利用反向线状印迹及高通量技术分别检测蜱体内梨形虫及全沟硬蜱体内细菌研究

发布时间：2018-03-23 05:00

  本文选题：梨形虫　切入点：反向线状印迹　出处：《中国农业科学院》2016年博士论文　论文类型：学位论文

【摘要】：梨形虫病是由顶复门、梨形虫目,巴贝斯属和泰勒属的宿主特异性血液原虫引起的原虫病的总称,巴贝斯属和泰勒属分别引起巴贝斯虫病和泰勒虫病。蜱是一类有害的吸血外寄生虫,是梨形虫病的主要传播媒介。蜱可以通过直接的接触和吸血过程,或者通过传播细菌,原生动物或病毒病对宿主造成寄生性损伤。在热带和亚热带地区,梨形虫病可导致家畜和野生动物的临床障碍。这种疾病造成的经济影响包括:肉类和奶制品产量降低,流产,生育能力降低,较高的预防与治疗成本,以及对全球动物和动物制品行业造成数十亿美元的经济损失。本文的研究结果包括两个方面:1.利用建立的反向线状印迹杂交技术对采自中国不同省份草地或动物体表的不同蜱种所携带的梨形虫虫种进行调查,2015年三月至六月对样品进行收集。结果表明,RLB检测方法具有高度的特异性,可以区分不同的梨形虫虫种,通过构建的重组质粒,计算其敏感性可达到102拷贝/μL。草地上收集的蜱样品可检测到单一或混合感染的情况,环形泰勒虫(33/408,8.1%)和新疆巴贝斯虫未定种(30/408,7.4%)的检出率最高。利用建立的RLB检测方法,对动物体表收集的蜱样品评估传播牛羊巴贝斯虫和泰勒虫虫种的潜在威胁。结果显示单一的环形泰勒虫(118/652,18.1%)感染和吕氏泰勒虫+中华泰勒虫(18/652,2.8%)混合感染的情况最为常见。许多蜱种所携带病原范围比以前报道的更广。2.利用建立的反向线状印迹杂交技术对采自我国黑龙江省的25只全沟硬蜱携带梨形虫种进行调查,同时在16S rRNA基因V4高变区域的基础上,利用高通量测序法对25只全沟硬蜱的细菌种类分布进行分析研究。RLB检测结果显示,所检测的全沟硬蜱没有发现感染或携带任何泰勒虫或巴贝斯虫虫种。通过全沟硬蜱样品的高通量测序显示,检测到200多个细菌属,其中一些对动物或人类具有潜在的致病重要性,一些细菌属可对环境造成污染。根据测序结果,我们发现了立克次氏体和伯氏疏螺旋体这些对人和动物具有致病性的蜱传细菌病原。综上所述,所建立的特异性RLB检测方法可以同时检测和鉴别不同巴贝斯虫和泰勒虫,该方法可作为一种潜在的诊断工具。同时,兽医从业人员应该对环形泰勒虫在中国的广泛流行提高意识,蜱对不同梨形虫种的传播能力需要通过动物传播实验进一步验证。同时应该对我国黑龙江省分布的携带多种病原体的全沟硬蜱对人及动物叮咬引发的疾病的威胁提高意识。
[Abstract]:Pyridiasis is a disease of host specific blood protozoa caused by the phylum, Pyridiidae, Babes and Taylor genera. Babes and Taylor cause Babes's disease and Taylor's disease respectively. Ticks are a kind of harmful extranasal parasites, which are the main vectors of piridiosis. Ticks can be transmitted by direct contact and bloodsucking, or by transmission of bacteria. Parasite damage to hosts caused by protozoa or viral diseases. In tropical and subtropical regions, pear worm disease can cause clinical disorders in domestic and wild animals. The economic effects of the disease include reduced meat and dairy production and miscarriages. Lower fertility, higher cost of prevention and treatment, The research results of this paper include two aspects: 1. Using reverse linear blotting technique to analyze grassland or animals collected from different provinces of china. Pyriform species carried by different ticks on the body surface were investigated, and samples were collected from March to June 2015. The results showed that the RLB assay was highly specific. Different piriform species can be distinguished. The sensitivity of the recombinant plasmid can reach 102 copies / 渭 L. single or mixed infection can be detected in ticks collected on grassland. The detectable rates of 33 / 4088.1th of the circular Taylor worm and 30 / 4087.4of the undetermined species of Babes in Xinjiang were the highest. Using the established RLB detection method, Ticks collected from animal surfaces were used to assess the potential threat to the transmission of cattle, sheep, Babes and Taylor species. The results showed that a single circular Taylor worm 118 / 652 / 18.1) and a mixed infection of Taylor lui's 18 / 652 / 2. 8) were the most common. Common. Many ticks carry a wider range of pathogens than previously reported. 2. Using the established reverse linear blotting technique to investigate 25 Pyriformis species collected from Heilongjiang Province, China. At the same time, based on the high variable region of 16s rRNA gene V4, the species distribution of 25 ticks were analyzed by high-throughput sequencing method. None of the tested ticks were infected or carried any Taylor or Babes species. Through high-throughput sequencing of the samples, more than 200 bacterial genera were detected. Some of them are potentially pathogenic to animals or humans, and some bacteria can cause environmental pollution. We have found that Rickettsiella burgdorferi and Borrelia burgdorferi, which are pathogenic to human and animal ticks, have been found. In conclusion, the established specific RLB assay can detect and differentiate different Babes and Taylor parasites at the same time. This method can be used as a potential diagnostic tool. At the same time, veterinary practitioners should raise awareness of the widespread prevalence of the ringworm in China. The transmission ability of ticks to different piriform species should be further verified by animal transmission experiments. Meanwhile, the awareness of the threat to human and animal bites caused by the distribution of various pathogens in Heilongjiang Province should be raised.
【学位授予单位】：中国农业科学院
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S852.7
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本文编号：1651974