贵州矮马主要传染性疾病的血清学和病原学研究

发布时间：2017-12-27 18:09

本文关键词：贵州矮马主要传染性疾病的血清学和病原学研究　出处：《贵州大学》2016年硕士论文　论文类型：学位论文

【摘要】：贵州矮马是目前全球仅存的少数天然矮马品种之一,是在自然生态环境和人工选择的双重影响下形成的珍贵遗传资源,主要分布在黔西南一带的偏远山区,其中以安顺市紫云县火花乡的矮马最为著名。2013年间,紫云县的马匹出现大量死亡现象,尤其是以懂桑村养殖的贵州矮马死亡率最高,发病率70.1%,死亡率60%,死亡病因不明。针对病马临床症状的分析,怀疑可能是马泰勒虫(Theileria equi)、马驽巴贝斯虫(Babesia caballi)、马传染性贫血病毒(Equine Infectious Anemia virus,EIAV)、马动脉炎病毒(Equine arteritis virus,EAV)或者马疱疹病毒I型(Equine herpesvirus type-1,EHV-1)所引发的传染性疾病。本研究采用血液涂片镜检、酶联免疫吸附实验(Enzyme-linked immunosorbent assay,ELISA)和聚合酶链式反应(Polymerase Chain Reaction,PCR)的方法对可疑的病原进行血清学和分子病原学研究,以期找到贵州矮马大量死亡的真正病因。采用竞争性酶联免疫吸附试验(cELISA)和PCR的方法对马泰勒虫病和马驽巴贝斯虫病的血清学和分子病原学进行研究,结果表明,cELISA方法检测贵州矮马、西南马和伊犁马血浆样品中马泰勒虫抗体阳性率分别为23.86%、11.43%、31.71%,而只有伊犁马样品中检测到马驽巴贝斯虫抗体,阳性率为85.37%。三个马群中均检测到马泰勒虫和马驽巴贝斯虫的18s rRNA基因,贵州矮马、西南马和伊犁马血液样品中马泰勒虫18s rRNA基因的检出率分别为78.41%、62.86%、68.29%,而马驽巴贝斯虫的18s rRNA基因检出率分别为11.36%、11.43%、90.24%。同时,对新鲜血液样品制作血液涂片,经显微镜观察,发现马泰勒虫和马驽巴贝斯虫虫体的存在,其大小及形态特征与文献中的描述相符。PCR和cELISA两种方法的检测结果存在一定的差异,PCR的阳性率普遍高于cELISA的阳性率,这可能是由于PCR方法检测灵敏度较高,另外,可能是由于采样时部分马匹正处于感染的早期阶段,还没有抗体的产生。有些样品的cELISA检测结果为阳性,但PCR方法检测却为阴性,这可能是由于马匹体内的马泰勒虫虫体已被自身的吞噬系统消除,但其抗体还会在体内存在一段时间。有部分样品cELISA检测结果为阴性,但PCR方法检测却为阳性,可能是因为寄主刚被感染,或者是在抗体消失后又出现了新的感染。以贵州矮马、西南马和伊犁马的血清为材料,采用ELISA的方法对EIAV、EAV和EHV-1进行血清学研究,3个马群中均未检测到EIAV抗体阳性样品;EAV的阳性率为48.08%、22.86%、10.81%;EHV-1的阳性率为99.38%、97.14%、100%。对EHV-1的DNA进行PCR扩增,贵州矮马、西南马和伊犁马EHV-1的检出率分别为94.23%、80%、83.78%。由于马传贫与马泰勒虫病的临床症状比较相似,所以EIAV抗体阴性的结果为马泰勒虫病的确诊提供了理论依据。EHV-1的阳性率很高,但是其所导致的疾病死亡率并不高,可能一种继发性感染。贵州矮马EAV血清学阳性率较西南马和伊犁马要高,而且马病毒性动脉炎的死亡率较高,表明EAV可能是导致贵州矮马大量死亡的病原之一。研究结果表明,马泰勒虫病和马病毒性动脉炎可能是导致贵州矮马大量死亡的主要原因。对尚存活的患病马匹根据其临床症状施以伊维菌素、血虫迪以及附红二合一药物进行及时的治疗,并同时施用抗生素药物对继发性感染进行治疗,直至马匹痊愈,最终有效的降低了当地马群的死亡率。综上,本研究应用显微镜、ELISA和PCR的方法对贵州矮马主要传染性疾病的血清学和病原学的研究,不仅减少了当地农户的经济损失,避免了贵州矮马这一珍贵品种即将灭绝噩梦的发生,更是为当地马属动物疾病的诊断、治疗和预防提供了可靠的理论依据。
[Abstract]:Guizhou pony pony is one of the few natural varieties currently the world's only, is the precious genetic resources formed in the double influence of natural ecological environment and artificial selection in the remote mountainous areas are mainly distributed in the southwest area, Ziyun County of Anshun City, which sparks township is the most famous pony. 2013 years, Ziyun County horses appear a large number of death phenomenon, especially to understand the Guizhou pony village of mulberry breeding the highest mortality rate, the incidence rate was 70.1%. The mortality rate was 60%, the death of unknown etiology. According to the analysis of the clinical symptoms of the disease of horses, is suspected of theileriaequi (Theileria equi), BABEI, Ma nu (Babesia, caballi) from equine infectious anemia virus (Equine Infectious Anemia virus, EIAV), equine arteritis virus (Equine arteritis virus, EAV) or equine herpes virus type I (Equine herpesvirus type-1, EHV-1) caused by infectious diseases. This study used blood smear microscopy, enzyme-linked immunosorbent assay (Enzyme-linked immunosorbent, assay, ELISA) and polymerase chain reaction (Polymerase Chain Reaction, PCR) method of serological and molecular pathogenic suspicious pathogen research, in order to find the real cause of death of a large number of Guizhou pony. By competitive enzyme-linked immunosorbent assay (cELISA) for Martel and BABEI, jade horse worm disease worm disease serology and molecular etiology research methods and results of PCR showed that the cELISA method for the detection of Guizhou