口蹄疫灭活疫苗与合成肽疫苗免疫效果对比试验

使用猪O型口蹄疫高效灭活疫苗与猪O型口蹄疫合成肽疫苗免疫28~35日龄商品猪,采用正向间接血凝试验和VP1结构蛋白抗体ELISA检测免疫前和首次免疫后14 d、28 d、35 d、42 d、56 d及二次免疫后7 d、14 d、28 d抗体水平。同时,对接种疫苗的实验猪进行免疫应激观察。结果显示:注射两种疫苗试验猪未出现不良免疫副反应;采用正向间接血凝试验检测,两种疫苗免疫抗体合格率差异不显著;运用VP1结构蛋白抗体ELISA进行检测,注射合成肽疫苗试验猪二免7 d后抗体水平快速上升,抗体检测合格率达到73%~94.12%。O型口蹄疫灭活免疫抗体不适宜用此检测方法。     

纳米磷酸钙作为猪瘟多肽疫苗佐剂的研究

为了探索纳米磷酸钙(NCaP)作为猪瘟多肽疫苗佐剂的可行性,本研究利用猪瘟多肽疫苗作为抗原,制备纳米磷酸钙吸附猪瘟多肽疫苗。将制备好的疫苗免疫兔后,分别检测兔抗体水平和攻毒后体温变化情况。检测结果显示,纳米磷酸钙对猪瘟多肽吸附率达70%;免疫后纳米磷酸钙佐剂组抗体水平低于弗氏佐剂,但与游离猪瘟多肽疫苗组相比,有较大提高;除弗氏佐剂组外,所有实验兔均有体温升高现象,纳米磷酸钙佐剂组体温升高的时间与空白组相比推迟16 h。结果表明,NCaP作为佐剂相对游离猪瘟多肽可提高其免疫效果,但其诱导的抗体水平低于弗氏佐剂组。     

猪O型口蹄疫合成肽疫苗及猪O型口蹄疫高效灭活疫苗免疫效果对比试验

使用猪O型口蹄疫合成肽疫苗与猪O型口蹄疫高效灭活疫苗免疫30-40日龄商品猪,免疫采用正向间接血凝试验、液相阻断ELISA及VP1结构蛋白抗体ELISA检测免疫前和首次免疫后14d、28d、35d、42d、56d及二次免疫后7d、14d、28d抗体水平。同时,对接种疫苗的实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;采用正向间接血凝试验检测,两种疫苗免疫抗体合格率差异不显著;运用液相阻断ELISA及VP1结构蛋白抗体ELISA进行检测,注射合成肽疫苗试验猪二免7d后抗体水平快速上升,抗体检测合格率达到73%-100%。与注射高效灭活苗组比较表明:合成肽疫苗临床使用的安全性较高,免疫抗体合格率明显优于猪O型口蹄疫高效灭活苗。     

山羊同时接种口蹄疫O-A-亚Ⅰ型三价灭活疫苗和小反刍兽疫活疫苗后30d内抗体动态研究

随机选择从未接种过小反刍兽疫(PPR)、口蹄疫(FMD)O-A-亚Ⅰ型三价灭活疫苗且经检测体内未含相关抗体的健康山羊40只,随机分成4组:第1组接种小反刍兽疫活疫苗,第2组接种口蹄疫O-A-亚Ⅰ型三价灭活疫苗,第3组同时接种小反刍兽疫活苗、口蹄疫O-A-亚Ⅰ型三价灭活疫苗,第4组不接种任何疫苗作为对照组,观察各组免疫副反应。在接种后7、14、21、28 d采血,用ELISA抗体检测试剂盒检测抗体动态。结果显示,接种后7 d时PPR单独免疫组和联合免疫组中PPR抗体阳性率均能达到80%;14~28 d时PPR抗体阳性率均能达到100%,PPR组和联合免疫组中同期PPR抗体OD450nm值之间比较差异均不显著;接种后第7天时FMD O-A-亚Ⅰ型三价灭活疫苗单独免疫组和联合免疫组中FMD O型抗体阳性率均能达到60%,14 d时FMD O型抗体阳性率均能达到100%;接种后14 d时FMD OA-亚Ⅰ型三价灭活疫苗单独免疫组和联合免疫组中亚FMD亚Ⅰ型和A型抗体阳性率均能达到国家规定的70%的标准。     

