Simulated overwintering of encephalitis viruses in diapausing female Culex tarsalis (Diptera: Culicidae)

Female Culex tarsalis Coquillett in reproductive diapause were infected per os or by intrathoracic inoculation with western equine encephalomyelitis (WEE) or St. Louis encephalitis (SLE) viruses during "fall," maintained over a simulated "winter," and then tested for virus infection and transmission in vitro and in vivo after "vernal" termination. Exposure of F1 progeny of field-collected females to cool temperatures and short daylength produced females in reproductive diapause that were reluctant to imbibe infectious virus from pledgets soaked with suspensions of virus, blood and sucrose (2.5% by volume). Those infected per os maintained virus at very low or undetectable titers. Some females that originally tested negative for WEE by plaque assay on Vero cell culture tested positive by reverse transcriptase-polymerase chain reaction (RT-PCR) and by Vero cell culture after passage in mosquito cells. Few females became infected orally with SLE, but these infected females developed elevated titers. Females inoculated with SLE retained their infection through winter and then transmitted readily in vitro and in vivo. Feeding on a vertebrate host after diapause termination significantly increased the titer of SLE in previously infected females. These experiments simulated how infections acquired either horizontally or vertically may provide mechanisms for WEE and SLE overwintering. Attempts to detect infected females during winter following a summer with enzootic WEE activity were negative by both RT-PCR and plaque assay.