淋病奈瑟菌感染动物模型的实验性研究和探讨

发布时间：2018-06-08 14:41

本文选题：淋病奈瑟菌 + 生殖道感染　；参考：《华中科技大学》2006年硕士论文

【摘要】： 淋病(gonorrhea)是由淋病奈瑟菌(Neisseria gonorrhoeae)引起的经典性病,1879年由德国人Neisser首次发现。人类是淋病奈瑟菌唯一的自然宿主,主要通过性接触而传播。淋病奈瑟菌可引起泌尿生殖道及多种附属器官和组织的化脓性感染,包括淋菌性尿道炎或生殖道炎、膀胱炎、前列腺炎、子宫颈炎、子宫内膜炎等多种疾病。妊娠期妇女感染淋病奈瑟菌,可发生羊膜腔内感染,包括胎儿感染。宫内感染可导致流产,约占自然流产的32%。孕妇感染后,80%以上无明显症状,如果没有得到及时治疗,在分娩过程中,产道的淋病奈瑟菌就会传播给新生儿,导致新生儿淋菌性结膜炎,或新生儿淋病。淋病是世界范围内发病率较高的性传播疾病之一,据WHO估计,全球每年新发病例超过6000万。我国近二十年16个城市监测发现,1987年淋病患者为11,115例,1997年上升为224,331例,1999年达340,960例,年增长率超过30%,淋病发病率为20.63/10万。2000年之后开始下降,淋病患者为285,661人,2001年为234,561人,发病人数仍位居性病之首。淋病奈瑟菌的唯一自然宿主是人类,一般情况下非常容易引起人类泌尿生殖道感染,但却不能感染所有常用实验动物。灵长类动物虽可部分复制出人类淋病感染过程,但由于经济及来源等方面的原因并不是理想的模型动物。科学家们曾进行了各种尝试试图找到合适的模型动物:早期曾用淋病奈瑟菌感染鸡胚,随后Arko等尝试用淋病奈瑟菌感染一系列的动物,包括兔、豚鼠等,但是都没有成功。经过长期艰苦的探索后,有研究表明:运用雌激素预处理小鼠,可短期制造出淋病奈瑟菌感染动物模型,为建立淋病实验动物模型提供了新思路。本研究在此基础上通过摸索尝试建立了一种简便、易掌握的淋病奈瑟菌感染动物模型,为研究淋病奈瑟菌致病机制及疫苗开发提供了合适的动物模型。本研究工作分以下二个部分: 一、淋病奈瑟菌感染动物模型的实验性研究采用野生型临床株淋病奈瑟菌生殖道接种17-β-雌二醇预处理的雌性BALB/c小鼠,同时腹腔注射抗生素(万古霉素、链霉素)以抑制生殖道共生菌的过度生长。小鼠生殖道拭子接种WGC选择培养基培养分离淋病奈瑟菌,取小鼠阴道分泌物涂片Gram染色观察,并用改良瑞氏染色和姬母萨染色观察并计数小鼠阴道分泌物涂片中多形核白细胞(polymorphonuclearleukocytes,PMN)的数目。受试小鼠生殖道中淋病奈瑟菌平均存活8天(2×106CFU接种量),阴道分泌物涂片Gram染色可见中性粒细胞中吞噬有革兰阴性双球菌,小鼠阴道拭子分离培养呈阳性,WGC选择培养基分离的可疑菌落经生化反应鉴定为淋病奈瑟菌,受试小鼠发生了炎症。二、淋病奈瑟菌主要外膜蛋白PⅠB疫苗免疫保护性研究选用清洁级雌性BALB/c小鼠,随机分为实验组和对照组。用淋病奈瑟菌PⅠB蛋白疫苗免疫实验组小鼠,经3次加强免疫后,用间接ELISA法测定实验组小鼠抗淋病奈瑟菌PⅠB蛋白IgG抗体水平,证实其具有较高抗体滴度后按第一部分介绍的方法建立淋病奈瑟菌感染实验动物模型,实验组淋病奈瑟菌的存活时间短于对照组,可认为经淋病奈瑟菌PⅠB蛋白免疫后,小鼠生殖道清除淋病奈瑟菌的速率明显加快,产生了免疫保护性。结论:成功建立了淋病奈瑟菌感染的动物模型,为研究淋病奈瑟菌的致病机制和疫苗开发研究奠定了基础。
[Abstract]:Gonorrhoeae (gonorrhea) is a classic venereal disease caused by Neisseria gonorrhoeae (Neisseria gonorrhoeae). It was first discovered by German Neisser in 1879. Human is the only natural host of Neisseria gonorrhoeae, which is transmitted mainly through sexual contact. Neisseria gonorrhoeae can cause a purulent infection of the genitourinary tract and a variety of accessory organs and tissues, including gonococcus Sexual urethritis or genital inflammation, cystitis, prostatitis, cervicitis, endometritis, and other diseases. Pregnant women infected with Neisseria gonorrhoeae, can occur amniotic infection, including fetal infection. Intrauterine infection can lead to abortion, about 80% of pregnant women with spontaneous abortion, more than 80% without obvious symptoms, if not in time. If not in time In the course of the treatment, Neisseria gonorrhoeae of the gonorrhoeae is transmitted to the newborn, resulting in gonococcal conjunctivitis, or neonate gonorrhea. Gonorrhea is one of the world's high incidence of sexually transmitted diseases. According to WHO estimates, more than 60 million new cases are found every year in the world. In China, 16 City monitoring found in the last twenty years, gonorrhea in 1987. The patients were 11115 cases, 224331 cases rose in 1997, 340960 cases in 1999, the annual growth rate was more than 30%. The incidence of gonorrhea fell after 20.63/10 million.2000 years, 285661 in gonorrhea and 234561 in 2001, and the number of diseases was still the first of venereal disease.