纳米氢氧化铝吸附禽流感病毒灭活疫苗的研制及其对肉鸡抗体效价的影响

自制纳米氢氧化铝佐剂,通过吸附H5N1灭活抗原,制备了纳米铝佐剂疫苗,按照中华人民共和国农业行业高致病性禽流感免疫技术规范标准,免疫肉鸡。经检测发现纳米铝佐剂疫苗组肉鸡产生有效保护抗体时间较油佐剂疫苗和常规铝佐剂疫苗提前3d和7d,抗体峰值达到时间较油佐剂疫苗和常规铝佐剂疫苗提前14d。结果表明,纳米氢氧化铝作为高致病性禽流感H5N1的免疫佐剂,可以更早的刺激动物产生有效保护抗体,纳米铝佐剂禽流感疫苗具有广泛的应用价值。     

接种猪口蹄疫O型合成肽疫苗的血清学调查分析

猪口蹄疫O型合成肽疫苗是采用固相多肽合成技术,在体外人工合成口蹄疫病毒主要抗原位点（合成肽）并连接人工T细胞位点,以此作为免疫原与进口佐剂混合配制而成油乳剂疫苗,它能够很好地预防猪O型口蹄疫的发生。传统灭活疫苗免疫后抗体的监测一般采用正向间接血凝法和液相阻断ELISA法,但由于灭活苗是全病毒疫苗,在一定程度上干扰了口蹄疫的诊断,     

猪口蹄疫O型合成肽疫苗免疫抗体消长规律研究

为了掌握遵义市猪口蹄疫O型合成肽疫苗免疫后抗体的消长规律,以便科学地使用猪口蹄疫O型合成肽疫苗,2014年在遵义市绥阳县某种猪场筛选30头55~73日龄的仔猪,注射猪口蹄疫O型合成肽疫苗,对其进行免疫抗体水平监测、免疫副反应和抗体维持时间调查。结果表明:仔猪免疫后7 d产生抗体,阳性率达73.33%,60 d达到峰值。猪口蹄疫O型合成肽疫苗免疫1次即可确保高效价抗体维持至150 d,说明猪口蹄疫O型合肽疫苗免疫原性高、生物安全性好、免疫不良反应小,在生产实践中值得推广。     

猪圆环病毒2型灭活疫苗(YZ株)的制备及对仔猪免疫效力的评价

运用不同佐剂,并配合不同剂型,制备猪圆环病毒2型(PCV2)灭活疫苗(YZ株),然后评价各疫苗对仔猪的免疫效力。制备的灭活疫苗包括:用10号白油制备油包水(W/O)剂型疫苗;用Montanide~(TM)ISA 206 VG佐剂制备水包油包水(W/O/W)剂型疫苗;分别用Montanide~(TM) ISA 15A VG和ISA 35 VG佐剂,制备水包油剂型(O/W)剂型疫苗;用Montanide~(TM) IMS 251C VG佐剂,制备水(W)剂型疫苗。除白油组外,其余4组疫苗安全性较好。5组疫苗免疫效力试验表明,ISA 206 VG(W/O/W)疫苗组首免后2周和3周PCV-2荧光抗体水平显著高于白油(W/O)和IMS 251C VG(W)疫苗组(P0.05);攻毒后猪血清中病毒PCR检测,ISA 206 VG(W/O/W)疫苗组14 d均为阴性,其他免疫组21~28 d转为阴性;猪淋巴结免疫组化检测,ISA 206 VG(W/O/W)、ISA 15A VG(O/W)疫苗组均未出现PCV-2阳性,且该两组中仔猪均未出现连续2 d以上的高热症状。以上结果表明,ISA 206 VG(W/O/W)PCV2灭活疫苗(YZ株)对仔猪具有较好的免疫保护效力,可进行后续的临床扩大试验。     

口蹄疫O型、A型二价灭活疫苗(OHM/02株+AKT-Ⅲ株)纯度、免疫持续期、攻毒保护的研究

用口蹄疫O型、A型二价灭活疫苗(OHM/02株+AKT-Ⅲ株)免疫12头6～9月龄口蹄疫抗体阴性牛,28日后二免,分别于接种后5 d、28 d、56 d、85 d、110 d采血,测定口蹄疫病毒非结构蛋白(NSPs)抗体及口蹄疫O型、A型ELISA抗体水平,并进行攻毒试验。试验牛免疫后5 d、28 d、56 d NSPs抗体阴性,疫苗纯度高。口蹄疫O型、A型ELISA效价28 d-110 d维持较高水平且达到99%以上保护效力,OHM/02株、A/XJNC/2010株攻毒100%保护,免疫过程安全性良好,无不良反应。     

