猪源A型多杀性巴氏杆菌灭活疫苗制备及免疫效果评估

Preparation of Inactivated Vaccine Against Porcine Pasteurella multocidaSerotype A and Evaluation of Its Immune Effect

【目的】制备猪源A型多杀性巴氏杆菌灭活疫苗,有效控制巴氏杆菌病的发生和流行。【方法】从发病猪病料中分离猪源A型多杀性巴氏杆菌,测定生长曲线,通过小鼠致病性试验筛选疫苗候选菌株。将筛选得到的疫苗候选菌株培养后进行灭活,无菌检验合格后,按1∶4的比例与SUMMIT-CB-200佐剂混合,制备猪源A型多杀性巴氏杆菌灭活疫苗。最后通过小鼠和仔猪免疫攻毒保护试验评估疫苗保护力。【结果】分离到11株猪源A型多杀性巴氏杆菌;生长曲线测定结果显示,11株菌均在培养至6 h时达到生长稳定期,活菌数范围为2.1×10^(9)~8.4×10^(9)CFU/mL;小鼠致病性试验结果显示,攻毒后小鼠出现精神沉郁、食欲降低、行动迟缓、被毛粗糙、扎堆、眼睛周围大量脓性分泌物、腹式呼吸、共济失调等症状,共筛选到A4、A8、A9和A11 4株疫苗候选菌株,其中最强毒株A4小鼠皮下攻毒半数致死量(LD50)为167 CFU;仔猪毒力试验结果显示,攻毒后仔猪出现精神沉郁、食欲降低、皮肤发红、呕吐、呼吸困难、腹式呼吸、体温升高、关节肿大、跛行等症状,A4菌株仔猪皮下攻毒LD50为1.72×10^(10)CFU;免疫攻毒保护试验结果显示,4株猪源A型多杀性巴氏杆菌灭活疫苗对小鼠和仔猪均具有一定保护力,其中A9保护力最强,对小鼠保护率达60%,对仔猪保护率达75%。【结论】本研究制备的灭活疫苗能对猪源A型多杀性巴氏杆菌强毒株提供较好的免疫保护力,具有一定的开发价值。

【Objective】The aim of the experiment was to prepare porcine Pasteurella multocida serotype A inactivated vaccine to effectively control the occurrence and prevalence of pasteurellosis.【Method】Pasteurella multocida serotype A was isolated from diseased swine samples,and the growth curves were determined.Then the vaccine candidate strains were screened by pathogenicity test in mice.The obtained vaccine candidate strains were cultured and inactivated.After the sterility test was qualified,the inactivated vaccine were prepared by mixing with SUMMIT-CB-200 adjuvant in a ratio of 1∶4.The vaccine protection ability was evaluated by immune challenge protection tests in mice and piglets respectively.【Result】11 strains of porcine Pasteurella multocida serotype A were isolated.The growth curve results showed that 11 strains reached a stable stage when cultured for 6 h,and the number of viable bacteria ranged from 2.1×10^(9) to 8.4×10^(9) CFU/mL.The pathogenicity test of mice showed that after challenge,the mice showed depression,decreased appetite,slow action,rough coat,clumping,large purulent secretions around the eyes,abdominal breathing,ataxia and other symptoms.Four vaccine candidate strains A4,A8,A9 and A11 were screened.The strongest strain A4 had a subcutaneous challenge median lethal dose(LD 50)of 167 CFU in mice.The results of virulence test of piglets showed that depression,loss of appetite,skin redness,vomiting,dyspnea,abdominal breathing,elevated body temperature,joint enlargement,lameness and other symptoms after challenge.The strain A4 had a subcutaneous challenge LD 50 of 1.72×10^(10) CFU.The results of immune challenge protection test showed that the four strains of inactivated vaccine had certain protective effect on mice and piglets,among which A9 had the strongest protection,with a protection rate of 60%for mice and 75%for piglets.【Conclusion】The inactivated vaccine prepared in this study could provide better immune protection against Pasteurella multocida serotype A,which had certain development value.