禽用DNA疫苗免疫效果增强的策略

对于人和动物的传染病来说,DNA疫苗免疫是一种很有希望的免疫方式。经过25年的发展,已经上市的核酸疫苗仍屈指可数。禽用核酸疫苗一直是禽类疫苗的研究热点,文中描述提高核酸疫苗免疫效果的方式：接种途径,疫苗剂量及首免时间,增加宿主细胞对质粒的摄入,添加免疫增强分子,优化质粒骨架和密码子,疫苗抗原的选择,异源性的首免-加强免疫策略。     

提高核酸疫苗免疫效果的策略

核酸疫苗因能诱导机体产生保护性免疫,且具有安全、热稳定性、价廉等优点而受到高度重视,如何提高核酸疫苗的免疫效果一直是核酸疫苗研究的热点和难点问题。文章主要从优化方法着手,就核酸疫苗研制中质粒载体的构建、抗原加工分子的联合应用、接种途径和剂量以及核酸疫苗在禽病上的应用做一综述。     

七种免疫调节剂对鸭接种禽流感疫苗后体液免疫应答的促进作用

为筛选出对鸭接种禽流感疫苗后具有免疫促进作用的免疫调节剂,对接种禽流感H5亚型油乳灭活疫苗后的鸭分别注射或投喂禽源干扰素、人源干扰素、黄芪多糖、益生素、人参皂苷、赐力健及增免散7种免疫调节剂,观察禽流感疫苗特异性的体液免疫的促进作用。研究结果表明,饲喂人参皂苷或注射禽源干扰素对接种禽流感疫苗的试验鸭有明显的促进作用,其中人参皂苷在首免后3周发挥促进作用,而禽源干扰素约在首免后5周表现出一定的促进作用。     

禽病的控制要点

正一、禽病难以控制的原因1.疫苗质量。用假、劣质疫苗免疫接种后,因其效价低、灭活不全或净化不好等原因,不能有效刺激被免禽免疫系统而不能产生相应种类和数量的抗体,从而因达不到预期的预防效果,使禽病的控制难度增大。灭活不全或净化不好时,甚至还能激发或诱发被免禽疫病的发生和流行。2.免疫程序。没有免疫程序、免疫程序不科学或不按时执行免疫程序,因免疫病种及其血清型、接种时间、接种途径、疫苗用量等使疫苗效     

鸡传染性鼻炎三价灭活疫苗安全性试验和效力检验初报

利用PageA、B、C型副鸡禽杆菌,研制鸡传染性鼻炎(IC)三价灭活疫苗。用6批IC疫苗进行了大剂量单次接种和单剂量多次接种的安全性试验,结果表明,IC三价灭活疫苗安全无副作用;用3批疫苗进行SPF鸡和普通鸡的免疫效力试验,结果对A、B、C型副鸡禽杆菌攻毒的保护率≥80%;商品鸡42日龄首免,110日龄二免,二免后约9个月对A、B、C型副鸡禽杆菌攻毒的保护率≥80%;用4℃～8℃保存一年的3批疫苗进行了SPF鸡的近期免疫效力试验,结果对A、B、C型副鸡禽杆菌攻毒的保护率≥80%。     

奶牛免疫布鲁氏菌A19疫苗后的抗体变化和排菌情况监测

为探究奶牛免疫布鲁氏菌A19疫苗(以下简称A19疫苗)后的抗体变化规律及向外排菌的风险,使用A19疫苗免疫30头3月龄奶牛,在首免和加强免疫后3个月内,通过虎红平板凝集试验(RBT)和试管凝集试验(SAT)进行抗体检测,采用荧光定量PCR方法,进行血液以及口鼻、阴道拭子的布鲁氏菌核酸检测。结果显示:首免后6 d,奶牛抗体开始出现阳转,60 d后绝大部分奶牛抗体转阴,而加强免疫后3d,又出现抗体阳转;仅在首免后2 d内的阴道拭子和首免后6 d内的血液中检测到布鲁氏菌核酸,而在口鼻拭子中以及其他时间点的血液和阴道拭子中均未检测到布鲁氏菌核酸。结果表明,A19疫苗免疫奶牛抗体阳转快,排菌期短,安全性高。     

