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The aim of this study was to evaluate, through qPCR, the prevalence of parasitemia in sick kennel dogs naturally infected by canine leishmaniasis. An evaluation of daily changes of the parasitic load in peripheral blood was also performed. A comprehensive clinical examination and the collection of several samples (blood, lymph node, skin, and conjunctiva) were performed in 140 dogs living in an endemic area. Among these, only the dogs with clinically evident leishmaniasis were enrolled (39/140; 27.9%). Twelve (30.8%) out of 39 showed parasitemia, with a low load (median: 4 Leishmania/ml) despite a high lymph node parasite load (median: 4000 Leishmania/ml) and high IFAT titers (≥ 1:640). Seven sick dogs were sampled every 4 h for 6 times during a 24-h period, in order to obtain light- and dark-span samples. Only one (14.3%) out of the seven serial sampled dogs showed Leishmania DNA in the peripheral blood in two samples (2/42; 4.8%). Surprisingly, Leishmania DNA was also detected in the peripheral blood of asymptomatic dogs, negative to both serology and PCR performed on samples other than blood (6/101; 5.9%). The present study confirms that in canine leishmaniasis parasitemia is uncommon and even transitory. Even if recommended, microscopic examination is confirmed as a low sensitivity method with a lower diagnostic utility in canine leishmaniasis than qPCR. Moreover, circulating Leishmania DNA can be found even in healthy dogs. This finding is important in clinical practice because in endemic areas it suggests a transfusion risk and a possible transmission to the vector. [ABSTRACT FROM AUTHOR]