山羊痘灭活疫苗免疫保护试验

将山羊痘GT4-STV42-56种毒在犊牛睾丸细胞上繁殖,收获毒液,用0.1%甲醛溶液灭活,加适量206佐剂,制成山羊痘油佐剂灭活疫苗.将疫苗以不同剂量皮下免疫接种不同种群的山羊.免疫后21 d,用AV40株强毒进行攻毒保护试验.结果显示,免疫剂量达0.5 mL时,疫苗对内蒙绒山羊的保护率为100%,对广西黑山羊的保护率为80%.实验表明,山羊痘灭活疫苗对山羊的免疫效果确实可靠.     

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。     

法氏囊疫苗免疫效果三种不同评价方法的比较

为了选出法氏囊疫苗免疫效果评估的合适方法,本研究使用酶联免疫吸附试验(ELISA)、琼脂糖扩散试验(AGP)和中和抗体检测3种方法对6种法氏囊灭活疫苗免疫后28 d的抗体水平进行评估,同时,对采血鸡进行攻毒,攻毒后4 d剖检所有鸡,观察法氏囊病变,评估3种抗体水平与攻毒保护之间的相关性;结果显示,ELISA和AGP方法检测免后血清结果相对较高,中和试验方法检测免后抗体值较低;ELISA和AGP方法检测疫苗免后抗体水平与攻毒保护结果基本一致,使用中和试验方法检测免后抗体与攻毒保护相关性较差;建议在实际生产中使用ELISA和AGP方法进行法氏囊疫苗免疫效果的评估。     

山羊痘灭活疫苗研究

用对山羊痘病毒敏感的原代或次代牛睾丸细胞进行病毒增殖培养,从中筛选出一株进行培养,将获得的病毒液用二乙烯亚胺作为灭活剂制备病毒灭活疫苗接种山羊。分别在接种后7、14、21、28及72d采集血清用琼脂扩散试验进行免疫抗体检测,均检测到较强的沉淀抗体;在72d对山羊进行强毒攻击试验未出现山羊痘病理变化。实验证明疫苗安全性良好,制备的山羊痘灭活疫苗对山羊有较强免疫保护作用。     

猪链球菌病氢氧化铝二价灭活苗免疫兔血清抗体消长规律

应用琼扩法 ( AGP)、间接 ELISA法检测猪链球菌病氢氧化铝二价灭活疫苗免疫兔的抗体水平。结果表明 ,免疫后 4d开始产生抗体 ,7d时免疫兔 1 0 0 %抗体阳转 ,1 5～30 d抗体效价达高峰。免疫后 35d血清抗体有所下降 ,二免后 5d抗体效价均显著升高 ,6个月后血清抗体在 1 :1 6 0 0。通过免疫抗体与攻毒保护相关性测试 ,抗体效价在 1 :80 0(含 1 :80 0 )以上能抵抗 1× 1 0 2 ～ 1× 1 0 3CFU马腺疫链球菌兽疫亚种强毒攻击。     

鸡传染性鼻炎二价油乳剂灭活疫苗免疫持续期的研究

应用3批鸡传染性鼻炎二价油乳剂灭活疫苗进行了免疫持续期的试验。结果表明，鸡群免疫1次后，对A型强致病力菌株的攻毒保护率为83%，对C型强致病力菌株的攻毒保护率为51.5%，在首免后4周左右进行二免，则免疫鸡群对A型菌的保护率在免疫后13个月仍然保持100%，对C型菌的保护率在免疫后9个月内可达到90.5%。免疫鸡血清抗体也均保持较高的水平，其中B-ELISA的A型抗体的检出率为83%-100%、C型的检出率为33%-87%，血凝抑制试验（HI）的A型抗体阳性检出率为87%-100%，C型抗体的检出率为62.8%-93%，血清平板凝集抗体的检出率为62.8%-100%，此结果充分显示了鸡传染性鼻炎二价油乳剂灭活疫苗的良好的免疫效力以及B-ELISA、HI及血清平板凝集试验较高的敏感性的特异性，研究还证实了HI抗体与攻毒保抗力的密切相关。     

