Black spot disease has been spreading over the past few years in canary flocks. The disease can be caused by various infectious agents including Circovirus, mycoplasma, and various parasites. Symptoms in neonatal canaries include black spot in the abdomen, abdominal enlargement, gall bladder congestion, death in the late period of the embryo, or the birth of the chicks carrying the virus. In adult canaries disease characterized by dullness, anorexia, lethargy and feather disorder, depression, fever, and rarely death. To conduct this study, samples collected from 10 canary flocks, each flock between 89 and 500 canaries. Specific PCR test was used for detection of canary Circovirus and Mycoplasma. Mmicrobial cultures were used for other bacterial or fungal infections. Intestinal smears were used for the detection of parasites infestations. Canary Circovirus was detected in 4 flocks. No other infections or infestations were found. According to the results of this study, canary Circovirus is a potential ethological agent for unilateral black spot on the right side of the bird body. It is a black spot due to the gall bladder congestion. This is the first report for the detection of canary Circovirus virus in Iran.