兽用疫苗中药佐剂发展与管理研究

本文聚焦兽用疫苗中药佐剂的研究与产业化发展，总结兽医领域在单味药与中药组份佐剂，以及配合疫苗使用的“免疫增强剂”效用，从现代免疫与中兽医理论两个角度分析作用机理，推动兽用疫苗中药佐剂在提高免疫效果、提高疫苗免疫应答方面的应用，提出产业发展对策与管理建议。     

细胞因子类佐剂在黏膜免疫中的作用

黏膜免疫和黏膜疫苗佐剂的研究是近年来免疫学研究的一个重要方面.由于细胞因子在免疫应答的产生和调节中具有重要作用,可作为免疫佐剂增强疫苗的免疫效果,所以,细胞因子在黏膜佐剂研究中引起了人们广泛的兴趣与关注.文章主要简述黏膜免疫应答的调节及细胞因子类佐剂在黏膜免疫的研究与应用.     

新型油佐剂A5与进口206油佐剂配制双相疫苗比较试验

佐剂能增强抗原的免疫原性, 提高抗原的特异性免疫效果. 早在1925年, 在免疫制剂中已开始使用佐剂. 随着生物技术的发展, 疫苗的更新换代, 佐剂已成为提高疫苗质量的重要部分,受到科技工作者的重视,并得到不断发展和提高.我们经数年与中牧股份公司兰州生物制药厂合作,共同开发研制出了双相佐剂-A5,经兰州生物制药厂质量检测实验室检测,各项检测指标合格,免疫效果良好.为了降低生产成本,提高疫苗质量,笔者等对新型油佐剂A5与进口206油佐剂配制的双相疫苗进行了比较试验,现介绍如下.     

细胞因子类佐剂在黏膜免疫中的作用

黏膜免疫和黏膜疫苗佐剂的研究是近年来免疫学研究的一个重要方面。由于细胞因子在免疫应答的产生和调节中具有重要作用，可作为免疫佐剂增强疫苗的免疫效果，所以，细胞因子在黏膜佐剂研究中引起了人们广泛的兴趣与关注。文章主要简述黏膜免疫应答的调节及细胞因子类佐剂在黏膜免疫的研究与应用。     

新型纳米乳佐剂在口蹄疫疫苗中的应用研究

本研究创制新型纳米佐剂与口蹄疫O型灭活抗原按1:1比例配制疫苗,并对该疫苗的物理性状、安全性以及免疫效果进行系统研究,结果表明,新型纳米佐剂疫苗流动性佳,易注射,稳定性好,免疫副反应小以及免疫效果均优于商品化ISA206佐剂疫苗,为纳米乳疫苗的后期研究与应用提供依据。     

弓形虫疫苗研究进展

弓形虫是细胞内寄生的一种原虫,弓形虫病是由弓形虫引起的一种重要人兽共患寄生虫病,弓形虫疫苗作为防控弓形虫病的重要措施是目前研究的热点。弓形虫进入机体后以细胞免疫为主,使得疫苗可以发挥最大功用。疫苗的免疫效果与候选抗原和佐剂的使用密切相关,不同类型的抗原和佐剂用于疫苗中能够产生不同的免疫效果。目前,弓形虫疫苗的候选抗原主要以虫体特异组分蛋白和新型表位基因为主,佐剂也从传统的弗氏佐剂拓展到了霍乱毒素、透明质酸酶和CpG-ODN等。综合考虑各方面因素进行疫苗研制,将成为生产具有理想保护性弓形虫疫苗的研究方向。     

不同佐剂对猪附红细胞体亚单位疫苗免疫效果的影响

为比较不同佐剂的猪附红细胞体亚单位疫苗的免疫效果,本试验提取猪附红细胞体抗原,并将其分别与白油佐剂、弗氏佐剂、明矾佐剂及铝胶佐剂混合,免疫小鼠。应用间接ELISA方法比较不同佐剂的猪附红细胞体亚单位疫苗的免疫效果。结果表明:白油佐剂组免疫效果最好,抗体水平在一周后有明显上升,其次为弗氏佐剂组、铝胶佐剂组和明矾佐剂组。该试验为猪附红细胞体亚单位疫苗的研究提供了可靠的理论依据,为该病的防治奠定了一定的基础。     

