Binding of western equine encephalomyelitis virus to brush border fragments isolated from mesenteronal epithelial cells of mosquitoes

Abstract

Brush border fragments (BBF) were isolated from mesenteronal epithelial cells of mosquitoes that are either susceptible (WS Culex tarsalis) or refractory (WR Cx. tarsalis; Culex pipiens) to peroral infection by western equine encephalomyelitis (WEE) virus. The isolated BBF were combined with radiolabeled WEE virus in a binding assay to compare the amount of virus bound by BBF from susceptible and refractory mosquitoes. BBF and WEE virus were mixed in a microcentrifuge tube, incubated for 1 h and centrifuged at 27,000 × g-30 min to pellet WEE virus bound to BBF. Optimal binding occurred at pH 7.2, 20°C and there was no requirement for divalent cations. BBF isolated from perorally susceptible mosquitoes (WS Cx. tarsalis) bound significantly greater amounts of radiolabeled WEE virus, compared to BBF isolated from refractory mosquitoes (WR Cx. tarsalis; Cx. pipiens), in all experiments. The binding of WEE virus to BBF from WS Cx. tarsalis appears to be specific, based on saturation and competitive binding studies; binding to BBF from WR Cx. tarsalis and Cx. pipiens is nonspecific. Scatchard analysis of the binding data for BBF from WS Cx. tarsalis yields an estimated 1.8–3.5 × 10 6 binding sites per mesenteronal epithelial cell with an affinity constant of ( K a) of 2.2 × 10 11 M −1.