The only natural host of Neisseria gonorrhoeae is human. In general, it is very easy to cause human genitourinary tract infection, but it can not infect all the common experimental animals. Though the primate can partially replicate the process of human gonorrhea infection, it is not an ideal model animal because of economic and source reasons. A variety of attempts were made to find the right model animals: early use of Neisseria gonorrhoeae to infect chicken embryos, and then Arko and other strains of Neisseria gonorrhoeae were tried to infect a series of animals, including rabbits and guinea pigs, but all failed. After a long and hard exploration, the study showed that the use of estrogen to pretreat mice could be produced in a short period of time. The animal model of Neisseria disease infected with Neisseria gonorrhoeae has provided a new idea for the establishment of an experimental animal model of gonorrhea. On this basis, a simple and easy mastered animal model of Neisseria gonorrhoeae infection was established on the basis of this study, and a suitable animal model was provided for the study of the pathogenesis of Neisseria gonorrhoeae and the development of the vaccine.
The research work is divided into two parts:
Experimental study on the animal model of Neisseria gonorrhoeae infection
17- beta estradiol pretreated female BALB/c mice were inoculated with Neisseria gonorrhoeae gonorrhoeae in the wild type clinical strain, and antibiotics (vancomycin, streptomycin) were intraperitoneally injected to inhibit the overgrowth of the reproductive tract symbionts. The mouse reproductive swabs were inoculated with WGC selection medium to isolate Neisseria gonorrhoeae, and the vaginal secretions were smeared to Gr in mice. Am staining was used to observe and count the number of polymorphonuclear leukocytes (polymorphonuclearleukocytes, PMN) in the smears of vaginal secretions in mice. The average survival rate of Neisseria gonorrhoeae in the reproductive tract of the mice was 8 days (2 x 106CFU inoculation), and Gram staining of vaginal secretions showed that neutrophils were swallowed in the vaginal secretions. With gram-negative diplococcus, the isolated culture of vaginal swabs in mice was positive. The suspicious colonies separated by the WGC selection medium were identified as Neisseria gonorrhoeae by biochemical reaction, and the mice were inflamed.
Two, the protective immunity of Neisseria gonorrhoeae major outer membrane protein P I B vaccine
The female BALB/c mice of clean grade were randomly divided into experimental group and control group. The mice were immunized with Neisseria gonorrhoeae P B protein vaccine in the experimental group. After 3 times of immunization, the level of IgG antibody of P I B protein of Neisseria gonorrhoeae in experimental group was determined by indirect ELISA method, and the method was proved to have high antibody titer and the method introduced in the first part. The experimental animal model of Neisseria gonorrhoeae infection was established. The survival time of Neisseria gonorrhoeae in the experimental group was shorter than that of the control group. It was considered that after immunization with Neisseria gonorrhoeae P I B protein, the rate of scavenging Neisseria gonorrhoeae in the mouse reproductive tract was obviously accelerated, and the immune protection was produced.
Conclusion: the animal model of Neisseria gonorrhoeae was successfully established, which laid a foundation for studying the pathogenic mechanism and vaccine development of Neisseria gonorrhoeae.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R-332

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