Equine abortion, stillbirth and neonatal death cause major economic losses to the equine industry worldwide. Both non-infectious and a wide range of infectious causes have been described. However, the relative contribution of pathogens to equine abortion, stillbirth and neonatal death is poorly documented, since available studies involve only a limited number of pathogens. Therefore, the objectives of the present retrospective monitoring study were to determine the prevalence of infectious agents associated with equine abortion and perinatal mortality in Belgium, and to set up a protocol usable under field conditions using polymerase chain reaction targeting. A real-time simple polymerase chain reaction for eight different abortifacient pathogens was conducted leading to the detection of at least one infectious agent in 37% of 105 analyzed cases. In the diagnosed cases, equine herpesvirus-1 was the most detected pathogen (49%), followed by Streptococcus equi subspecies zooepidemicus (28%), Coxiella burnetii (18%), Leptospira interrogans (3%) and Neospora caninum (3%). None of the analyzed cases was positive for equine viral arteritis, equine herpesvirus-4 and Chlamydophila spp. In this study, PCR targeting is shown to have its value to detect a (co)-infectious cause in equine abortion, stillbirth and neonatal death, especially in field conditions where autolysis and contamination might preclude a full post-mortem examination protocol based on classical microbiological examination.