Characterisation of serum IgG(T) responses to potential diagnostic antigens for equine cyathostominosis.

Abstract Cyathostomins are ubiquitous parasitic nematodes of horses. These worms spend substantial periods as intestinal wall stage encysted larvae, which can comprise up to 90% of the total burden. Several million larvae have been reported in individuals. Emergence of these larvae from the gut wall can lead to life-threatening colitis. Faecal egg count tests, increasingly used by horse owners to inform anthelmintic treatments, do not correlate with the intra-host burden of cyathostomins; this represents a key gap in the diagnostic toolbox. Previously, a cyathostomin Gut Associated Larval Antigen (Cy-GALA) was identified as a promising marker for the intra-host stages of infection. Here, Cy-GALA and an additional protein, Cyathostomin Immuno-diagnostic (Cy-CID) antigen, were investigated to examine their value in providing information on cyathostomin burden. ELISA analyses examined serum IgG(T) responses to recombinant proteins derived from individual cyathostomin species. Receiver Operator Characteristic (ROC) curve analysis was performed on the ELISA data; proteins with the highest Area Under the Curve (AUC) values were selected to test protein combinations to investigate which were the most informative in identifying the infection status of individuals. Three cocktail (CT) combinations were tested, comprising: a) Cy-GALA proteins from two species and a Cy-CID protein from a third species (CT3), b) Cy-GALA proteins from five species (CT5), and c) all CT5 components, plus a Cy-CID protein from an additional species (CT6). The best predictive values for infection were obtained using CT3 and CT6, with similar values achieved for both. Proteins in CT3 are derived from the most commonly reported species, Cyathostomum catinatum, Cylicocyclus nassatus and Cylicostephanus longibursatus. This combination was selected for future development since it represents a more commercially viable format for a diagnostic test.