Development Of Detection Methods Of Leishmania Spp And Distribution And Proliferation Of Leishmania In BALB/c Mice

Kala-azar is amphixenosis that leishmania spp infect human and animals by biting ofinsect vectors, called again leishmaniasis. Leishmania spp belong to Trypanosoma. Lots ofcases of leishmaniasis have been reported in the world, leishmaniasis is one of the tropicaldiseases which the World Health Organization strengthenly controls[1].Depending on thelocation of Leishmania parasite, leishmaniasis is divided into visceral leishmaniasis,cutaneous leishmaniasis and mucosal leishmaniasis,in which visceral leishmaniasis is themost harmful to human and animals[2].Visceral leishmaniasis is the major leishmaniasis inChina that is caused by Leishmania donovani. After the Sichuan earthquake, someresearchers have detected Leishmania parasites dogs at Wenchuan, Jiuzhaigou and Heishuiin Sichuan in2008.The result indicates that positive rate is more than20%.So Leishmaniavaccine is imminently made to prevent of leishmaniasis. Some scholars believe thatleishmania tarentolae is not harmful to animals which is isolated from the lizard[3], andtherefore can be used to make leishmaniasis vaccine. Leishmania tarentolae is easily culturedin vitro, and similar to function that protein translation, processing and modification in themammalian cell. At the same time, leishmania tarentolae have high protein expression levelsand targeting, so it is often used to research non-pathogenic parasitic insect proteinexpression system.In this experiment, Leishmania donovani and Leishmania tarentolae are selected asresearch objects. There is the different pathogenicity between leishmania donovani andleishmania tarentolae, so we have studied their morphological by Giemsa staining andnegative staining.The results show two kinds of Leishmania have no significant difference intheir morphology. Then we have injected respectively10~8/ml leishmania donovani andleishmania tarentolae via intraperitoneal way in BALB/c mice,and use indirectimmunohistochemical methods detects their distribution in mice.The results show thatleishmania donovani and leishmania tarentolae distribute in heart, liver, spleen, lungs, andkidneys in BALB/c mice. Secord, we have established real–time quantitative PCR methor,and have used it to detect proliferation of Leishmaniadonovani and leishmania tarentolae inthe liver in BALB/c mice.The results show that leishmania tarentolae’s relative growth amount is less than leisin the liver of mice less than leishmania donovani. At the same time,we have studied the difference of tissue pathology in mice infect leishmania donovani andleishmania tarentolae. The results show that: Leishmania donovani lead to rupture of themyocardial fibers, interstitial widening; Evacuation of liver cells, increase of fibers of livercell cord, some liver cell atrophy; Spleen tissues happened spotty hemorrhage and necrosis,increas of eosinophils, megakaryocytes, macrophages and lymphocytosis. Leishmaniatarentolae lead to increase of megakaryocytes in the spleen and liver in mice. In addition, wehave used fluorescent dye to stain Leishmania tarentolae that has fluorescent protein.Andleishmania tarentolae can be clearly seen in mice tissue. Real time quantitative PCR studiedthe proliferation of Leishmania tarentolae in the blood in mice. Results show thatLeishmania tarentolae is wavy proliferation in mice.This experiment verified Leishmania tarentolae is no pathogenicity to mammalian, andthis theory provides a scientific basis for the development of kala-azar vaccine, treatment ofleishmaniasis, and Leishmania tarentolae protein expression vector.