新疆南疆图兰扇头蜱鉴定及其携带部分病原检测

发布时间：2018-01-04 03:36

本文关键词：新疆南疆图兰扇头蜱鉴定及其携带部分病原检测　出处：《塔里木大学》2017年硕士论文　论文类型：学位论文

【摘要】：图兰扇头蜱(Rhipicephalus turanicus)是Pomerantzev于1940年命名的蜱种,研究发现该蜱是立克次体、无浆体、莱姆病、巴贝虫和森林脑炎病毒的传播媒介,这些病原给畜牧业的健康发展带来了极大的损害。为了对新疆南疆图兰扇头蜱进行鉴定并对其可能传播携带的病原进行检测。本试验对新疆南疆采集的疑似扇头蜱属蜱虫,先通过传统的形态学分类学方法对采集的蜱进行分类鉴定,选取初步鉴定的31个采集点扇头蜱属蜱为研究对象,进一步选取蜱线粒体12S rDNA和16S rDNA基因扩增特异性的基因片段,最终通过传统形态学分类与现代分子生物学方法相结合对图兰扇头蜱进行鉴定。为了对该蜱携带的病原进行检测,利用聚合酶链式反应(PCR)技术,对图兰扇头蜱立克次体omp A、omp B、g1tA和scal基因、嗜吞噬细胞无浆msp4基因、绵羊无浆体线粒体16S rDNA基因,巴贝斯虫线粒体18S rDNA基因、宫本疏螺旋体16S rDNA、布鲁氏菌线粒体16S rDNA基因进行检测,将PCR检测阳性产物进行测序,测序结果在NCBI/BLAST比对平台进行比对,通过同源性比对和系统进化关系的分析来确定病原的基因型。结果显示本次试验共采集扇头蜱属蜱1144只,通过形态学与分子生物学相结合的鉴定方法,经鉴定所有的蜱均为图兰扇头蜱,存在有尾凸或无尾凸形态学差异。蜱传病原检测发现,立克次体OmpA基因阳性率为16.1%(15/93)、立克次体gltA基因阳性率为12.9%(12/93)、立克次体sca1基因阳性率为16.1%(15/93)、立克次体OmpB基因阳性率为16.1%(15/93),采用MEGA5.0软件使用邻接法构建系统进化树,结果显示:存在三种立克次体Candidatus Rickettsia barbariae、Rickettsia massiliae和Rickettsia africae,系统进化关系显示Candidatus Rickettsia barbariae构成一单独分支,既不属于斑点热群(SFG)又不属于斑疹伤寒群(TG),其中Rickettsia africae为新疆南疆首次报道;无浆体检总阳性率为29.03%,其中绵羊无浆体的阳性率为9.68%(9/93),噬吞噬细胞无浆体的阳性率为19.35%(18/93);巴贝斯虫的阳性率为9.68%(9/93);宫本疏螺旋体阳性率为3.32%(3/93),该病原也为新疆南疆首次报道;布鲁氏菌的检测中未检测到阳性。本研究利用传统的物种鉴定与分子生物学相结合的方法,对新疆南疆图兰扇头蜱进行了准确的物种分类鉴定;对其携带立克次体、无浆体、巴贝斯虫、宫本疏螺旋体和布鲁氏菌病原进行核酸PCR检测,从分子生物学及分子流行病学角度探讨这些病原体在蜱体内存在及流行情况,为科学防御蜱传病提供依据。
[Abstract]:Turan Rhipicephalus (Rhipicephalus turanicus Pomerantzev) is a tick species named in 1940, the study found that this tick is Rickettsia Anaplasma, Lyme disease, BABEI, insect and tick borne encephalitis virus media, these pathogens to the healthy development of animal husbandry has brought great harm to South Xinjiang. Turan Rhipicephalus were the identification and the possible spread of the pathogen carrying were detected. The test of the southern Xinjiang collection of suspected tick Rhipicephalus genera, morphological first through the traditional taxonomy method for identification of collected ticks, select 31 collection points preliminary identification Rhipicephalus genera ticks as the research object, further selected gene fragment tick rDNA and 16S mitochondrial 12S rDNA gene amplification specificity, the traditional morphological classification and modern molecular biology methods combined with the identification of Turan Rhipicephalus. In order to carry on the tick The pathogen was detected by polymerase chain reaction (PCR) technique, the Turan Rhipicephalus Rickettsia OMP A, OMP B, g1tA and scal genes, up a cell no pulp MSP4 gene of Anaplasma ovis mitochondrial 16S rDNA gene, rDNA gene, insect mitochondrial 18S BABEI, Miyamoto 16S of Borrelia rDNA, detection the mitochondrial 16S rDNA gene of Brucella, the detection of PCR positive products were sequenced, the sequencing results were compared in the NCBI/BLAST on the platform, through the analysis of homology and phylogenetic relationships to determine the genotype of the pathogen. The result shows that this test were collected 1144 ticks Rhipicephalus genera, identified through the combination of morphology and molecular biology the identified all ticks were Tulan Rhipicephalus has no tail tail convex or convex morphological differences. Detection of tick borne pathogens, the positive rate of OmpA gene of Rickettsia was 16.1% (15/93), Patrick Once the gltA gene positive rate was 12.9% (12/93), the positive rate of SCA1 gene of Rickettsia was 16.1% (15/93), the positive rate of OmpB gene of Rickettsia was 16.1% (15/93), using MEGA5.0 software, the phylogenetic tree constructed using the neighbor joining method showed that there are three kinds of Rickettsia barbariae Rickettsia massiliae encoding the Candidatus, Rickettsia and africae, system evolution the relationship between Candidatus Rickettsia barbariae constitute a separate branch, does not belong to the spotted fever group (SFG) and does not belong to the typhus group (TG), where Rickettsia africae is reported for the first time in South Xinjiang; total positive rate of slurry examination was 29.03%, the positive rate of Anaplasma ovis was 9.68% (9/93), positive rate paste for 19.35% macrophage phagocytic cells (18/93); the positive rate was 9.68%, from BABEI (9/93); the positive rate was 3.32% Miyamoto sparse spiral (3/93), the pathogen is reported for the first time in South Xinjiang; The detection of Brucella was not detected in this study. The positive method of species identification and molecular biology combined with the traditional, in the south area of Xinjiang Turan Rhipicephalus for exact identification and classification of species; carrying Rickettsia, Anaplasma, Babes worm, Miyamoto sparse spiral body and nucleic acid detection of Brucella PCR. To investigate these pathogens in ticks and epidemic situation from the perspective of molecular biology and molecular epidemiology of tick borne diseases, to provide the basis for scientific defense.

【学位授予单位】：塔里木大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S852.7

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