猪口蹄疫O型合成肽疫苗免疫效果试验

为了更好地了解猪口蹄疫O型合成肽疫苗与灭活疫苗免疫后抗体水平之间的差异．在梅小市选择2个规模化养猪场,每个猪场分2组,对试验猪分别免疫猪口蹄疫O型合成肽疫苗和猪口蹄疫O型灭活疫苗．对接种疫苗的试验猪进行免疫应激观察,结果显示：注射合成肽疫苗试验猪未出现不良免疫副反应．说明用合成肽疫苗临床使用的安全性较高;采用猪O型口蹄疫VP1抗体检测试剂盒和正向间接血凝试验分别对使用上述两种口蹄疫疫苗免疫21d后的猪血清进行检测,结果表明：免疫猪口蹄疫O型合成肽疫苗用猪O型口蹄疫VP1抗体检测试剂盒检测．抗体阳性合格率达到92．50％,免疫猪口蹄疫O型灭活疫苗用正向间接血凝试验检测,免疫合格率为35．83％,说明猪口蹄疫O型合成肽疫苗接种后的免疫效果明显优于传统灭活疫苗。     

猪五号病浓缩苗和基因苗的免疫效果

应用本实验室建立的间接ＥＬＩＳＡ检测猪乙型五号病病毒抗体的方法，对浓缩型五号病来活苗（以下简称浓缩苗）和猪五号病病毒基因工程多肽苗（以下简称基因工程苗）的免疫效果进行监测。结果表明，两种疫苗均可达到较好的免疫效果。     

超前免疫──猪免疫上的新技术

超前免疫──猪免疫上的新技术时常发生注射过疫苗的猪在有效免疫期内仍染上猪瘟的现象。原因是免疫程序不科学。仔猪由于吸吮母猪初乳而获得的母源抗体干扰了猪瘟疫苗，可实行仔猪超前免疫，克服母源抗体干扰。超前免疫是仔猪产下后，完成擦身、断脐、剪牙、称重等接产工...     

蓝耳疫苗对猪群猪瘟抗体水平影响分析

应用酶联免疫吸附试验（ELISA）检测了12个猪场317份血清中的猪瘟和蓝耳抗体,其中免疫蓝耳疫苗的猪群猪瘟和蓝耳抗体阳性率分别为34．64％和90．2％,未免蓝耳疫苗的猪群猪瘟和蓝耳抗体阳性率分别为31．37％和74．39％,结果表明猪群蓝耳病毒的隐性感染和持续感染仍很严重。进一步对免疫和未曾免疫蓝耳疫苗的猪群进行猪瘟和蓝耳抗体阳性率比较,结果发现育肥猪和仔猪的猪瘟和蓝耳病抗体阳性率存在着反比关系,因此,蓝耳疫苗（活苗）的免疫对猪瘟抗体的产生有一定的抑制作用。     

福建省部分规模猪场猪圆环病毒病疫苗免疫效果评价与分析

为了解免疫不同圆环病毒病疫苗的抗体水平,给养殖户提供合适的免疫依据,对福建省不同地区8个猪场在使用不同猪圆环病毒病疫苗和免疫程序后猪群的抗体水平进行检测。结果显示:免疫场和未免疫场各阶段种猪群的猪圆环病毒病抗体阳性率皆为100%,差异不显著(P>0.05);但各阶段种猪群的S/P平均值,免疫场较未免疫场更高;同时通过比较变异系数发现,未免疫场种猪的抗体水平更活跃;14日龄免疫亚单位疫苗后抗体水平较高且维持时间较长,而免疫全病毒灭活苗后抗体水平仍持续下降。综上,种猪群免疫圆环病毒病疫苗是必要的,亚单位疫苗比全病毒灭活苗的免疫效果好。     

猪口蹄疫O型基因工程疫苗免疫保护试验

选取在1000L发酵罐规模的中试车间生产的3批猪口蹄疫O型基因工程疫苗,按照测定PD50的方法进行免疫保护攻毒试验.结果显示,3批猪口蹄疫O型基因工程疫苗的效力均在3个PD∞以上.试验表明,该猪口蹄疫O型基因工程疫苗可以进行大规模产业化生产.     

