禽大肠杆菌分离、鉴定及三价灭活疫苗的研制

发布时间：2018-07-16 12:08

【摘要】：鸡大肠杆菌病是由大肠埃希氏菌的某些血清型菌株引起一类传染病的总称。本病既可通过种蛋垂直传播,也可通过被大肠杆菌污染的饲料、饮水、空气经消化道和呼吸道感染,由于感染途径不同,临床上可引起急性败血症、气囊炎、心包炎、肝周炎、输卵管炎、卵黄性腹膜炎、关节炎、肉芽肿、死胚、肠炎、脑炎、眼炎等许多病型。近几年来,随着我国集约化养鸡业的迅速发展,其发病也日趋严重,危害也越来越明显,目前已成为危害我国养鸡业发展的重要疾病之一,能否有效防治鸡大肠杆菌病,己成为决定养鸡业成败的关键问题。由于大肠杆菌极易产生耐药性,随着大量高效抗菌药物的普遍应用和耐药性的产生,使得药物治疗大肠杆菌病的效果越来越差。因此研制有效的疫苗来预防鸡大肠杆菌病,越来越受到重视并被广泛的采用。由于鸡致病性大肠杆菌抗原结构复杂,血清型众多,不同血清型菌株之间缺乏完全交叉保护,加之不同地区的优势血清型又存在很大的差异,很难研制一种对全国各地鸡大肠杆菌病都有预防作用的疫苗,给本病的免疫防制带来了较大困难。目前被广泛采用的方法是研制现地多价疫苗,现地应用,实践证明,应用这种方法收到了很好的免疫效果。我们从山东省聊城、菏泽、诸城、潍坊、烟台、济宁等地区进行禽大肠杆菌流行病学调查,成功分离到了130株大肠杆菌,血清学结果表明,分离的大肠杆菌分属13个血清型,分别为O78、O1、O2、O38、O88、O9、O24、O127、O15、O82、O35、O18、O114,占所分离大肠杆菌菌株的92.3%;未定型分离株10株,占7.7%。其中,O78、O1、O2为优势血清型,其中O78型40株,O1型21株,O2型15株,分别占所分离菌株的30.8%、16.2%、11.5%。我们进行动物回归试验,挑选选出毒力强的O1、O2、O78禽大肠杆菌菌株进行毒力试验,筛选出毒力强、免疫原性好的O1型QL-003株、O2型QL-035株、O78型QL-057株做为制苗用菌种进行三价疫苗的制备。我们进行了禽大肠杆菌O1型QL-003株、O2型QL-035株、O78型QL-057株生产工艺研究、毒力及免疫原性试验,以此为基础,制备禽大肠杆菌三价灭活疫苗,并进行了疫苗性状、无菌、安全、效力、甲醛、汞类防腐剂残留量测定等检验,结果均合格。将3批疫苗免疫后2个月、4个月、6个月进行攻毒,对O1型O78型QL-057株、O2型QL-035株和O78型QL-057株的攻毒保护率均不低于80%(8/10),对照鸡均100%(10/10)发病,表明我们研制的3价灭活疫苗免疫期至少为6个月。
[Abstract]:Escherichia coli is a kind of infectious disease caused by some serotype strains of Escherichia coli. The disease can be transmitted vertically by breeding eggs, or through food contaminated by E. coli, drinking water and air through digestive tract and respiratory tract infections. Because of the different routes of infection, it can cause acute septicemia, airbag inflammation, pericarditis, and pericarditis. Salpingitis, yolk peritonitis, arthritis, granuloma, dead embryo, enteritis, encephalitis, ophthalmitis and many other diseases. In recent years, with the rapid development of intensive chicken industry in China, the incidence of chicken disease is becoming more and more serious, and the harm is becoming more and more obvious. At present, it has become one of the important diseases that endanger the development of chicken industry in China. Has become the key to determine the success or failure of the chicken industry. As Escherichia coli is easy to develop drug resistance, with the widespread use of a large number of highly effective antimicrobial agents and the emergence of drug resistance, the efficacy of drugs in the treatment of Escherichia coli is getting worse and worse. Therefore, more and more attention has been paid to the development of effective vaccine to prevent chicken E. coli disease. Due to the complex structure of pathogenic Escherichia coli antigen and numerous serotypes, there is a lack of complete cross-protection among different serotype strains, and there are great differences in dominant serotypes in different regions. It is very difficult to develop a vaccine which can prevent chicken colibacillosis all over the country. At present, the widely used method is to develop a field multivalent vaccine, which has been applied in the field. It has been proved that the application of this method has a very good immune effect. We carried out epidemiological investigation of Escherichia coli in poultry from Liaocheng, Heze, Zhucheng, Weifang, Yantai and Jining areas of Shandong Province, and successfully isolated 130 strains of Escherichia coli. The serological results showed that the isolated Escherichia coli belonged to 13 serotypes. They were O78-O1O1O2O3O3O8O88O9, O24O127O127O127O127O15O82O35O3O18O114, respectively, accounting for 92.3x of the Escherichia coli strains isolated, and 10 strains of unshaped isolates, accounting for 7.7% of Escherichia coli strains isolated. Among them, O78, O1, O2 were the dominant serotypes, among which 40 strains of O78, 21 strains of O1 and 15 strains of O2, accounted for 30.8% of the isolated strains, 16.2and 11.5T, respectively. The animal regression test was carried out and the virulence test was carried out by selecting the highly virulent strains of avian Escherichia coli O1O2O78. The highly virulent and immunogenic QL-003 strain O1 QL-035 strain O78 QL-057 strain were selected for the preparation of trivalent vaccine for vaccine production. The production process, virulence and immunogenicity of avian Escherichia coli O1 QL-003 strain O1 QL-035 strain O78 QL-057 strain were studied. Based on these tests, avian Escherichia coli trivalent inactivated vaccine was prepared. The results of determination of formaldehyde and mercury preservative residues were all up to standard. Two months, four months and six months after the three batches of vaccines were immunized, the protection rates of O1 O78 QL-057 QL-035 and O78 QL-057 QL-057 strains were not less than 80% (8 / 10), 100% (10 / 10) of the control chickens, respectively. The results showed that the 3-valent inactivated vaccine had been immunized for at least 6 months.
【学位授予单位】：山东农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S855.12

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