Detection of Leishmania infantum in Dogs by PCR with Lymph Node Aspirates and Blood

The PCR technique was applied to the diagnosis of leishmaniasis in dogs, both serologically negative and positive. DNA was taken from lymph node aspirates and blood. The primers 13a and 13b, derived from Leishmania amazonies and Leishmania braziliensis kinetoplast DNA (kDNA), also amplified Leishmania infantum IPT1 constant region of minicircle kDNA. The amplified fragment is 116 bp long. It was cloned and the sequence was determined. A 70-bp inner fragment was designed and used as a probe in dot blot hybridization. A group of 124 dogs was examined, 37 of which showed typical symptoms of disease. PCR was performed on 124 blood samples and 52 lymph node aspirates. Using microscopic examination as the "gold standard," we calculated sensitivity, specificity, and positive and negative predictive values of 100% using lymph node aspirates and values of 85, 80, 95, and 57%, respectively, using blood samples. We found that 40% of the animals without lesions and 38% of the animals with clinical signs gave false-negative results by indirect immunofluorescence antibody testing. These animals could contribute to the spreading of infection among dogs, and represent a potential risk for human health as well.