Comparison of Skin Prick Tests (SPT), Intradermal Tests (IDT) and In Vitro Tests in the Characterization of Insect Bite Hypersensitivity (IBH) in a Population of Lusitano Horses: Contribution for Future Implementation of SPT in IBH Diagnosis

SIMPLE SUMMARY: Insect Bite Hypersensitivity (IBH) prevalence in Portugal and in Lusitano horses is not known, but environmental characteristics are favorable to the activity of Culicoides and common knowledge shows high occurrence in Lusitano stud farms. This study aimed to compare skin tests and in vitro allergy tests for the diagnosis of IBH in Lusitano horses. The results of our study showed that skin prick tests presented a higher discriminatory diagnostic potential in IBH diagnosis. This increases our knowledge about IBH in Lusitano horses and could represent a step forward in the future development of specific immunotherapy. ABSTRACT: Thirty controls (C) and 30 IBH-affected (T) Lusitano horses were evaluated. T horses were included based on anamnesis and physical examination, supported by questionnaires. All horses were submitted to skin tests, Intrademal (IDT) and Skin Prick Tests (SPT), on the neck with 14 specific allergens, 13 recombinant proteins (r-proteins) from Culicoides nubeculosus (Cul n) and Culicoides obsoletus (Cul o) salivary glands and Culicoides nubeculosus Whole Body Extract (Cul n WBE). Addicionally, a cluster of six T and six C horses were also tested with Cul n 3 and Cul n 4 produced in insect cells and barley, as well as E. coli produced Cul o 3 and Cul o WBE. Allergen concentrations were 10 µg/mL for IDT and 100 µg/mL for SPT, and wheal diameters assessed at 20 min, 6 and 48 h. IDTs were considered positive when wheal diameter was ≥50% of the histamine wheal and SPT’s ≥ 0.9 cm. In vitro tests, allergen-specific serum IgE and sulfidoleukotriene (sLT) release assay were also carried out. Results showed that Cul n WBE, Cul n 7, 8, 9, Cul o1P and Cul o 2P were the best performing allergens for SPTs (p ≤ 0.0001) for the 1st allergen panel and Cul o WBE, Cul n 3 Bar and Cul n 4 Bac (p ≤ 0.05) for the 2nd, presenting a higher discriminatory diagnostic potential than IDTs, at a concentration of 100 µg/mL, with readings assessed at 20 min. Regarding in vitro tests overall, the sLT release assay performed best.