西藏牛羊住肉孢子虫种类鉴别及进化分析

Species Identification and Evolution Analysis of Sarcocystis parasitize in Cattle or Sheep in Lhasa,Tibet

住肉孢子虫病严重影响畜牧业健康发展,危害肉制品和公共卫生安全。为研究西藏拉萨市临近各县牛羊住肉孢子虫流行情况及虫种鉴定,对西藏拉萨市当雄县和尼木县的牦牛、绵羊、山羊、黄牛、犏牛等共62头(只)牛羊的心肌、膈肌、腿肌、食道肌进行采样。采用压片镜检法对采集的牛羊膈肌进行观察检测,若发现住肉孢子虫包囊即判定为阳性,否则为阴性。采用PCR技术对住肉孢子虫线粒体细胞色素C氧化酶第一亚基(COX1)进行扩增,对62头(只)牛羊的心肌、膈肌、腿肌、食道肌进行检测。结果显示,PCR检测的住肉孢子虫病的阳性率与压片镜检法结果一致,藏牛羊的住肉孢子虫阳性率70.97%;对膈肌PCR产物进行克隆测序,在NCBI上进行同源性比对,共检出4个住肉孢子虫虫种,分别为肉孢子虫(Sarcocystis pilosa)、山羊犬住肉孢子虫(Sarcocystis capracanis)、羊柔嫩住肉孢子虫(Sarcocystis tenella)和家山羊犬肉孢子虫(Sarcocystis hircicanis),对应样品编号分别为5C、17C、19C和27C,其同源性分别为92.10%、99.71%、99.81%和99.26%;种系发育树结果显示,样品5C、17C、19C、27C与其NCBI上同源性最高的住肉孢子虫虫种Sarcocystis pilosa、Sarcocystis capracani、Sarcocystis tenella和Sarcocystis hircicanis处于同一分支上,且系统发育树中的Bootstrap值较高,很好地证明了研究结果中所确定的分类地位,两者确定为同一种属。另外,在牦牛身上检出了Sarcocystis pilosa,为进一步了解Sarcocystis pilosa生活史提供了线索。

The purpose of this paper was to study the prevalence and species identification of cattle and sheep Sarcocystis in neighboring counties of Lhasa city.In this study,a total of 62 domestic animals(yaks,sheep,goats,yellow cattle and dzo)in Dangxiong and Nimu counties of Lhasa city,Tibet were sampled and tested.The diaphragmatic muscles of the collected animals were observed and tested by compression microscopy,and the cysts were found to be positive,otherwise they were negative.The first subunit of cytochrome C oxidase(COX1)was amplified by PCR.The myocardium,diaphragm,leg muscle and esophagus muscle of 62 cattle and sheep were detected.The results showed that the positive rate of PCR detection was consistent with that of compression microscopy,and the positive rate of Tibetan cattle and sheep was 70.97%.Diaphragm PCR products for cloning sequencing and homology in NCBI,spore parasite species were detected 4 living meat,meat sporozoan respectively(Sarcocysti spilosa),goats,dogs live meat sporozoan(Sarcocystis capracanis)and soft sheep meat sporozoan(Sarcocystis tenella)and home dog meat goats sporozoan(Sarcocystis hircicanis),respectively corresponding to sample number 5C,17C,19C and 27C.The homology was 92.10%,99.71%,99.81%and 99.26%,respectively.The phylogenetic tree results showed that samples 5C,17C,19C and 27C were on the same branch with Sarcocystis pilosa,Sarcocystis capracani,Sarcocystis tenella and Sarcocystis hircicanis,which had the highest homology in NCBI,and the Bootstrap value in the phylogenetic tree was high,which well proved the taxonomic status determined in the research results,and the two were identified as the same genus.In addition,Sarcocystis pilosa has been detected in yaks,providing clues for scientists to further understand the life history of Sarcocystis pilosa.