鸭坦布苏病毒病灭活疫苗(HB株)免疫产生期和持续期的试验

评价鸭坦布苏病毒病灭活疫苗(HB株)免疫16日龄北京鸭的效力、免疫产生期和持续期。利用实验室制备的3批鸭坦布苏病毒病灭活疫苗(201401、201402和201403批),以0.5 mL/只的剂量分别经肌肉或皮下注射途径免疫16日龄DTMUV抗体阴性北京鸭,首次免疫后14 d按照同样的剂量和途径进行二次免疫。分别于首免后14 d,二次免疫后7、14、42、60d和100 d采血分离血清,用ELISA方法检测抗体。采血后以0.5 mL/只含500 DID50的剂量经胸部肌肉注射鸭坦布苏病毒进行攻毒,攻毒后2 d采血,分离血清进行病毒分离。3个批次疫苗免疫16日龄北京鸭,二次免疫后7 d均可检测到血清抗体,至60 d检测时抗体阳性率在80%以上。三批疫苗二次免疫后7 d即对试验鸭产生保护,保护率可达到60%以上,其中皮下注射途径的免疫持续期为60 d,肌肉注射途径的免疫持续期为100 d。鸭坦布苏病毒病灭活疫苗对16日龄北京鸭采用二次免疫的程序可获得良好的保护效果,胸部肌肉注射途径优于皮下注射途径。     

鸡新城疫灭活疫苗免疫攻毒试验与血清学(HI)试验的平行关系研究

对19批鸡新城疫(ND)灭活疫苗进行了效力检验,在测定PD50的同时,进行了HI抗体效价的测定,以寻找血清学与抗攻毒保护的平行关系.结果表明,免疫剂量与GMHI抗体效价和攻毒保护率呈正相关:免疫1/12.5剂量组共保护54/60(90%),GMHI抗体效价为5.6 log2;免疫1/25剂量组共保护152/191(79.6%),GMHI抗体效价为4.9 log2;免疫1/50剂量组共保护123/183(67.2%),GMHI抗体效价为4.0 log2;免疫1/100剂量组共保护46/139(33%),GMHI抗体效价为3.0 log2;对照组保护0/89(0),GMHI抗体效价≤2.0 log2.采用PD50测定的18批疫苗中,有15批合格,3批不合格;按照我国现有质量标准,即免疫1/25剂量攻毒的方法进行检验,19批疫苗中有16批合格,3批不合格;血清学方法检验19批疫苗中有17批合格,2批不合格.其中免疫攻毒效检方法与PD50测定方法的不合格疫苗批次完全相同,其中2批为免疫胚制造的疫苗,1批为进口的灭活疫苗,血清学检验不合格的2批疫苗包含在其中,三种检验方法结果基本一致.试验还发现,HI抗体效价和抗攻毒保护呈正相关:未免疫对照组鸡血清HI抗体效价≤2.0 log2,攻毒保护率为0(0/89);免疫ND灭活疫苗后如果血清HI抗体效价＜3.0 log2,攻毒保护率为3.5%(4/113);HI为3.0 log2,攻毒保护率为16.1%(14/87);HI为4.0 log2,攻毒保护率为86.7%(86/99);HI达到5.0 log2,攻毒保护率为97.2%(103/106),HI≥6.0 log2,攻毒保护率为100%(168/168).     

猪流感(H3N2亚型)油乳剂灭活疫苗的免疫及攻毒试验

用不同剂量的猪流感H3N2亚型油乳剂灭活疫苗免疫15头断奶仔猪,每组5头,四周后二免。另外3头为非免疫攻毒对照组。首免后第10周用同源病毒攻毒,分别在免疫前及免疫后2、3、4、6、8、10、11周采血,分离血清,用HI试验检测其抗体滴度。攻毒后通过免疫猪和非免疫攻毒对照猪临床、病理变化及病毒分离等方面的比较,评价疫苗的保护效果。结果中、高剂量组的抗体滴度明显高于低剂量组;免疫组和对照组攻毒前后体温无显著差异(P>0.05),免疫组病理保护明显好于非免疫攻毒对照组;鼻液和肺中的病毒分离率比较,免疫组明显低于非免疫攻毒对照组。本研究表明,用该疫苗免疫后猪对同亚型的病毒攻击有一定的保护作用。     

猪链球菌病氢氧化铝二联灭活疫苗猪体血清抗体消长规律

应用琼扩法 (AGP)、间接ELISA法检测猪链球菌病氢氧化铝二联灭活疫苗免疫猪的抗体水平。结果表明 ,免疫后 4天开始产生抗体 ,7天时免疫猪 10 0 %抗体阳转 ,15～ 30天抗体效价达高峰。免疫后 35天进行二免 ,二免后 5天抗体效价均显著升高 ,6个月后血清抗体仍维持在 1∶16 0 0。母猪二免可使仔猪获得高水平且持勺的抗体 ,35日龄其母源抗体仍保持在 1∶80 0。通过免疫抗体与攻毒保护相关性测试 ,抗体效价在 1∶80 0 (含 1∶80 0 )以上能抵抗 1× 10 7CFU马腺疫链球菌兽疫亚种和 1×10 8CFU猪链球菌 2型的强毒攻击     

