Relevant Day/Night Temperatures Simulating Belgian Summer Conditions Reduce Japanese Encephalitis Virus Dissemination and Transmission in Belgian Field-Collected Culex pipiens Mosquitoes

Abstract

Japanese encephalitis virus (JEV), a zoonotic mosquito-borne Flavivirus, can be considered an emerging infectious disease. Therefore, vector competence studies with indigenous mosquitoes from regions where JEV is not yet endemic are of great importance. In our study, we compared the vector competence of Culex pipiens mosquitoes emerged from Belgian field-caught larvae under two different temperature conditions: a constant 25 °C and a 25/15 °C day/night temperature gradient representing typical summer temperatures in Belgium. Three- to seven-day-old F0-generation mosquitoes were fed on a JEV genotype 3 Nakayama strain spiked blood-meal and incubated for 14 days at the two aforementioned temperature conditions. Similar infection rates of 36.8% and 35.2% were found in both conditions. The observed dissemination rate in the gradient condition was, however, significantly lower compared to the constant temperature condition (8% versus 53.6%, respectively). JEV was detected by RT-qPCR in the saliva of 13.3% of dissemination positive mosquitoes in the 25 °C condition, and this transmission was confirmed by virus isolation in 1 out of 2 RT-qPCR positive samples. No JEV transmission to saliva was detected in the gradient condition. These results suggest that JEV transmission by Culex pipiens mosquitoes upon an accidental introduction in our region is unlikely under current climatic conditions. This could change in the future when temperatures increase due to climate change.

Belgian Anopheles plumbeus Mosquitoes Are Competent for Japanese Encephalitis Virus and Readily Feed on Pigs, Suggesting a High Vectorial Capacity

“…plumbeus of up to 2 months [ 3 ]. Mortality in this study was also remarkably higher than when working with Culex pipiens using the same protocol, in a previous study [ 30 ]. A possible explanation could be that An.…”

“…For our experiments, a pre-existing eighth passage of JEV genotype 3 Nakayama strain with a titer of 10 7.7 TCID50/mL was used for our experiments. For the detailed protocol, we refer to a previous study by our team [ 30 ].…”

“…Oral infection, dissection and salivation were performed according to the standardized conditions described in [ 12 ] and applied in [ 30 ]. In short, three- to seven-day-old F0 generation An.…”

“…After 14 days (14 dpi), the surviving JEV-exposed mosquitoes were cold anesthetized for approximately two minutes, and samples were collected to determine infection, dissemination and transmission rate according to the protocol described previously [ 30 ]. Briefly, the mosquito body was collected for infection; the legs, wings and head were collected for dissemination, and saliva was collected for transmission.…”

“…RT-qPCR-positive samples were additionally subjected to virus isolation attempts on Vero cells. We refer the reader to our previous study for the detailed protocol [ 30 ].…”