Location: Foreign Arthropod Borne Animal Disease Research
Project Number: 3022-32000-024-020-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Aug 19, 2024
End Date: Aug 18, 2025
Objective:
Transovarial vertical transmission by Aedes mosquitoes has been implicated in maintaining RVFV during the long inter-epidemic seasons. Field studies have suggested vertical transmission of RVFV in several mosquito species. In these studies, field collected eggs and larvae along with adult mosquitoes reared from collected eggs, have been positive for RVFV screening. Additionally, the detection of RVFV in field collected male mosquitoes suggests that they were infected as an egg via vertical transmission from an infected female. While field studies have suggested that vertical transmission of RVFV can occur in mosquitoes, laboratory confirmation of transovarial transmission under controlled conditions has yet to be done. To date, Culex tarsalis mosquitoes have been investigated for vertical transmission in a laboratory environment. However, the other Culex spp and Aedes aegypti of North America have yet to be studied for this phenomenon. Understanding the potential of mosquito species to vertically transmit RVFV to the next generation is needed to identify potential impact on outbreaks in endemic locations like Senegal and in naïve locations.
The objectives of this collaboration are to (1) identity additional mosquito species that can transmit RVFV through the transovarial route and (2) characterize how RVFV persists in the endemic cycle in Senegal.
Approach:
Examination of vertical transmission of RVFV will utilize Culex quinquefasciatus and Aedes aegypti mosquitoes. Both species are chosen for this study as they are natural vectors in endemic areas and potential vectors for the spread and establishment of the virus in North America. The initial studies to examine vertical transmission of RVFV, will assess the ability of RVFV to disseminate to the ovaries of Cx. quinquefasciatus and Ae. aegypti mosquitoes. The presence of RVFV in the ovaries will expose the oocyte to the virus during its development, thereby supporting the theory of transovarial transmission. Adult, female Cx. quinquefasciatus and Ae. aegypti will be provided an artificial blood meal containing RVFV or a sham. Engorged females with visible blood in the abdomen will be collected and housed for 14 days. At days 3, 7, and 14 post engorgement, the ovaries from the mosquitoes will be dissected, collected, and transferred to tubes containing cell culture media. The mosquito tissue will be homogenized, and the homogenates will undergo three blind passages in susceptible cell culture for the detection of infectious virus.
To examine transmission of RVFV to the offspring, adult, female mosquitoes will be provided an artificial blood meal as described above. Fully engorged mosquitoes will be collected and housed in a carton with a cup of water and a small paper towel for female mosquitoes to deposit egg rafts from first gonotrophic cycle (E1). The female mosquitoes will be allowed to feed two more times. All mosquitoes will be allowed to feed on non-infectious blood for one hour at day 7 and 14 post infectious blood-meal. Percent of infected mosquitoes will be calculated and engorged females will be housed as stated above yielding egg rafts from the second and third gonotrophic cycle (E2 and E3). Each egg raft will be homogenized and assayed for infectious virus. Mosquitoes will be reared from the remaining rafts and collected at different life stages. Once larvae emerge, the egg husks will be collected and assayed for infectious virus. Larvae will be pooled and assayed for infectious virus by performing blind passages as described above. The remaining larvae will be reared to adults. Saliva will be collected from the adult females and assayed for infectious virus by performing blind passages. Adult mosquitoes (male and female) will be individually homogenized and assayed for infectious virus via standard plaque assay. Determination of vertical transmission is based on the presence and absence of virus.
This project will also include a field component in Senegal to characterize how RVFV persists in the native transmission cycle. This field component will investigate the biodiversity and ecology of arbovirus vectors, screen livestock and wildlife animals (bats, rodent) as host and reservoir of arboviruses, assess vector competence of local Aedes and Culex spp. for arboviruses, and develop integrated decision support system using modeling to predict RVFV emergence risk.
