Histamine release factor from Dermanyssus gallinae (De Geer): characterization and in vitro assessment as a protective antigen

A cDNA encoding a 174-amino-acid orthologue of a tick histamine release factor (HRF) was identified from the haematophagous poultry red mite Dermanyssus gallinae. The predicted D. gallinae HRF protein (Dg-HRF-1) sequence is highly conserved with the tick HRFs (identity 52-54%) and to a lesser degree with translationally controlled tumour proteins (TCTP) from mammals and other invertebrates (range 38-47%). Phylogenetically, Dg-HRF-1 partitions with the tick HRF clade suggesting a shared linage and potentially similar function(s). A recombinant Dg-HRF-1 protein (rDg-HRF-1) was produced and shown to induce degranulation of rat peritoneal mast cells in vitro, confirming conservation of the histamine-releasing function in D. gallinae. Polyclonal antibodies were generated in rabbits and hens to rDg-HRF-1. Western blotting demonstrated that native Dg-HRF is a soluble protein and immunohistochemical staining of mite sections revealed that the distribution of Dg-HRF, although ubiquitous, is more common in mite reproductive, digestive and synganglion tissues. A survey of hens housed continuously in a mite-infested commercial poultry unit failed to identify IgY specific for recombinant or native Dg-HRF, indicating that Dg-HRF is not exposed to the host during infestation/feeding and may therefore have potential as a vaccine using the concealed antigen approach. To test the protective capability of rDg-HRF-1, fresh heparinised chicken blood was enriched with yolk-derived anti-Dg-HRF IgY antibodies and fed to semi-starved mites using an in vitro feeding system. A statistically significant increase in mortality was shown (P = 0.004) in mites fed with anti-Dg-HRF IgY after just one blood meal. The work presented here demonstrates, to our knowledge for the first time, the feasibility of vaccinating hens with recombinant D. gallinae antigens to control mite infestation and the potential of rDg-HRF-1 as a vaccine antigen.