猪附红细胞体对仔猪猪瘟免疫效果的影响

为探讨猪附红细胞体对仔猪猪瘟免疫效果的影响，本试验利用猪瘟抗体检测ELISA试剂盒，对已注射猪瘟疫苗的69头感染猪附红细胞体的仔猪和31头无猪附红细胞体感染的健康仔猪进行了猪瘟抗体检测．结果表明，感染猪附红细胞体仔猪的猪瘟抗体水平低下，其猪瘟疫苗整体免疫合格率（49．2％）明显低于健康仔猪（93．5％），且显性感染仔猪的免疫合格率（41．6％）明显低于隐性感染仔猪（53．5％）、说明，猪附红细胞体严重干扰了猪瘟疫苗的免疫效果，且干扰程度随着猪附红细胞体感染程度的加深而更为明显。     

猪口蹄疫O型合成肽疫苗的免疫效果调查

应用猪口蹄疫病毒VP1结构蛋白抗体酶联免疫吸附试验（VP1-ELISA）对驻马店境内部分接种猪口蹄疫O型合成肽疫苗的猪群进行了血清学抗体检测,以掌握合成肽疫苗的免疫效果。4个规模化猪场血清检测结果显示其抗体合格率为96．6％,其中种猪95．3％、育肥猪97．8％,说明口蹄疫O型合成肽疫苗免疫效果较好。     

猪口蹄疫疫苗临床免疫效果评估

为了解2款口蹄疫疫苗免疫效果的差别,采用蔗糖密度梯度离心法测定疫苗抗原含量,并采用ELISA方法对猪群免疫前后进行相应抗体检测.结果表明,疫苗X的抗原含量显著高于疫苗Y.免疫疫苗X的母猪群,抗体阳性率及离散度变化均优于疫苗Y;仔猪免疫疫苗X后,抗体的阳性率快速升高并达到较高的效价,但是仔猪免疫疫苗Y后,抗体的阳性率出现...     

猪蓝耳病疫苗免疫剂量研究

在漯河市某养殖场随机选取2栋(保育13和保育17)健康状况良好的保育猪进行猪蓝耳病(PRRS)疫苗不同免疫剂量的效果试验,每栋舍中猪只随机分为四组(A、B、C、D组),分别免疫不同剂量的蓝耳病疫苗。每组猪群均随机选取15头仔猪标记,分别在免疫前、免疫后7、14、21、28 d进行采血,采用酶联免疫(ELISA)检测试剂盒检测PRRSV抗体水平。结果显示,猪群首次免疫蓝耳病疫苗后第2周抗体水平达到高峰,之后有所下降;不同剂量的疫苗免疫,猪群整体抗体变化趋势一致,说明低剂量的蓝耳病毒即可引起猪群的感染。     

O型口蹄疫抗体金标试纸盒检测免疫动物抗体效价与攻毒保护关系

本研究目的在于用O型口蹄疫金标试纸条研究免疫动物抗体效价与攻毒保护之间的对应关系以及免疫牛抗体消长规律.用金标试纸条检测了口蹄疫O型疫苗质量标准规定的牛、羊和猪免疫和攻毒血清抗体效价及不同免疫程序的牛血清效价.结果显示,当牛、猪和羊免疫血清稀释≥1∶8时,试纸条是阳性结果,表示有99％的被免疫动物属于抗体水平保护范围;被检测血清稀释≤1∶2时,是阴性结果,表示被免疫动物抗体水平属于不保护范围;血清稀释1∶2～1∶8时,是阳性结果,表示有50％的被免疫动物属于抗体水平保护范围.牛在首次免疫8周后加强免疫可以获得良好的保护率和保护时间.通过本试验确定了金标试纸条检测免疫动物抗体效价与攻毒保护之间的关系及牛加强免疫最佳的时间.     

猪囊虫病基因工程疫苗的研制及应用

从猪带绦虫六钩蚴cDNA文库中筛选目的基因并进行克隆表达，重组抗原用血清学方法和猪体免疫试验进行鉴定，筛选保护性抗原基因。将具有免疫保护作用的重组抗原纯化，并与免疫刺激复合物佐剂结合，制成猪囊虫病基因工程疫苗。对工程菌的培养发酵、重组抗原的下游纯化及疫苗的配制、疫苗的中间试制进行了研究，确定了一套比较完善的生产工艺。应用昆明小鼠建立了猪囊虫病的实验动物模型，应用该模型进行上述疫苗的免疫预防试验，免疫1次和2次的免疫保护率分别为96．9％和98．4％。本动物的免疫预防试验显示，疫苗安全性好，免疫保护率达92．2％，且免疫组发现的囊虫多数已死亡。在流行区进行了田间和区域试验。免疫猪未有不良反应，囊虫感染率分别由20％降为1．1％和5．4％降为0．21％。用该疫苗对人工感染的囊虫病猪进行免疫治疗，发现在感染早期的治疗效果较好。免疫动物的体液免疫和细胞免疫的检测结果显示，该疫苗刺激机体产生抗体的时间早、持续时间长、效价高；可显著提高淋巴细胞转化率及E－RFC和Ea-RFC细胞、ANAE^ 细胞和粗粒型ANAE^ 细胞、抑制／杀伤性T细胞亚群的数量。以上结果表明，用大肠杆菌表达的重组抗原制备的猪囊虫病基因工程疫苗安全、高效，且成本低廉，可规模化生产，有成为预防猪囊虫病的一种新型生物制剂。     

