Neospora caninum is a protozoan parasite that causes abortion, perinatal mortality, and subfertility in cattle worldwide. Despite the presence of the DNA of the parasite in semen of infected bulls, the effect on semen quality has not been extensively studied. This study aimed to investigate the effect of a natural Neospora caninum infection on fresh and frozen semen quality parameters in Belgian Blue bulls. Two hundred and fourteen bulls were serologically screened with an indirect ELISA-test specific for anti-Neospora caninum antibodies, every two months during one year. In addition to serological screening, semen was collected twice weekly using an artificial vagina. The following semen quality parameters were assessed: ejaculate volume, concentration, total motility of fresh semen samples, as well as morphology, total and progressive motility for frozen/thawed semen samples. Bulls were semen sampled throughout the whole year, but only semen samples of bulls that had six consecutive positive or negative ELISA-test results were included in our dataset (n = 98). Generalized linear and binomial mixed models were used for statistical analysis of each outcome variable. In these models the explanatory variables were defined as: age, barn location, mean Temperature Humidity Index (THI) during sperm production (14–42 days before sampling), maximum daily THI at collection, season of sperm production, season at collection and the Neospora caninum antibody test results. Initially, individual explanatory variables were tested in univariable models for each outcome variable. Akaike information criterion (AIC) values were used to select explanatory variables to build a multivariable model, where the Neospora caninum test result was forced in all models. The present study reveals an overall apparent seroprevalence of Neospora caninum of 9,2% in the study population. No significant associations were detected between natural neosporosis, substantiated by ELISA-antibody levels, and any of our tested outcome variables on fresh ...