Morhpological And Molecular Characteristics Of Sarcocystis Species From Pigs In Yunnan Province

Sarcocysts were detected in 6 of 51 pigs(11.76%)in farm market in Kunming City and Baoshan City.Two species of Sarcocystis(Sarcocystis miescheriana and Sarcocystis suihominis)were identified based on the morphological characteristics.Sarcocysts of S.miescheriana were found in 2 of 51 pigs(3.92%),Sarcocysts of S.suihominis in 4 of 51 pigs(7.84%).Using Light microscop,sarcocysts of S.miescheriana appeared milky white color,and fusiform shape.The sarcocyst wall had numerous finger-like protrusions.Sarcocysts of S.suihominis were milky white color,and appeared fusiform shape.The sarcocyst wall had a thick hair-like protrusions.By transimission electron microscope,sarcocysts of S.miescheriana had finger-like protrusions,and a few microtubules scattered distribted within the protrusion;sarcocysts of S.suihominis had hair-like protrusions,and within the protruisosn,numorous microstubules arragend parallel,which extendied from the tip of the protrusion into the ground substances.18S rDNA,28 S rDNA,mitochondrial COX1 gene and ITS-1 gene were amplified using DNAs extracted from individual sarcocysts of each of the 2 species of Sarcocystis isolated from porks.All PCR products were purified,cloned,sequenced and analyzed.The sequences of 18 S rRNA gene for sarcocysts of S.miescheriana and S.suihominis of sarcocyst were about 1860 bp in size,and the sequences identity among the clones of S.miescheriana was 99.6%(n=5),99.2%(n=8)identiy for different clones of S.suihominis The identity between the 2 parasits was 97.2%(n=13).The sequences of 28 S rRNA gene for S.miescheriana and S.suihominis were about 3450 bp,the identities among S.miescheriana and S.suihominis were 98.6%(n=8)and 98.9%(n=8),respectively.The identity between the 2 parasites was 95.9%(n=16).The COX1 gene sequences for S.miescheriana and S.suihominis were about 710 bp,and the identities of intra-species were 99.5%(n=12)for S.miescheriana,and 99.4%(n=9)for S.suihominis.The identy between the 2 parasites was 81.1%(n=21).The sequences of ITS-I gene for S.miescheriana and S.suihominis were about 860 bp,and the identity of different clones for S.miescheriana was 97.2%(n=12),95.0%(n=12)iendity for different clones of S.suihominis.The identiy between the 2 parasites was 73.5%(n=24).Therefore,the mitochondrial COX1 gene and ITS-I gene are more effective than the 18 S rRNA and 28 S rRNA genes in distinguishing two Sarcocystis of pigs among the above four kinds of genetic markers,and they are excellent molecular markers for the molecular identification of Sarcocystis of pigs.The phylogenetic tree inferred from seqeucnes of 18 S rRNA gene,28 S rRNA gene,COX1 gene and ITS-I gene by MP,ML and NJ methods showed that S.miescheriana and S.suihominis formed two independent evolutionary branches,but have a close relationship.