Leading anti-tick vaccine targets are variably conserved in cattle fever ticks

Rhipicephalus (Boophilus) microplus causes significant cattle production losses worldwide because it transmits Babesia bovis and B. bigemina causative agents of bovine babesiosis. Control of these ticks primarily has relied on treatment of cattle with chemical acaricides, but frequent use, exacerbated by the one-host life cycle of these ticks, has led to high-level resistance to multiple classes of acaricides. Consequently, new approaches for control, such as anti-tick vaccines, are critically important. Key to this approach is targeting highly conserved antigenic epitopes to reduce the risk of vaccine escape in heterologous tick populations. We evaluated amino acid conservation within 14 tick proteins across 167 R. microplus collected from geographically diverse locations in the Americas and Pakistan using PCR amplicon sequencing and in silico translation of exons. We found that amino acid conservation varied considerably across these proteins. Only one target, the voltage-dependent anion channel, was fully conserved in all 167 R. microplus samples (protein similarity 1.0). Five other proteins were highly conserved: the aquaporin RmAQP1 (0.989), vitellogenin receptor (0.985), serpin-1 (0.985), and subolesin (0.981). In contrast, the glycoprotein protease Bm86 was one of the least conserved (0.889). The Bm86 sequence used in the original Australian TickGARD vaccine carried many amino acid replacements compared to the R. microplus populations examined here, supporting the hypothesis that this vaccine target is not optimal for use in the Americas. By mapping amino acid replacements onto predicted 3D protein models, we also identified amino acid changes within several small peptide vaccines targeting portions of the aquaporin RmAQP2, chitinase, and Bm86. These findings emphasize the importance of thoroughly analyzing protein variation within anti-tick vaccine targets across diverse tick populations before selecting candidate vaccine antigens. When considering protein conservation alone, RmAQP1, vitellogenin receptor, serpin-1, subolesin, and especially the voltage-dependent anion channel rank as high priority anti-tick vaccine candidates for use in the Americas and perhaps globally.