In vitro evaluation of a cysteine protease from poultry red mites, Demanyssus gallinae, as a vaccine antigen for chickens.

Poultry red mites (PRMs, Dermanyssus gallinae) are hematophagous ectoparasites that negatively affect egg production, which causes serious economic losses to the poultry industry worldwide. Currently, the emergence of acaricide-resistant PRMs has impeded PRM control in poultry farms. Several alternatives for acaricide use have been described for managing PRM-caused problems. Vaccination is among the methods for controlling PRMs in poultry houses. Currently, several candidates for vaccine antigens have been identified. This study identified a cysteine protease, Deg-CPR-2, which differs from 2 other previously reported cysteine proteases in PRMs, from previously obtained data from RNA-sequencing (RNA-seq) analysis. We investigated the characteristics of Deg-CPR-2 and assessed its efficacy as a vaccine antigen in vitro. Phylogenetic analysis revealed that Deg-CPR-2 belonged to a different cluster from those of other cysteine proteases in PRMs. This cluster also included cathepsin L-like proteases, enzymes thought to be involved in hemoglobin digestion in ticks. Expression analysis revealed Deg-CPR-2 expression in midguts and all the life-stages; however, there were differences in the expression levels across the life-stages. The enzyme activity of recombinant Deg-CPR-2 was inhibited in the presence of a cysteine protease inhibitor, which suggests that Deg-CPR-2 functions as a cysteine protease in PRMs. Finally, there was an in vitro increase in the mortality of PRMs, mainly protonymphs that were artificially fed with plasma from chickens immunized with Deg-CPR-2. These findings suggest that Deg-CPR-2 may contribute to protein digestion in the midgut of PRMs and is crucially involved in physiological processes in PRMs. Additionally, immunization with Deg-CPR-2 may reduce the number of protonymphs, and Deg-CPR-2 should be considered as a candidate antigen for anti-PRM vaccine development.