Acetylcholinesterase of Rhipicephalus (Boophilus) microplus and Phlebotomus papatasi: Gene identification, expression, and biochemical properties of recombinant proteins

The southern cattle tick Rhipicephalus (Boophilus) microplus (Bm) is a vector of bovine babesiosis and anaplasmosis. Tick resistance to organophosphate (OP) acaricide involves acetylcholinesterase (AChE) insensitivity to OP and metabolic detoxification. Sequencing and in vitro expression of Bm genes encoding AChE allowed biochemical characterization of three BmAChEs expressed in tick synganglion. rBmAChE1, rBmAChE2 and rBmAChE3 exhibited substrate preference for acetylthiocholine, high substrate inhibition and sensitivity to AChE-specific inhibitors. OP-insensitivity mutations were demonstrated in rBmAChE1 and rBmAChE3. Gene silencing suggested functional complementation of BmAChEs in vivo. BmAChE genes were amplified in copy number and multiple transcript polymorphisms were expressed in individual tick synganglia for each of the BmAChEs, suggesting allelic diversity within individuals. Studies also identified a gene encoding AChE of the sand fly, Phlebotomus papatasi, a vector of leishmaniasis in humans and animals. Expression of recombinant P. papatasi AChE (rPpAChE) enabled biochemical characterization and identification of effective inhibitors that selectively target rPpAChE.

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► Recombinant tick and sand fly acetylcholinesterases were expressed in baculovirus.
► Tick AChE mutations associating with organophosphate resistance were identified.
► rBmAChE1 mutant constructs were insensitive to coumaphos oxon in vitro.
► Phenotypic resistance to organophosphate is highly complex and multigenic.