New Standard Miniature Incandescent Light Trap, model 1012. After morphological identification, mosquitoes were separated according to Culex pipiens species. Pools of 30 mosquitoes per pool were made, being tested for the presence of the West Nile virus. The advanced dPCR method was used to detect West Nile virus. Digital™ PCR (dPCR™) is an innovative technology that provides ultrasensitive nucleic acid detection and absolute quantification. It is very effective for resolving low-abundance targets, such as very small amounts of virus inside mosquito vectors.