Development of an indirect ELISA test using an affinity purified surface antigen (P38) for sero-diagnosis of canine Neospora caninum infection

Neospora caninum is an intracellular protozoan that infects domestic and wild canids, as well as many warm blooded animals as shown by the isolation of viable parasites and the detection of parasite DNA in naturally infected hosts. N. caninum is described as a cause of neuromuscular disease and death in dogs. The dog is also known as a definitive host, shedding oocysts involved in the transmission of the infection to intermediate hosts. This study was conducted to develop an indirect ELISA test using an affinity purified 38 kDa N. caninum surface antigen (P38) for the sensitive and specific diagnosis of this infection in dog populations. To define a suitable cut-off, serum samples from 233 dogs were analyzed using an N. caninum-specific indirect fluorescent antibody test. All of these serum samples were subjected to the newly designed P38-ELISA. The Two-graph Receiver Operating Characteristics (TG-ROC) of the serum ELISA was determined to examine the performance of the test. TG-ROC analysis showed an area under curve (AUC) of 0.996 that indicates the test results being highly accurate. Optimal sensitivity and specificity (100% and 97.9%, respectively) were determined for SI n cut-off point of 0.23. To examine possible cross-reactions with other parasites affecting dogs, sera of dogs positive for antibodies against Toxoplasma gondii ( n = 17) and Leishmania infantum ( n = 11) infections were tested. These revealed negative results when tested in the new ELISA.