pony, southwest Martel horse and Yili horse in plasma samples of antibody positive rate were 23.86%, 11.43%, 31.71% but, only Yili horse was detected in the samples of BABEI, Ma Nu antibody positive rate was 85.37%. Three horses were detected in 18S rRNA gene of theileriaequi horse and jade BABEI worm, Guizhou pony, southwest Mataile horse and Yili horse blood samples from 18S rRNA gene were detected in 78.41%, 62.86%, 68.29%, and 18S rRNA gene, insect jade horse BABEI detection rate was 11.36%, respectively. 11.43%, 90.24%. At the same time, the fresh blood samples making blood smears, through a microscope observation, found of theileriaequi and jade horse BABEI worm body exists, its size and morphology and the literature description. There are some differences between the results of PCR and cELISA of the two methods, the positive rate of PCR is higher than cELISA, this may be because the PCR method has higher detection sensitivity, in addition, may be due to sampling when some horses are in the early stages of infection, no antibodies. CELISA test results of some samples were positive, but PCR detection is negative, it may be because Martel purpose of horses in vivo has been their phagocytic system eliminated, but the antibody will continue to exist for some time in the body. Some samples were negative for cELISA test, but PCR method was positive, probably because the host was just infected, or after the antibody disappeared, there was a new infection. In Guizhou, Yili and southwest horse pony horse serum as the material, uses the method of ELISA serological research on EIAV, EAV and EHV-1, the 3 horses were not detected in EIAV antibody positive samples; the positive rate of EAV was 48.08%, 22.86%, 10.81%; the positive rate of EHV-1 was 99.38%, 97.14%, 100%. The EHV-1 DNA was amplified by PCR, the Guizhou pony, southwest horse and Yili horse EHV-1 detection rates were 94.23%, 80%, 83.78%. As the clinical symptoms of horse transmission and Martel's disease are similar, the results of EIAV antibody negative provide a theoretical basis for the diagnosis of Martel's disease. The positive rate of EHV-1 is high, but the mortality of the disease is not high, and it may be a secondary infection. The positive rate of serum EAV is southwest of Guizhou Pony Ma and Yili horse must be high, and equine viral arteritis with high mortality, suggest that EAV may be one of the causes of death of a large number of pathogenic Guizhou pony. The results show that the Martel worm disease and equine viral arteritis was likely to cause the death of a large number of Guizhou pony. For the remaining sick horses according to their clinical symptoms with ivermectin, blood worms and drug combo with red Di timely treatment and application of antibiotics for the treatment of secondary infection, until the horse recovered, eventually reduce the death rate of local horses. In summary, this study used microscope and ELISA and PCR on the main Guizhou pony infectious disease serology and etiology, not only reduces the farmers' economic loss, avoid the Guizhou pony this precious varieties will be extinct nightmare, is when the diagnosis, equine animal diseases provide theory a reliable basis for the prevention and treatment of.
【学位授予单位】：贵州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S858.21

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