Morphology And Molecular Characteristics Of Sarcocystis Species From Horse And Donkey

The genus Sarcocystis(Apicomplexa)is widely parasitic in homothermal animal cells.Horses and donkeys as intermediate host,the species identification,amount and naming of Sarcocystis are still controversial.The present study aims to investigate the relationships between morpholoy and phylogeny of Sarcocystis from local horses and donkeys of Kunming,Yunnan province.Two types of sarcocyst from horse and donkey muscle respectively were identified as Sarcocystis bertrami and our study showed that the S.bertrami infection rate in 9 samples of horse was 22.2%,and the Sarcocystis asinus infection rate was 6.8%in 73 samples of donkey.Moerover,light microscopy analysis showed that there is little morphological differences between S.bertrami and S.asinus,for example,they shared the white and fusiform sarcocyst,banana-like bradyzoites and finger-like protrusion.The protrusions was 2.81 μm(n=32)and bradyzoites were 11.18×4.99 μm(n=25)of S.bertrami;and the protrusions were 1.63 μm(n=22)and bradyzoites were 14.72×4.64 μm(n=17)of S.asinus.By transmission electron microscopy,S.bertrami and S.asinus had villar protrusions and the protrusions contain microtubules,the microtubules from the protrusions extended to the ground substance.The single sarcocyst of S.bertrami and S.asinus parasitic in muscle of horse samples or donkey samples were collectied respectively and the total DNA was extracted from the sarcocyst,and then partial 18S rRNA gene,partial 28S rRNA gene and complete COI gene sequences were amplified through PCR and clone.The 18S rRNA gene of both S.bertrami and S.asinus of sarcocyst was about 1600bp,they shared high identy,99.6%(n=l5),98.2%(n=36),respectively,the identy between them was 95.1%(n=54).The size of the 28S rRNA gene of both S.bertrami and S.asinus of sarcocyst was about 1620bp,they shared identy of 98.9%(n=21),98.9%(n=28)respectively and the identy between them was 96%(n=56).The size of the COI gene of S.bertrami and S.asinus of sarcocyst was about l000bp,they shared high identy,99.1%(n=36),99.3%(n=66)respectively and the identy between them was 96%(n=108).Despite S.bertrami and S.asinus presented similar morphological characteristics,and gathered together in the phylogenetic tree with a close genetic relationship,but on the basis of this form an independent 2 groups and three gene markers separated them,suggesting that they may be different species.In conclusion,the present study is the first to compare and analyze the molecular markers of Sarcocystis in horses and donkeys,and our study showed that they may be different species based on these datas.