猪囊虫病基因工程疫苗的研制

从猪带绦虫六钩蚴ｃＤＮＡ文库中筛选目的基因并进行克隆表达,重组抗原用血清学方法和猪体免疫进行鉴定。钭具有免疫保护作用的重组抗原纯化,与免疫刺激复合物疫苗佐剂混合,制备成猪囊虫病基因工程疫苗,免疫仔猪并攻虫。该疫苗安全,无毒副作用,免疫猪减虫率９２．２％,完全保护率５５．５％,且免疫组发现的囊虫多数已死亡。     

规模化猪场种猪猪瘟免疫方式和剂量的探讨

对福建某公司2010年、2011年"一刀切"免疫(一年春、秋各免疫1次,每头每次免疫剂量1.5头份猪瘟兔化弱毒脾淋苗)的4561份种猪血清样品采用HerdChek猪瘟抗体检测试剂盒进行抗体检测,评价种猪猪瘟免疫效果,保证猪瘟免疫合格率达到80%以上,为规模化猪场种猪进行猪瘟免疫提供科学依据。结果显示:2010年、2011年种猪抗体检测合格率为86.70%、84.32%,对抗体不合格种猪再一次加强免疫后抗体检测总合格率91.28%、89.74%。由此表明:种猪猪瘟全群采用"一刀切"免疫猪瘟兔化弱毒脾淋苗,可以起到良好的免疫效果。     

Current status of foot-and-mouth disease vaccines including the use of genetic engineering

SUMMARY Foot-and-mouth disease virus (FMDV) vaccines are used to protect animals against infection by the 7 FMDV serotypes composed of greater than 60 FMDV subtypes. Because of problems of both live attenuated and inactivated FMDV vaccines and also because of the very large market for an effective safe vaccine, research into other types of vaccines has been undertaken. One of the 4 virus structural proteins, VP1, is believed to be the main protein that stimulates virus neutralising antibodies and studies have concentrated on its potential as a subunit vaccine. Genetic engineering has been used to clone the VF1 gene of FMDV and VP1 synthesised from the cloned gene has been used in experimental vaccine studies. The studies in small numbers of cattle and pigs demonstrated that 2 vaccinations with genetically engineered VP1 could confer protection against FMDV challenge. However, there are a number of areas that need further research before such a genetically engineered vaccine could be used commercially. The use of chemically synthesised antigenic fragments of VP1 has recently been reported, and these synthetic fragments appear to be potentially better at producing immunity to FMDV than the whole genetically engineered VP1 protein, perhaps because of conformational problems in the presentation of whole VP1. Other possible future directions in the research and in the development of safe, effective FMDV vaccines are discussed. In conclusion, although very significant progress has been made in cloning FMDV-VP1 genes, we are still far from a genetically engineered VP1-FMDV subunit vaccine. In the meantime, properly inactivated and safety-tested FMDV vaccines will continue to be used and to be of benefit to the livestock industry in countries where foot-and-mouth is endemic or in combating introductions of the disease.     

规模化种猪场猪瘟免疫情况调研

猪瘟是由猪瘟病毒引起的一种以高热、出血为主要特征的烈性、高度接触性传染病,至今仍在中国广泛流行。接种疫苗是防控该病发生的最根本的方法,为了查清山西省规模化种猪场猪瘟疫苗免疫情况,课题组采用ELISA试剂盒,对8个地市42个规模化种猪场465头哺乳猪、456头保育猪、436头育肥猪、419头母猪进行了猪瘟免疫抗体检测。查明了哺乳猪抗体阳性率平均为70.11%,保育猪抗体阳性率平均为40.57%,育肥猪抗体阳性率平均为50.22%,母猪抗体阳性率平均为69.69%,证明被检猪群免疫抗体不理想,尤其是保育猪抗体水平较低。同时对12个规模化种猪场,4类不同免疫方法免疫猪瘟疫苗的133头哺乳猪、110头保育猪、105头育肥猪、135头母猪进行了免疫抗体检测。结果表明:乳猪在吃奶前超免猪瘟高效细胞苗6头份,21日龄时二免猪瘟高效细胞苗4头份,60日龄三免猪瘟高效细胞苗2头份,母猪产后21 d跟胎免疫效果最好,使保育猪免疫抗体阳性率达到89.29%,有些猪场使用该方法免疫后,使保育猪的死亡率有了明显的降低,生长发育逐渐走向正常;而采用仔猪在断奶后28日龄至35日龄时首免猪瘟普通细胞苗4头份,60日龄二免猪瘟普通细胞苗2头份,母猪产后28 d跟胎免疫的方法效果最差,不宜推广应用。通过对山西一些规模化种猪场猪瘟免疫情况的调研,基本查清了规模化种猪场猪瘟免疫效果和最佳免疫方法,可为养猪场防控猪瘟提供依据。     

