Abstract
Neosporosis, caused by N. caninum, is an emerging protozoan disease responsible for significant economic losses in the global dairy and meat industries, primarily due to abortion in cattle. Dogs serve as both definitive and intermediate hosts and play a key role in the parasite transmission cycle. Currently, effective control strategies remain limited, partly due to insufficient information on infection status. In Vietnam, data on N. caninum infection are scarce and mainly limited to cattle and buffalo. In this study, an indirect enzyme-linked immunosorbent assay (iELISA) based on recombinant NcGRA4 protein was applied and evaluated for the detection of anti-N. caninum antibodies in dogs. A total of 142 shelter dogs from Hanoi, northern Vietnam, were tested to determine seroprevalence. The NcGRA4-based iELISA detected an overall seroprevalence of 28.87% (41/142), whereas the indirect fluorescent antibody test (iFAT) showed a lower prevalence of 14.08% (20/142), indicating substantial exposure to N. caninum among shelter dogs in this region. Using iFAT as the reference method, the NcGRA4-based iELISA demonstrated a sensitivity of 90.00%, a specificity of 81.15%, and an overall accuracy of 82.39%. These findings indicate that the NcGRA4-based iELISA is a suitable screening tool for seroepidemiological surveillance of N. caninum infections in dogs. Univariable and multivariable logistic regression analyses showed no significant associations between N. caninum seropositivity and the investigated variables, including age, sex, breed, and housing conditions. This study also provides the first serological evidence of canine exposure to N. caninum in Vietnam.