Synergistic effect of silencing the expression of tick protective antigens 4D8 and Rs86 in Rhipicephalus sanguineus by RNA interference

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Abstract
Tick proteins have been shown to be useful for the development of vaccines which reduce tick infestations. Potential tick protective antigens have been identified and characterized, in part, by use of RNA interference (RNAi). RNAi allows for analysis of gene function by characterizing the impact of loss of gene expression on tick physiology. Herein, we used RNAi in Rhipicephalus sanguineus to evaluate gene functions of two tick protective antigens, 4D8 and Rs86, the homologue of Bm86, on tick infestation, feeding and oviposition. Silencing of 4D8 alone resulted in decreased tick attachment, survival, feeding and oviposition. Although the effect of Rs86 RNAi was less pronounced, silencing of this gene also reduced tick weight and oviposition. Most notably, simultaneous silencing of 4D8 and Rs86 by RNAi resulted in a synergistic effect in which tick survival, attachment, feeding, weight and oviposition were profoundly reduced. Microscopic evaluation of tick tissues revealed that guts from dual injected ticks were distended with epithelial cells sparsely distributed along the basement membrane. These results demonstrated the synergistic effect of the silencing expression of two tick protective genes. Inclusion of multiple tick protective antigens may, therefore, enhance the efficacy of tick vaccines.