Flow cytometry to evaluate the level of Babesia gibsoni parasitemia in vivo and in vitro by using the fluorescent nucleic acid stain SYTO16

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In the present study, we employed flow cytometry to evaluate the level of parasitemia of Babesiagibsoni infecting canine erythrocytes in vivo and in vitro by using fluorescent nucleic acidstaining. Peripheral blood samples from a B. gibsoni-infected dog and cultured B. gibsoni parasitizingin canine erythrocytes were stained with a membrane-permeable fluorescent nucleicacid stain, SYTO16. In this study, we utilized normal canine erythrocytes (LK erythrocytes)and canine erythrocytes containing high concentrations of potassium, reduced glutathione,and some free amino acids (HK erythrocytes) as host cells for culture. Parasitized cells in vivowere discriminated completely from unparasitized cells and a correlation (r = 0.998) betweenthe percentage of SYTO16-positive cells and parasitemia in vivo was observed. On the otherhand, erythrocytes in vitro could not be divided clearly into parasitized and unparasitized cells.However, when LK erythrocytes were used as host cells, the percentage of SYTO16-positivecells was almost the same as, and was well correlated (r = 0.932) with, the level of parasitemia.When HK erythrocytes were used as host cells, the percentage of SYTO16-positive cells was almosthalf of, but was correlated (r = 0.982) with, the level of parasitemia. Therefore, we attemptedto observe the changes in the percentage of parasitized cells after treatment with antiprotozoaldrug or mitochondria inhibitors by using flow cytometry. The changes in the percentageof SYTO16-positive cells corresponded well with the changes of the level of parasitemiawhen the parasites in HK erythrocytes were cultured with each compound. The present resultssuggest that flow cytometric detection using SYTO16 is a rapid and reliable method for monitoringparasitemia both in vivo and in vitro.
The Graduate School of Veterinary Medicine, Hokkaido Universityの論文
2008-03-07

Flow cytometry to evaluate the level of Babesia gibsoni parasitemia in vivo and in vitro by using the fluorescent nucleic acid stain SYTO16

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