2610. A Deadly Intrusion: Competitive Strain Displacement among Dengue Virus Strains in Sri lanka

Abstract Background Mosquito-borne dengue virus (DENV), the agent of dengue hemorrhagic fever (DHF), is genetically diverse, and new strains regularly invade distant locations and displace existing strains. Invasive strains often cause higher rates of DHF than displaced strains, so it is critical to identify the mechanisms that enable invasion. We tested the hypothesis that invasive strains are less susceptible to RNA interference (RNAi), the major antiviral defense in mosquitoes, than displaced strains. Methods We knocked-down (KD) RNAi in vivo in Aedes aegypti, the DENV vector, by injecting mosquitoes with double-stranded RNA against Argonaute 2 (Ago2), a key enzyme in the RNAi pathway, or a control dsRNA. Ago2 KD and control mosquitoes were fed bloodmeals containing 1 of 3 isolates each of 3 different strains of DENV that had undergone sequential competitive displacement in Sri Lanka, termed, in order of displacement, Pre-DHF, Post-DHF and Ultra-DHF. We predicted that the Pre-DHF strain, which we have previously shown to be less infectious for mosquitoes than the other two strains, would show a greater increase in infectivity than those strains. Engorged mosquitoes were incubated for 10 days, homogenized, and assayed for virus. Results Ago2 KD efficiency ranged from 79% to 98%, as determined by semi-quantitative PCR and band densitometry. The percentage of mosquitoes infected following Ago2 vs. control KD was not significantly different (33% vs. 47%; paired t-test, DF = 8, P = 0.08). However, among infected mosquitoes, virus titer was significantly higher in Ago2 KD mosquitoes (3.98 vs. 3.38 log10 plaque forming units/body; t-test, DF = 14, P = 0.02). Contra our prediction, a two-factor ANOVA did not reveal a significant interaction between the effect of virus strain and treatment (DF = 5, P = 0.58), indicating that Pre-DHF viruses did not show a larger response to Ago2 KD than Post and Ultra-DHF viruses. Conclusion These data support the role of RNAi as a key mosquito defense against virus replication in mosquitoes but indicate that the differences in competitive success among the 3 DENV strains studied are not due to differences in interactions with Ago2 during initial stages of mosquito infection. Disclosures All authors: No reported disclosures.