IBD疫苗的正确使用

1.确定最佳首免日龄种蛋蛋黄的抗体滴度与种母禽血清的抗体效价几乎是一致的。维禽母源抗体一般是随着蛋黄的吸收越来越高。至一定日龄达到高峰值,之后逐渐下降,约20天降至为零。而IBD的母源抗体一日龄最高,约18天之后降至为零。母源抗体直接影响着疫苗的免疫效果,所以IBD(甘保罗病法氏囊炎病)疫苗的首免日龄以14天接种为宜。 减少母源抗体的干扰作用在生产实践中无母源抗体的雏禽几乎是不存在的。雏禽母源抗体对油佐剂灭能疫苗的影响很少,首免接种油苗似手可能解     

鸡传染性鼻炎(三价)和新城疫二联灭活疫苗的研究

利用Page A型、B型、C型副鸡禽杆菌和新城疫病毒La Sota株,研制鸡传染性鼻炎(三价)和新城疫二联油乳剂灭活疫苗。用3个批次疫苗进行单剂量(0.5mL/次)3次接种和大剂量(2.0mL)单次接种的安全性试验、SPF鸡的免疫效力试验、商品鸡的免疫持续期试验和疫苗的保存期试验。结果表明,3批疫苗对试验鸡安全无副作用;免疫接种SPF鸡只30d后对A、B、C型副鸡禽杆菌攻毒的保护率≥80%,用20μL/只疫苗免疫接种SPF鸡21d后新城疫的平均HI抗体24;商品鸡42日龄首免,110日龄二免,二免后9个月对A、B、C型副鸡禽杆菌攻毒的保护率≥70%,对新城疫病毒强毒100%保护;用4℃~8℃保存12个月和15个月的3批疫苗进行了SPF鸡的近期免疫效力试验,结果对A、B、C型副鸡禽杆菌攻毒的保护率≥70%,用20μL/只疫苗免疫SPF鸡,免疫接种后21d新城疫病毒的平均HI抗体24,对新城疫病毒强毒的攻毒保护率70%。说明研制的疫苗安全有效。     

动物核酸疫苗的研究现状及发展前景

核酸疫苗是近年来备受人们关注的一种新型疫苗。核酸疫苗以其特有的可诱导机体产生全面的免疫应答,对不同亚型的病原体具有交叉防御作用,以及安全、可靠、生产方便等优点被称之为“疫苗的第三次革命”。核酸疫苗由编码能引起保护性免疫反应的病原体抗原的基因片段和载体构建而成,包括DNA疫苗和RNA疫苗,目前研究较多的是DNA疫苗。DNA疫苗是指含有编码抗原基因的真核表达质粒DNA,经直接接种体内后,可被体细胞摄取,并转录、翻译、表达出相应的抗原,然后通过不同途径刺激机体产生针对此种抗原的应答。作者简单介绍了动物核酸疫苗的特点、免疫机制、免疫影响因素及在畜禽传染病中的应用,此外还分析了核酸疫苗的发展前景等问题,从而为核酸疫苗的发展提供了新思路和新途径。     

猪传染性胃肠炎核酸疫苗免疫小鼠后的血清抗体IgG检测

本试验旨在对猪传染性胃肠炎核酸疫苗(含有TGEV-S基因质粒)开展免疫原性研究,为科学评价该TGE核酸疫苗的免疫效果提供依据。通过灌胃口服免疫TGE核酸疫苗诱导小鼠机体产生特异性免疫,定期采样监测其血清抗体产生规律,采用间接ELISA方法测定免疫小鼠血清抗体IgG的抗体效价。结果发现:TGE核酸疫苗免疫小鼠后有效地激发了TGEV特异性抗体,首免后第二周开始就能检测到血清抗体应答,且该组抗体水平一直都与空载体组和PBS空白对照组之间呈极显著差异(P0.01),表现出良好的免疫原性。     

禽类DNA疫苗研究进展

新型疫苗的研究对于禽类传染病的防制意义重大。传统疫苗是基于抗原刺激机体产生特异性抗体的原理,它们大多数激发机体的体液免疫,很难启动细胞免疫。脱氧核糖核酸(DNA)疫苗作为第3代疫苗,具备传统疫苗和其它基因工程苗不可比拟的优点,能够诱导全方位的免疫反应且使用更安全更方便,DNA疫苗是将外源基因与真核质粒重组后直接导入细胞内,使外源基因在宿主细胞内表达合成保护性抗原蛋白。这是模拟病毒自然感染提呈过程,既能产生细胞免疫,又能产生体液免疫。文章对DNA疫苗在禽类应用的可行性和应用研究新进展作了综述。     