不同佐剂对猪附红细胞体亚单位疫苗免疫效果的影响

为比较不同佐剂的猪附红细胞体亚单位疫苗的免疫效果,笔者通过试验提取猪附红细胞体抗原,并将其分别与白油佐剂、弗氏佐剂、明矾佐剂及铝胶佐剂混合,免疫小鼠。应用间接ELISA方法比较不同佐剂的猪附红细胞体亚单位疫苗的免疫效果。结果表明：白油佐剂组免疫效果最好,抗体水平在一周后有明显上升,其次为与弗氏佐剂组、铝胶佐剂组和明矾佐剂组。该试验为猪附红细胞体亚单位疫苗的研究提供了可靠的理论依据,为该病的防治奠定了一定的基础。     

疫苗佐剂的研究发展和前景展望

佐剂对于疫苗的免疫效果具有至关重要的作用,本文对目前常用佐剂及纳米佐剂的类别及其免疫机制的研究发展进行了系统性的综述。纳米技术作为一门新兴的科学技术,在佐剂的应用和研究发展中具有十分可观的潜质。     

3种不同类型佐剂的猪圆环病毒2型亚单位灭活疫苗免疫效果的比较研究

为制备猪圆环病毒2型商品化亚单位疫苗选择良好的佐剂,对由水性佐剂GEL 01、白油佐剂以及氢氧化铝胶佐剂分别制备的3种猪圆环病毒2型亚单位疫苗的免疫效果进行比较研究。本实验选取3~4周龄猪圆环病毒2型抗体阴性猪30头,根据试验疫苗接种情况分为6组,分别为:3种不同佐剂制备的亚单位疫苗(A组、B组和C组)、商品化疫苗(D组)以及空白对照组和攻毒对照组,然后进行临床观察、抗体水平监测和免疫效力评价,观察各组仔猪的不良反应情况、抗体水平以及攻毒保护情况。结果显示:3种不同类型佐剂制备的猪圆环病毒2型亚单位疫苗接种猪后均无任何不良反应、能刺激机体产生良好的ELISA抗体水平和免疫效果,但水性佐剂GEL 01制备的疫苗与另外2种佐剂制备的疫苗相比,综合抗体水平、免疫效果等因素,水性佐剂GEL 01制备的疫苗在临床使用上更具优势,为商品化亚单位疫苗佐剂的选择提供了重要的数据支持。     

鸡球虫病免疫学研究进展

鸡球虫病是严重危害养禽业的一种寄生虫病。一直以来,鸡球虫病主要依靠药物来进行防治。但是,随着球虫抗药性问题的日益严重,抗球虫新药开发困难及人们对食品安全的关注,药物在球虫病控制中的作用越来越受到限制,因而用免疫方法来控制鸡球虫病日益受到重视,并有望成为主要的技术手段。目前鸡球虫免疫学研究主要集中在鸡球虫免疫原性、宿主免疫应答及寻找有效的鸡球虫保护性抗原等方面。近年来随着分子生物学和现代生物技术的发展,鸡球虫基因工程疫苗以其独特的优势显示其在未来球虫病控制中的诱人前景,并可能取代传统疫苗而成为鸡球虫病防治的主要手段。     

In vivo effects of CpG oligodeoxynucleotide on Eimeria infection in chickens

Poultry coccidiosis is the major parasitic disease of poultry and, until now, no recombinant vaccine has been developed. Short oligodeoxynucleotides (ODNs) containing unmethylated CpG motifs (CpG ODNs) have been shown to be effective immunoprotective agents and vaccine adjuvants in mammalian systems. Their use in poultry to protect against intracellular parasites has not been reported to date. The present work investigated the effects of CpG ODN treatment on host susceptibility to Eimeria infection in two chicken strains with different genetic background, SC and TK. The data show that CpG ODN enhanced the birds' resistance to coccidiosis in a normally susceptible chicken strain (TK), as shown by reduced oocyst shedding and improved weight gain. CpG treatment had a differential effect on body weight gains and serum antibody responses, depending on the chicken strain and ODN dose, delivery route, and backbone. This study shows for the first time that CpG ODNs could be used as immunoprotective agents in Eimeria-infected chickens to enhance resistance to the pathogen and improve performance. Future research is needed to optimize their use alone and as vaccine adjuvants that may lead to better and more efficient vaccine applications.     

