Assessing the performance of TRX and DUF148 antigens for detection of prepatent Guinea worm (Dracunculus medinensis) infection in dogs

Guinea worm (GW, Dracunculus medinensis) is a nematode that causes a painful and debilitating neglected tropical disease in humans. The GW Eradication Program has decreased human infections by >99% over the last 40 years. However, GW emergence in animal hosts, particularly dogs, has hampered eradication efforts. Currently, there is no method for diagnosing GW infection in animals during the prepatent period, before the adult female worms emerge. Previous works have identified two GW proteins, TRX and DUF148, as immunoreactive antigens with GW-positive human and dog sera. This study developed and validated indirect enzyme-linked immunosorbent assays (ELISA) using each antigen alone or in a combination of both antigens. Using serum samples from experimentally exposed dogs, TRX and DUF148 showed reactivity at 9- and 11-weeks post-exposure, respectively. In an experimentally infected ferret, TRX and DUF148 showed reactivity at 13- and 15-weeks post-exposure, respectively. These antigens were further validated using sera of dogs from endemic villages in Chad (n=47) and shelter dogs from the non-endemic United States (n=492). DUF148 showed better reactivity and sensitivity of 76.6.% in detecting GW infection in prepatent sera compared to TRX. However, DUF148 cross-reacted with one serum sample from Brugia pahangi experimental infection and several shelter dog sera. The anti-DUF148 titer was significantly higher in the shelter dogs positive for gastrointestinal nematodes than in negative dogs. To mitigate this cross-reaction, we produced 3 peptides of DUF148. Peptide 3 from the C-terminal was more reactive with prepatent sera and had a sensitivity of 83%; however, the specificity was not superior to DUF148 whole antigen. The antibody response to DUF148 in Chad dogs with the history of GW emergence waned overtime but was detectable until two years post-GW-emergence. Our findings could facilitate the development of diagnostic methods for early detection of GW infection in dogs in endemic countries.