Immune Effect of Inactivated Vaccines against Foot-and-Mouth Disease and Their Interference on Differential Diagnosis

Vaccination with inactivated vaccine is an important measure to prevent and control foot-and-mouth disease (FMD), however, the immune effect and antigenic purity of inactivated vaccines are two major concerns for the establishment and evaluation of FMD free zones with vaccination. In this study, four groups of FMD type O and type A bivalent inactivated vaccines from 3 FMD vaccine manufacturers (designated as A, B and C) were selected to inoculate healthy juvenile cattle of FMD free. All cattle were immunized 3 or 4 times at a 1-month interval. Serum samples were collected before and after 1 month of every vaccination to determine the level of antibody to structural protein and non-structural protein. Results：(1) The qualified rates of antibody to structural protein: in group a1 (vaccine from company A, different batches), the antibody qualified rate could reach 100% for type O and type A, respectively after each vaccination. In group a2 (vaccines from company A, same batch), the antibody qualified rates were 36.7%, 98.3% and 100% for type O, and 15%, 86.7% and 100% for type A after the first to the third vaccination, respectively. In group b (vaccine from company B, same batch), the antibody qualified rates were 18.3%, 97% and 100% for type O, and 1.7%, 45% and 53.3% for type A after the first to the third vaccination, respectively. In group c (vaccines from company C, same batch), the antibody qualified rates were 26.7%, 96.7% and 100% for type O, and 21.7%, 71.7% and 100% for type A after the first to the third vaccination, respectively. (2) Antibody positive rate to non-structural protein 3ABC (confirmed with second ELISA test): In group a1, the positive rates were 0.7%, 1.4%, 9.5% and 4.8% after the first to the fourth vaccination, respectively; In group a2 and c, no 3ABC antibody-positive animal was detected after 3 repeated vaccination; In Group b, only one animal with a positive rate of 0.6% was detected after the third vaccination. The antibody qualified rates to the structural protein of FMDV in 3 of the 4 groups were far less than 70% after the primary vaccination, however, those were increased significantly after boost and repeated vaccination. The antigen purity of vaccines in three groups (a2, b and c) can meet the requirement of OIE standard on the FMD vaccine, however, the seroconversion to 3ABC antibody was obvious in animals from group a1 after repeated vaccination, which would cause some extent of interference to differential diagnosis. Also, a combination of a primary screening test and a confirmatory ELISA test can further improve the accuracy of differential diagnosis. This study provides an important scientific basis to make a rational program for establishment and evaluation of FMD free zone with vaccination.     

口蹄疫灭活疫苗的免疫效果及其对鉴别诊断的干扰

灭活疫苗免疫是防控口蹄疫的重要措施,但是灭活疫苗的免疫效果及其对感染与免疫鉴别诊断的干扰一直是口蹄疫免疫无疫区建设评估需要明确的重要问题。本研究中选择了3个企业（代号A、B与C）的4组口蹄疫O型与A型二价灭活疫苗,分别免疫口蹄疫抗体阴性健康未成年牛,免疫3～4次,测定免疫前后结构蛋白和非结构蛋白抗体的应答水平。结果显示：（1）结构蛋白抗体合格率：a1组（A企业多批次疫苗）4次免疫后O型和A型均为100%;a2组（A企业同批次疫苗）一~三免O型为36.7%、98.3%与100%,A型为15%、86.7%与100%;b组（B企业疫苗）一~三免O型为18.3%、97%与100%,A型为1.7%、45%与53.3%;c组（C企业疫苗）一~三免O型为26.7%、96.7%与100%,A型为21.7%、71.7%与100%。（2）非结构蛋白3ABC抗体阳性率（两种方法复核结果）：a1组一~四免分别为0.7%、1.4%、9.5%与4.8%;a2组和c组三次免疫均未检测到阳性;b组仅三免后阳性率为0.6%。3组灭活疫苗首次免疫牛的抗体合格率远不及70%,但加强免疫后抗体合格率均显著提高;非结构蛋白抗体检测结果表明有3组疫苗的抗原纯净度符合OIE的要求,但a1组灭活疫苗免疫后,仍然对感染与免疫鉴别诊断存在干扰;采用两种非结构蛋白抗体检测方法进行复核检验,可以提高感染与免疫鉴别诊断的准确性。本研究为口蹄疫免疫无疫评价方案的制定提供了科学依据。     