Comparison of tissue reactions produced by Haemophilus pleuropneumoniae vaccines made with six different adjuvants in swine.

Tissue damage caused by six different adjuvants incorporated in a Haemophilus pleuropneumoniae vaccine was compared in swine. The adjuvants compared were four mineral oil compounds, one peanut oil compound and aluminum hydroxide. Inoculations were given in the neck, quadriceps and semitendinosus muscles. The mineral oil adjuvants were highly irritant and caused extensive areas of granulomatous inflammation that were present at eight weeks after injection. The aluminum hydroxide produced smaller lesions that also persisted for eight weeks. Only the peanut oil adjuvant did not produce significant lesions at the site of injection. At two and four weeks, but not at eight weeks postinoculation, lesions in the quadriceps and semitendinosus muscles were approximately twice as extensive as those in the muscles of the neck.     

Immunoadjuvant effects of bacterial genomic DNA and CpG oligodeoxynucleotides on avian influenza virus subtype H5N1 inactivated oil emulsion vaccine in chicken

This study investigated the immunoadjuvant effects of three types of bacterial genomic DNA and CpG oligonucleotides (CpG ODN) on the avian influenza virus (AIV) subtype H5N1 inactivated oil emulsion vaccine under two immunization strategies. The genomic DNA extracted from Escherichia coli O₂, Staphylococcus aureus, Streptococcus faecalis FQ68, and synthetic CpG ODN were used as adjuvants, and their effects on the AIV oil emulsion vaccine were examined in chickens. The results indicated that when administered separately from the vaccine, adjuvants induced lower haemagglutination inhibition (HI) titres and serum IgG titres but resulted in higher concentrations of IFN-γ and IL-10. In contrast, when combined with the oil emulsion vaccine prior to inoculation, CpG ODN induced higher HI, IgG titres and IFN-γ concentration but resulted in lower IL-10 concentration. These data suggest that, depending on the immunization approaches, adjuvants may exert distinct immune effects in chickens receiving AIV H5N1 oil emulsion vaccine: the prior incorporation of CpG ODN into the vaccine may augment both the humoral and Th1 type immune responses, while separate inoculation of adjuvants has not shown better adjuvanticity.     

三种佐剂对鸡体液免疫影响的比较

佐剂是疫苗的重要组成部分,不同佐剂对体液免疫会产生不同的作用.选择合适的佐剂不仅可以增强疫苗的免疫效果,而且可以提高机体的抵抗力.本试验选用白油佐剂、弗氏佐剂和蜂胶佐剂分别与灭活的新城疫病毒制成疫苗,免疫雏鸡,探讨不同佐剂对鸡体液免疫的影响.结果表明,试验组鸡抗体水平高,饲养至61日龄均未发生新城疫,其中弗氏佐剂的效果最好,蜂胶佐剂的效果次之,油佐剂的效果最次.     

Effects of Different Adjuvants on the Immunogenicity of PCV2-Cap Protein

The study was aimed to prepare vaccines with different adjuvants,and research its effects on immunogenicity.The PCV2 Cap gene with its signal peptide removed was connected to pET-28a vector,and then was induced to express,using sodium deoxycholate（DOC）and low concentration of urea to dissolve the inclusion body.Different adjuvants,such as alum-based adjuvants,liposome adjuvants,propolis adjuvants,white oil adjuvants and Freund adjuvant were prepared,together with protein purified to make up subunit vaccines,and commercial inactivated vaccine as positive control,immuning mice,and ELISA method were used to detect changes in concentrations of animal serum antibody and cytokines,evaluated the immune protective effect.Results showed that the expression product in the form of inclusion body,using the DOC could omit dissolving inclusion body protein renaturation steps,and the purification method was simple,the capsid protein obtained had high purity.ELISA assays showed that PCV2-Cap had good immunogenicity.And we found that the water system adjuvants had high immune activity,this could provide important previous experimental data for the commercialization of the subunit vaccine.     

