Detection of Neospora caninum in the semen and blood of naturally infected bulls

Abstract

A prospective study was designed to investigate the presence of Neospora caninum in semen and blood of eight bulls seropositive to N. caninum using nested-PCR procedures. Positive semen and blood samples were bioassayed in a BALB/c nu/ nu mouse model. Specific anti- N. caninum serological and interferon-gamma (IFN-γ) responses were also studied. In parallel, five seronegative bulls acted as non-infected controls. All bulls were located in a collaborating AI centre and monitored for 22 weeks. Six of eight seropositive bulls showed N. caninum DNA in their semen and/or blood samples at some time during the course of the study. In all positive semen samples, we consistently found Neospora-DNA in the cell fraction and not in seminal plasma. Parasite load, as determined by a real-time PCR in nested-PCR positive semen samples, ranged from 1 to 10 parasites/ml. We found no association between the presence of N. caninum DNA in semen and blood. N. caninum could not be detected in the BALB/c nu/ nu mice inoculated with PCR-positive semen or blood samples. Specific IgG antibody levels in seropositive bulls fluctuated over time, at times falling below cut-off level. The response was predominantly IgG2, with significant differences compared to control bulls ( P < 0.05). The overall mean specific IFN-γ response in seropositive bulls was also higher than those observed in the control group ( P < 0.05), although extensive variation in individual responses was observed among bulls and over time. No significant association was found between bulls showing Neospora DNA in semen, blood, or both, and specific IgG, IgG1, IgG2, IgM and IgA levels or IFN-γ response. This study is the first to report the presence of Neospora DNA in semen and blood of naturally-infected bulls. Our observations indicate intermittent presence of N. caninum in blood and semen and shedding in semen in low numbers.