兔出血症-巴氏杆菌病二联灭活苗的研制

筛选出了具有良好免疫原性的兔瘟强毒株GMH881和兔巴氏杆菌C51-17株,研制出了兔出血症-巴氏杆菌二联灭活苗,对疫苗进行了免疫剂量、免疫产生期、兔出血症抗体测定、免疫保护期、保存期等试验：确定出二联苗免疫剂量1.0 mL;二联苗在免疫后5 d对兔病毒性出血症强毒的保护率达到100%,二联苗免疫后7d对兔巴氏杆菌的保护率均达到80%以上;二联苗的兔病毒性出血症抗体测定,结果表明,二联苗免疫家兔后7d HI抗体均可达22.75,15 d明显上升,二联苗HI效价30 d达到最高峰,可达28.0,二联苗在60~120 d均可维持在27.0~25.5水平,到180 d时抗体水平面略有下降为25.25,二联苗分别在免疫后4个月、6个月用兔出血症强毒攻击均产生100%保护,在6个月用2个MLD的兔巴氏杆菌攻击保护率可达70%以上,二联苗4~8℃条件下保存期暂定为1年,25℃条件下暂定为半年。     

上海地区活禽批发市场H9亚型禽流感病毒调查

为弄清上海地区活禽批发市场中H9禽流感病毒（Avian influenza virus,AIV）的流行情况及鸡群的免疫情况,2009年对上海三大活禽批发市场进行了采样监测。采用HI试验检测H9 AIV抗体、荧光RT-PCR试验和鸡胚接种分离鉴定病毒。共采集110批次1 646份血样和喉头泄殖腔棉拭样品,平均抗体合格率为60.27%,分离到H9病毒134株,其中4-6月和9-11月为全年中病毒分离的2个高峰期（样品带毒率均超过了10.00%）,明显比其它月份要高（其他月份均低于5.00%）,样品带毒率平均为8.14%。不同市场、不同地区采集的样品其抗体合格率和样品的带毒率也存在一定的差异。在30批分离到病毒的样品中,13批次已免疫H9N2油乳剂灭活苗且抗体合格率均大于70.00%的样品中分离到45株病毒（45/195）,其中6批次抗体合格率达到100%的样品中也分离到了病毒（8/90）,但带毒率明显比未经疫苗免疫的样品（79/255）低。调查结果表明养殖户对肉鸡群H9N2油乳剂灭活苗免疫重视程度不够,鸡群中带毒现象较普遍。疫苗免疫后能产生较高的免疫抗体,且抗体能减轻临床症状,降低带毒率,但不能完全阻止病毒复制,存在高抗体下带毒现象。     

Dose-response effects of inactivated Newcastle disease vaccines: influence of serologic assay, time after vaccination, and type of chickens

Knowledge of the dose-response relation of inactivated vaccines and of the factors that influence this relation is essential for the evaluation of existing vaccine potency assays and the development of new potency assays that are based on the antigen content of the inactivated vaccines. We quantified the relation between vaccine dose, serologic response, and clinical protection after vaccination for three different inactivated Newcastle disease (ND) vaccines. Qualitatively, similar dose-response curves were obtained for the three vaccines when either the serologic response or the clinical protection of specific-pathogen-free (SPF) chickens was plotted against the different vaccine doses applied. However, the vaccines differed quantitatively: doses of vaccines that induced similar antibody titers or clinical protection differed 2-8-fold. In contrast with the narrow range of antibody titers induced by a full vaccine dose, a very broad range of titers was obtained after dilution of the vaccines. At least 95% of the SPF chickens with detectable antibody in the serum were protected against a challenge with virulent Herts ND virus. The relation between the dosage of two different ND vaccines and the serum antibody titers remained markedly constant between 3 and 18 wk after vaccination. Vaccination of broilers instead of layers with a dilution series of inactivated ND vaccine resulted in significantly lower antibody levels and less clinical protection against virulent challenge. In conclusion, despite quantitative differences, we found comparable dose-response relations for the three inactivated ND vaccines studied.     