Research Progress on Novel Vaccine of Avian Influenza

Avian influenza (AI) is an acute respiratory disease caused by influenza A virus.Avian influenza virus (AIV) can infect poultry,wild birds and some mammals including human.AI is a big threat to both poultry and human health because the virus can cross the species barrier to get the capacity of transmitting from poultry to human.Vaccination is the most efficient measure against AI outbreaking.Traditional vaccines include inactivated vaccine based on chick embryo and attenuated vaccine.Although the traditional vaccines play important roles in the past AI epidemics,many disadvantages have been proved to exist in traditional vaccines.Forced by major drawbacks of traditional vaccines,several studies focused on the development of novel vaccines.In this review,we reviewed recombinant live vector vaccine,subunit vaccine,DNA vaccine and virus-like particle vaccine of AI in order to provide some references for prevention and control of AI.     

禽流感新型疫苗研究进展

禽流感是由A型流感病毒引起的一种急性呼吸道传染病,家禽、野鸟和部分哺乳动物均可感染.禽流感给中国养殖业造成了巨大的损失,同时,随着病毒的种间传播,人类的生命安全也受到了严重威胁.目前,疫苗免疫仍是防控禽流感最主要的手段,传统的疫苗主要有鸡胚灭活苗和禽流感弱毒疫苗.虽然在过往几次禽流感暴发过程中,传统疫苗发挥了重要作用,但其自身却存在诸多弊端,因此研制新型疫苗来弥补传统疫苗的不足是很有必要的.文章主要对禽流感重组活载体疫苗、基因工程亚单位疫苗、DNA疫苗和病毒样颗粒疫苗等新型疫苗的研究进展进行综述,旨在为禽流感的防控提供参考.     

Potency of an experimental DNA vaccine against Aujeszky's disease in pigs

Intradermal vaccination with plasmid DNA encoding envelope glycoprotein C (gC) of pseudorabies virus (PrV) conferred protection of pigs against Aujeszky's disease when challenged with strain 75V19, but proved to be inadequate for protection against the highly virulent strain NIA-3. To improve the performance of the DNA vaccine, animals were vaccinated intradermally with a combination of plasmids expressing PrV glycoproteins gB, gC, gD, or gE under control of the major immediate-early promotor/enhancer of human cytomegalovirus. 12.5 microg per plasmid were used per immunization of 5-week old piglets which were injected three times at biweekly intervals. Five out of six animals survived a lethal challenge with strain NIA-3 without exhibiting central nervous signs, whereas all the control animals succumbed to the disease. This result shows the increased protection afforded by administration of the plasmid mixture over vaccination with a gC expressing plasmid alone. A comparative trial was performed using commercially available inactivated and modified-live vaccines and a mixture of plasmids expressing gB, gC, and gD. gE was omitted to conform with current eradication strategies based on gE-deleted vaccines. All six animals vaccinated with the live vaccine survived the lethal NIA-3 challenge without showing severe clinical signs. In contrast, five of six animals immunized with the inactivated vaccine died, as did two non-vaccinated controls. In this test, three of six animals vaccinated with the DNA vaccine survived without severe clinical signs, whereas three succumbed to the disease. Comparing weight reduction and virus excretion, the DNA vaccine also ranged between the inactivated and modified-live vaccines. Thus, administration of DNA constructs expressing different PrV glycoproteins was superior to an adjuvanted inactivated vaccine but less effective than an attenuated live vaccine in protection of pigs against PrV infection. Our data suggest a potential use of DNA vaccination in circumstances which do not allow administration of live attenuated vaccines.     

