Abstract
Parasitic sea lice represent a major sanitary threat to marine salmonid aquaculture, an industry accounting for 7% of world fish production. Caligus rogercresseyi is the principal sea louse species infesting farmed salmon and trout in the southern hemisphere. Most effective control of Caligus has been obtained with macrocyclic lactones (MLs) ivermectin and emamectin. These drugs target glutamate-gated chloride channels (GluCl) and act as irreversible non-competitive agonists causing neuronal inhibition, paralysis and death of the parasite. Here we report the cloning of a full-length CrGluClα receptor from Caligus rogercresseyi. Expression in Xenopus oocytes and electrophysiological assays show that CrGluClα is activated by glutamate and mediates chloride currents blocked by the ligand-gated anion channel inhibitor picrotoxin. Both ivermectin and emamectin activate CrGluClα in the absence of glutamate. The effects are irreversible and occur with an EC50 value of around 200 nM, being cooperative (nH = 2) for ivermectin but not for emamectin. Using the three-dimensional structure of a GluClα from Caenorabditis elegans, the only available for any eukaryotic ligand-gated anion channel, we have constructed a homology model for CrGluClα. Docking and molecular dynamics calculations reveal the way in which ivermectin and emamectin interact with CrGluClα. Both drugs intercalate between transmembrane domains M1 and M3 of neighbouring subunits of a pentameric structure. The structure displays three H-bonds involved in this interaction, but despite similarity in structure only of two these are conserved from the C. elegans crystal binding site. Our data strongly suggest that CrGluClα is an important target for avermectins used in the treatment of sea louse infestation in farmed salmonids and open the way for ascertaining a possible mechanism of increasing resistance to MLs in aquaculture industry. Molecular modeling could help in the design of new, more efficient drugs whilst functional expression of the receptor allows a first stage of testing of their efficacy.
Highlights
Sea lice are marine ectoparasite copepods of the Caligidae family that attach to host marine fish and feed on their epidermal tissue, and blood
Caligus rogercresseyi is the principal sea louse species infesting farmed fish in the southern hemisphere. Successful control of these parasites has been achieved using macrocyclic lactones (MLs), but resistance has emerged over time
Our molecular modeling of the protein in complex with the drugs suggests a novel way in which ivermectin and emamectin exert their effects on CrGluCl due to a lack of conservation at interaction sites identified in the crystal structure of the receptor from C. elegans
Summary
Sea lice are marine ectoparasite copepods of the Caligidae family (order Siphonostomatoida) that attach to host marine fish and feed on their epidermal tissue, and blood. The Caligidae family is constituted by hundreds of species belonging to the Caligus and Lepeoptherius genera. Increasing interest in these parasites has arisen owing to the ravages they produce on fish aquaculture leading to morbidity and mortality with extremely high economic impact in the industry. A different sea louse, Caligus rogercresseyi is the most important parasite affecting Atlantic salmon and rainbow trout sea water farming in Chile. This Caligus was described as a separate species only in the year 2000 [2] and much remains to be known about its biology. As in the Northern hemisphere, Caligus rogercresseyi infestation within the nationally important Chilean aquaculture industry is associated with increased costs and decreased productivity with high social impact [Alvial et al cited in 3]
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