This study performed clinical, serological and parasitological assessments in dogs from Vale do Rio Doce, in Minas Gerais State, Brazil, a region considered as a 'controlled endemic' area for canine visceral leishmaniosis (CVL). Nevertheless, there are signs that CVL in dogs may be re-emerging as a programme to control the disease was interrupted in the 1990s. The majority of the animals examined presented various symptoms associated with CVL. The enzyme-linked immunosorbent assay test indicated 13.7 and 12.4% of positivity of dogs from the urban and rural areas, respectively. According to indirect immunofluorescence assay test and TRALd tests, 18.2 and 42.2% of dogs in the rural area were seropositive, respectively. Parasitism in seropositive dogs was confirmed by in vitro tissue culture. Sand flies of the genus Lutzomyia, which are able to transmit both cutaneous and visceral leishmaniosis, were found in the area. The results provide a strong evidence of the re-emergence of CVL in this region.