A combined bovine herpesvirus 1 gB-gD DNA vaccine induces immune response in mice

Although DNA vaccines have several advantages over conventional vaccines, antibody production and protection are often not adequate, particularly in single plasmid vaccine formulations. Here we assessed the potential for a combined vaccine based on plasmids encoding the membrane-anchored or secreted forms of bovine herpesvirus type 1 (BHV-1) glycoprotein B and D (gB and gD) to induce neutralizing and cell mediated immune responses in mice. Animals were injected by intramuscular, subcutaneous and intranasal routes. Mice immunized with the combined vaccine containing the secreted forms of BHV-1 glycoproteins developed higher titers of anti-BHV-1 neutralizing antibodies, compared to wild type gB/gD combined plasmids and to single plasmid injected groups. Cellular immunity was also developed in mice immunized with combined vaccines, whereas low or no response were observed in single plasmid injected animals. The data suggest the potential use of this combined vaccine in in vivo trials of calves, in order to evaluate its protective efficacy.     

The effect of eukaryotic expression vectors and adjuvants on DNA vaccines in chickens using an avian influenza model

Vaccination of poultry with naked plasmid DNA has been successfully demonstrated with several different poultry pathogens, but the technology needs to be further developed before it can be practically implemented. Many different methods can conceivably enhance the efficacy of DNA vaccines, and this report examines the use of different eukaryotic expression vectors with different promoters and different adjuvants to express the influenza hemagglutinin protein. Four different promoters in five different plasmids were used to express the hemagglutinin protein of an H5 avian influenza virus, including two different immediate early cytomegaloviruses (CMVs), Rous sarcoma virus, chicken actin, and simian virus 40 promoters. All five constructs expressed detectable hemagglutinin protein in cell culture, but the pCI-neo HA plasmid with the CMV promoter provided the best response in chickens when vaccinated intramuscularly at 1 day of age on the basis of antibody titer and survivability after challenge with a highly pathogenic avian influenza virus at 6 wk postinoculation. A beneficial response was observed in birds boostered at 3 wk of age, in birds given larger amounts of DNA, and with the use of multiple injection sites to administer the vaccine. With the use of the pCI-neo construct, the effects of different adjuvants designed to increase the uptake of plasmid DNA, including 25% sucrose, diethylaminoethyl dextran, calcium phosphate, polybrene, and two different cationic liposomes, were examined. Both liposomes tested enhanced antibody titers as compared with the positive controls, but the other chemical adjuvants decreased the antibody response as compared with the control chickens that received just the plasmid alone. The results observed are promising for continued studies, but continued improvements in vaccine response and reduced costs are necessary before the technology can be commercially developed.     

双顺反子表达质粒在DNA疫苗研究中的应用

简述了DNA疫苗的研究概况，介绍了真核双表达质粒的结构特点及其在基因佐剂和二价DNA疫苗中的应用情况，总结分析了双表达质粒的优点和存在的问题，并对其今后在DNA疫苗中的研究方向和前景进行了展望。     

禽流感DNA疫苗研究进展

禽流感（avian influenza,AI）是由禽流感病毒（AIV）引起的一种禽类烈性综合征,威胁动物和人类公共健康,严重影响中国养禽业发展,接种疫苗一直是控制禽流感病毒传播最有效的手段。基于基因工程技术的不断发展,各种新型疫苗相继研发并投入使用。其中,禽流感DNA疫苗具有安全性高、制备方法简单、易于储藏和运输等优点,受到了广泛关注。常见的禽流感疫苗有HA DNA疫苗、NA DNA疫苗、M DNA疫苗、NP DNA疫苗等。禽流感DNA疫苗是将含有目的基因序列的重组质粒导入动物细胞,诱导动物机体产生体液和细胞免疫应答。为了提高禽流感DNA疫苗的免疫效果,国内外学者通过添加合适的佐剂、将目的基因导入理想质粒载体、对抗原序列优化,增强DNA疫苗的转染效率和基因表达水平,取得了一定的研究成果。自DNA疫苗开始研发至今,H1、H3、H5、H7、H9等众多亚型禽流感DNA疫苗逐步研发。2018年,由中国农业科学院哈尔滨兽医研究所研制的禽流感H5亚型DNA疫苗获得国家一类新兽药证书,是中国首个获得批准的禽流感DNA疫苗,极大地推动了DNA疫苗的发展。文章主要论述了禽流感DNA疫苗的载体构建、免疫机制、佐剂和载体选择以及疫苗研发等方面的研究进展和创新,并对其应用前景进行简要分析,旨在为科研工作者研制新型禽流感疫苗提供新的思路和参考。