Dengue viruses binding proteins from Aedes aegypti and Aedes polynesiensis salivary glands

Abstract

Dengue virus (DENV), the etiological agent of dengue fever, is transmitted to the human host during blood uptake by an infective mosquito. Infection of vector salivary glands and further injection of infectious saliva into the human host are key events of the DENV transmission cycle. However, the molecular mechanisms of DENV entry into the mosquito salivary glands have not been clearly identified. Otherwise, although it was demonstrated for other vector-transmitted pathogens that insect salivary components may interact with host immune agents and impact the establishment of infection, the role of mosquito saliva on DENV infection in human has been only poorly documented. To identify salivary gland molecules which might interact with DENV at these key steps of transmission cycle, we investigated the presence of proteins able to bind DENV in salivary gland extracts (SGE) from two mosquito species. Using virus overlay protein binding assay, we detected several proteins able to bind DENV in SGE from Aedes aegypti (L.) and Aedes polynesiensis (Marks). The present findings pave the way for the identification of proteins mediating DENV attachment or entry into mosquito salivary glands, and of saliva-secreted proteins those might be bound to the virus at the earliest step of human infection. The present findings might contribute to the identification of new targets for anti-dengue strategies.

Figure 1

DENV-binding proteins from Ae aegypti salivary glands detected with anti-DENV polyclonal antibodies. Total proteins from Ae aegypti salivary gland extracts were separated on a SDS-PAGE (Electr) and transferred onto a nitrocellulose membrane. Membrane sheets were then incubated with either: DENV reference strains (DEN1 to DEN4); a semi-purified non-inoculated suckling mouse brain extract (NEG); or a DEN4 clinical isolate. Virus binding was detected using anti-DENV HMAF. Migration of the molecular weight markers and the estimated size of the DENV-binding proteins are indicated in kilodaltons (kDa), respectively on the left and on the right side of the figure.

Figure 2

DENV-binding proteins from Ae aegypti salivary glands detected with anti-E monoclonal antibodies. Total proteins from Ae aegypti salivary gland extracts were treated as described in Figure 1. After transfer, membrane sheets were incubated with either: DEN1 or DEN4 reference strains. Virus binding was then detected using anti-E Mabs. The estimated size of the DENV-binding proteins are indicated in kilodaltons (kDa) on the right side of the figure.

Figure 3

DENV-binding proteins from Ae polynesiensis salivary glands detected with anti-E monoclonal antibodies. Ae polynesiensis salivary gland extracts were treated as described for Ae aegypti in Figures 1 and 2. After the membrane sheets had been incubated with either the DEN1 or DEN4 reference strains, DENV binding was detected using anti-E type specific Mabs. Migration of the molecular weight markers and the estimated size of the DENV-binding proteins are indicated in kilodaltons (kDa), respectively on the left and on the right side of the figure.