不同佐剂对猪圆环病毒衣壳蛋白免疫原性的影响

试验旨在研究猪圆环病毒不同佐剂疫苗的制备及其对免疫原性的影响。将去除信号肽的PCV2（porcine circovirus type 2,PCV2）Cap蛋白基因连接在pET-28a载体上,进行诱导表达,用脱氧胆酸钠（DOC）和低浓度尿素对表达产物进行溶解,制备氢氧化铝胶体佐剂、脂质体佐剂、弗氏佐剂、白油佐剂、蜂胶佐剂,并与纯化的PCV2-Cap蛋白混合制成亚单位疫苗,以商品化的灭活疫苗作为阳性对照,免疫小鼠,并使用ELISA方法检测动物血清中抗体及细胞因子含量的变化,评估其免疫保护效果。结果显示,表达产物以包涵体的形式存在,使用DOC溶解包涵体可省略蛋白复性步骤,且纯化方法简单,获得纯度较高的衣壳蛋白;ELISA检测结果表明PCV2-Cap蛋白能诱导产生特异性较高的抗体;水系佐剂制备的亚单位疫苗具有较高的免疫活性,为亚单位疫苗的商品化提供重要的数据支持。     

An improved Corynebacterium pseudotuberculosis vaccine for sheep

Extensive experiments in mice confirmed that the immunogenicity of a Corynebacterium pseudotuberculosis vaccine could not be significantly improved with the use of various adjuvants. Immunity against C. pseudotuberculosis likewise could not be enhanced by incorporating various immunostimulants into the vaccine or by the use of live vaccines. However, a combination of aluminium hydroxide gel and saponin as adjuvant did have a beneficial effect. This vaccine was tolerated better, and a smaller dose apparently protected sheep more effectively against intralymph node challenge than the currently available alum-precipitated vaccine.     

Effects of adjuvants on the immune response of staphylococcal alpha toxin and capsular polysaccharide (CPS) in rabbit

This study was performed to isolate a vaccine strain of S. aureus from clinical or subclinical mastitis and to choose the most optimal adjuvant for immune response of alpha toxin and capsular polysaccharide (CPS) of field strain. Of thirty strains of S. aureus isolated from milk of clinical or subclinical mastitis, V112 strain isolated from milk of gangrenous mastitis was used in this vaccine. Twenty one of rabbits were allocated into 5 groups based on adjuvants and immunized twice every 2 weeks for 8 weeks. This vaccine was composed of alpha toxin (10 hemolytic units) and formalinized whole cells (1 x 10(11) cells/ml. Five rabbits received PBS solution as a control group. The highest antibody titers against alpha toxin and CPS were observed in dextran sulfate- and aluminium hydroxide-adjuvant group at 8 weeks after immunization, respectively. These results of the study showed that one adjuvant could not induce strong and long-term immune response of alpha toxin and CPS antigens. Therefore, the use of combined adjuvants in subunit vaccine may be useful and feasible.     

Strategies and hurdles using DNA vaccines to fish

DNA vaccinations against fish viral diseases as IHNV at commercial level in Canada against VHSV at experimental level are both success stories. DNA vaccination strategies against many other viral diseases have, however, not yet yielded sufficient results in terms of protection. There is an obvious need to combat many other viral diseases within aquaculture where inactivated vaccines fail. There are many explanations to why DNA vaccine strategies against other viral diseases fail to induce protective immune responses in fish. These obstacles include: 1) too low immunogenicity of the transgene, 2) too low expression of the transgene that is supposed to induce protection, 3) suboptimal immune responses, and 4) too high degradation rate of the delivered plasmid DNA. There are also uncertainties with regard distribution and degradation of DNA vaccines that may have implications for safety and regulatory requirements that need to be clarified. By combining plasmid DNA with different kind of adjuvants one can increase the immunogenicity of the transgene antigen – and perhaps increase the vaccine efficacy. By using molecular adjuvants with or without in combination with targeting assemblies one may expect different responses compared with naked DNA. This includes targeting of DNA vaccines to antigen presenting cells as a central factor in improving their potencies and efficacies by means of encapsulating the DNA vaccine in certain carriers systems that may increase transgene and MHC expression. This review will focus on DNA vaccine delivery, by the use of biodegradable PLGA particles as vehicles for plasmid DNA mainly in fish.