SEROLOGICAL RESPONSES IN PIGS VACCINATED WITH INACTIVATED PORCINE PARVOVIRUS

The safety and immunogenicity of inactivated porcine parvovirus (PPV) vaccines were investigated. Both beta-propiolactone and formalin successfully inactivated virus without destroying immunogenicity, which was considerably enhanced by incorporation of a gel adjuvant in the vaccine. Using the formalised-gel vaccine, initial antibody responses were demonstrated in susceptible piglets and adult pigs at 7 days after vaccination. These antibody responses persist at significant levels for at least 6 months after vaccination. Antibody levels increased up to 16 fold when revaccination was carried out. Vaccination of gilts with low level (passive) immunity resulted in antibody responses comparable to those recorded in susceptible pigs. The vaccine was safe as determined by absence of residual virus in the vaccine, absence of viraemia and excretion in vaccinted stock, and absence of effect on litters of sows vaccinated at different gestational ages. Vaccine stored at 4 degrees C for 6 months was as immunogenic as fresh vaccine.     

兔病毒性出血症、巴氏杆菌病二联蜂胶灭活苗的研制

用兔出血症毒(RHDV)和巴氏杆菌研制成蜂胶佐剂灭活苗,用1.0mL免疫试验兔,免疫后第5天,对RHDV保护率达100%;免疫后第7天对巴氏杆菌的保护率为72.5%,通过HI法监测兔病毒性出血症抗体水平,结果表明:蜂胶苗产生抗体时间较早,第7天后HI效价25.0,第20天效价显著上升达29.25以上,第30天达到最高峰,可达211.0,60~120 d均可维持在27.0~25.5水平,到第180天时抗体水平面略有下降为25.25,蜂胶苗分别在免疫后4个月、6个月用兔出血症强毒攻击均产生100%保护,在6个月用2个MLD的兔巴氏杆菌攻击保护率可达70%以上,二联苗4~8℃条件下保存期暂定为1年,25℃条件下暂定为半年。     

禽流感油乳剂灭活疫苗的研究

将6种不同亚型的禽流感病毒（AIVH2N9、H3N8、H5N1、H5N2、H7N1、H9N2）分别接种鸡胚,收获尿囊液,经甲醛灭活,以矿物油为佐剂制成油乳剂灭活疫苗。疫苗接种4和8周龄SPF鸡,注苗后均无不良反应。每种亚型疫苗免疫后14天和21天攻毒保护率均达90％-100％。分别用H5N1和H9N2亚型灭活疫苗免疫8周龄SPF鸡、25和28周龄健康商品蛋鸡,免疫后7天产生免疫力,14天保护率达100％,21天后抗体达高峰,仔鸡接苗后最高血凝抑制（HI）几何平均滴度（GMT）为7.3-8.0log2,蛋鸡为8.0-10.5log2。免疫后180天,抗体效价不低于6.5log2。免疫后180天分别以AIV攻击,H5亚型疫苗组,用强毒攻击无一发病和死亡,对照鸡全部发病死亡;H9亚型疫苗组,对照鸡在攻毒后72小时停产,免疫鸡无一发病且产蛋正常,对同源攻毒的保护率达100％。     

Response of feral cats to vaccination at the time of neutering

OBJECTIVE: To determine whether administration of inactivated virus or modified-live virus (MLV) vaccines to feral cats at the time of neutering induces protective serum antiviral antibody titers. DESIGN: Prospective study. ANIMALS: 61 feral cats included in a trap-neuter-return program in Florida. PROCEDURES: Each cat received vaccines against feline panleukopenia virus (FPV), feline herpes virus (FHV), feline calicivirus (FCV), FeLV, and rabies virus (RV). Immediately on completion of surgery, vaccines that contained inactivated RV and FeLV antigens and either MLV or inactivated FPV, FHV, and FCV antigens were administered. Titers of antiviral antibodies (except those against FeLV) were assessed in serum samples obtained immediately prior to surgery and approximately 10 weeks later. RESULTS: Prior to vaccination, some of the cats had protective serum antibody titers against FPV (33%), FHV (21%), FCV (64%), and RV (3%). Following vaccination, the overall proportion of cats with protective serum antiviral antibody titers increased (FPV [90%], FHV [56%], FCV [93%], and RV [98%]). With the exception of the FHV vaccine, there were no differences in the proportions of cats protected with inactivated virus versus MLV vaccines. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that exposure to FPV, FHV, and FCV is common among feral cats and that a high proportion of cats are susceptible to RV infection. Feral cats appeared to have an excellent immune response following vaccination at the time of neutering. Incorporation of vaccination into trap-neuter-return programs is likely to protect the health of individual cats and possibly reduce the disease burden in the community.