The equine parvovirus-hepatitis (EqPV-H), recently identified in association with serum hepatitis in horses (also known as Theiler's disease), has been so far described in horses from North America, Asia and Europe. There is no information regarding its circulation in South America. Our retrospective study (2013)(2014)(2015)(2016) screened by EqPV-H nested-PCR a total of 96 Brazilian horses grouped according to previous status of infection: Known to be positive for one or more horse "hepatitis viruses" (equine hepacivirus, equine pegivirus-EPgV and Theiler's disease-associated virus) and known to be negative. Serum biochemical parameters (aspartate aminotransferase, gammaglutamyl transferase and glutamate dehydrogenase) were evaluated in EqPV-H positive horses. Molecular characteristics of the isolates were analyzed by DNA sequencing and phylogenetic analysis. EqPV-H DNA was detected in 12.5% (12/96) of horses from 46.6% (7/15) of the farms evaluated. Similar results were obtained between coinfected group (12.3%, 7/57) and non-coinfected group (12.8%, 5/39). Coinfection with EPgV was the most frequent (5/7). Altered serum biochemical parameters suggested a subclinical hepatopathy in some animals (3/12), but the majority presented no clinical or laboratory signs of infection. Nucleotide identity was higher than 94% in comparison with previous isolates. In conclusion, we demonstrated, for the first time in South America, the circulation of EqPV-H. The Brazilian isolates presented a low genetic variability, thus corroborating previous evidence.