表达口蹄疫病毒P1基因的重组伪狂犬病病毒TK/gG-/PgG-P1的构建

为了构建口蹄疫与伪狂犬病二价基因工程疫苗株 ,将口蹄疫病毒 (FMDV) P1基因插入到伪狂犬病病毒 (PRV)通用载体 p Pg G- uni中 ,得到 PRV转移载体 p Pg G- P1。将转移载体与 Eco R 线性化后的 PRV弱毒疫苗株 TK- /g G- / lac Z 基因组 DNA共转染 PK- 15细胞 ,转染产物经多次空斑纯化和 PCR鉴定 ,获得了纯化的重组 PRV TK- /g G- / Pg G- P1,重组病毒基因组 DNA经酶切鉴定进一步表明 ,FMDV的 P1基因已成功地整合到 PRV弱毒疫苗株的基因组中。Western blot试验表明 ,FMDV P1基因在重组 PRV中得到表达。该研究为进一步研制口蹄疫与伪狂犬病二价基因工程疫苗奠定了坚实的基础     

猪口蹄疫○型合成肽疫苗诱导的免疫抗体的监测

设3组猪群,首免和二免分别注射生理盐水1 mL、猪口蹄疫0型合成肽疫苗1 mL(说明书推荐免疫剂量)及猪口蹄疫0型合成肽疫苗3 mL(说明书推荐免疫剂量3倍量),二免28天后检测猪群的口蹄疫抗体,结果显示3组猪群的抗体合格率分别为33%、78%和100%,疫苗诱导的抗体水平达到农业部规定的动物强制免疫标准。     

Difference in protective immunity of the tongue and feet of guinea pigs vaccinated with foot-and-mouth disease virus type A12 following intradermolingual and footpad challenge

Immunity to Foot-and-Mouth Disease Virus (FMDV) type A12 was developed in guinea pigs by vaccinating them with varying doses of inactivated FMDV inoculated by different routes. Vaccinated animals and FMDV convalescent animals were then tested for differences in protective immunity by challenging each animal with 10³ median guinea pigs infectious doses of FMDV intradermally into the tongue and the heelpad of a rear foot. Protected immunity was evaluated by examining the tongue and feet for the development of vesicles. Protective immunity of the tongue was found to be different from that of the feet. This led to the finding that three levels of protective immunity (LPI) could be distinguished. Convalescent guinea pigs demonstrated the highest level of protective immunity (LPI-1). They did not develop vesicles after a tongue or heelpad challenge. The second level of protective immunity (LPI-2) was shown by animals vaccinated with high doses of FMDV. After a tongue and heelpad challenge, they developed vesicles on the inoculated heelpad, but not on the tongue. The lowest level of protective immunity (LPI-3) was shown by animals vaccinated with lower doses of FMDV. After a tongue and heelpad challenge, they developed vesicles on the tongue and the inoculated foot, but FMDV did not spread to the uninoculated feet. Anti-FMDV antibody titers could not predict the level of protective immunity. Other experimental results show that protective immunity in the heelpad is complex and more difficult to induce than that of the tongue. In addition, the results show the feasibility of using double site challenges of the tongue and a foot for measuring protective immunity in guinea pigs.     

Evaluation of a 3A-truncated foot-and-mouth disease virus in pigs for its potential as a marker vaccine

Foot-and-mouth disease (FMD) is a highly contagious and economically devastating disease of cloven-hoofed animals in the world. The disease can be effectively controlled by vaccination of susceptible animals with the conventional inactivated vaccine. However, one major concern of the inactivated FMD virus (FMDV) vaccine is that it does not allow serological discrimination between infected and vaccinated animals, and therefore interferes with serologic surveillance and the epidemiology of disease. A marker vaccine has proven to be of great value in disease eradication and control programs. In this study, we constructed a marker FMDV containing a deletion of residues 93 to 143 in the nonstructural protein 3A using a recently developed FMDV infectious cDNA clone. The marker virus, r-HN/3A_93–143, had similar growth kinetics as the wild type virus in culture cell and caused a symptomatic infection in pigs. Pigs immunized with chemically inactivated marker vaccine were fully protected from the wild type virus challenge, and the potency of this marker vaccine was 10 PD₅₀ (50% pig protective dose) per dose, indicating it could be an efficacious vaccine against FMDV. In addition, we developed a blocking ELISA targeted to the deleted epitope that could clearly differentiate animals infected with the marker virus from those infected with the wild type virus. These results indicate that a marker FMDV vaccine can be potentially developed by deleting an immunodominant epitope in NSP 3A.