口蹄疫纳米乳疫苗免疫持续期的试验研究

试验用同一批次相同抗原含量的口蹄疫病毒灭活抗原,分别采用纳米乳佐剂和ISA206佐剂配制口蹄疫O型、A型双价灭活疫苗,并对仔猪进行免疫,免疫后6个月内用液相阻断ELISA方法检测O型、A型免疫抗体效价(lg)。结果:免疫后6个月内纳米乳疫苗免疫猪产生的抗体lg平均值均高于ISA206佐剂疫苗,抗体水平在免疫后1个月达到最高。免疫后第6个月,纳米乳疫苗O型抗体lg平均值为2.19,A型抗体lg平均值为2.175,按液相阻断ELISA试剂盒的判定标准免疫猪抗体lg≥1.8,具有99%以上保护效力,纳米乳疫苗属于完全保护范围,而ISA206佐剂疫苗属于不完全保护范围。     

Development of a water-in-oil-in-water adjuvant for foot-and-mouth disease vaccine based on ginseng stem-leaf saponins as an immune booster

There has been an increasing interest in finding new formulations that qualify as vaccine adjuvants, which must be safe, stable, and have the capacity to stimulate a strong immune response. In this study, a basic formulation of a water-in-oil-in-water (W/O/W) adjuvant CV13 was developed, and ginseng stem-leaf saponins (GSLS) were added as an immune booster into oil phase. The physicochemical properties of the adjuvant were tested. Furthermore, the immune activity and the adjuvant effects, as indicated by the foot-and-mouth disease virus (FMDV) antigen were evaluated. The results showed that CV13 was similar in appearance to ISA 206 and could package FMDV antigen into a stable W/O/W emulsion. The FMD vaccine prepared with CV13 alone or CV13 containing GSLS achieved pharmaceutical characteristics comparable to a vaccine prepared with ISA 206, moreover the structural stability of the CV 13 vaccine was found to be better. Mice that were immunized with the FMD vaccine prepared with CV13 containing GSLS presented a significantly higher LPBE antibody titer and splenocyte proliferation rate than those immunized with a vaccine prepared with CV13 alone (p < 0.05). In addition, there was no significant difference between the groups that were immunized with FMD vaccine prepared with CV13 containing GSLS and ISA206 in terms of cellular and humoral immune response. In this paper, CV13 containing GSLS shows excellent immunologic adjuvant effect in mice model, and this new adjuvant may provide a potential choice for FMD vaccine production in the future.     

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.     

复方中药偶蹄康对奶牛口蹄疫疫苗抗体水平、血清细胞因子及生化指标的影响

为研究复方中药偶蹄康对奶牛口蹄疫O型、A型、亚洲Ⅰ型3个血清型疫苗抗体水平、10种血清细胞因子含量及生化指标的影响,本试验将健康成年奶牛分为药物组和空白对照组,每组100头,连续拌料给药7 d后注射口蹄疫三联苗,注射疫苗后0、14、28、56 d分别采血,检测血清疫苗效价、血清细胞因子含量及生化指标。结果显示,注射疫苗后0、28和56 d药物组与空白对照组抗体效价无显著差异（P>0.05）;注射疫苗后14 d药物对口蹄疫3种血清型抗体效价的提升作用均极显著（P<0.01）。药物组血清总胆红素（T-Bil）降低,0、28 d差异显著（P<0.05）,56 d差异极显著（P<0.01）;28、56 d药物组与空白对照组葡萄糖含量差异显著（P<0.05）;0 d血磷含量差异极显著（P<0.01）,但未超出正常生理值范围,且后续检测无显著差异（P>0.05）。注射疫苗后14 d药物组血清CD4含量显著高于空白对照组（P<0.05）;注射疫苗后28 d药物组血清CD4、IL-2、IL-6、IL-10、IL-12、IL-18、IFN-γ含量极显著高于空白对照组（P<0.01）;注射疫苗后56 d药物组血清IL-6、IL-12含量极显著高于空白对照组（P<0.01）,CD4、IL-2、IL-4、、IL-18含量显著高于空白对照组（P<0.05）。综上可知,偶蹄康对奶牛口蹄疫O型、A型、亚洲Ⅰ型3个血清型疫苗效价有显著提升,对奶牛肝脏和肾脏功能无明显的损伤,且有一定的利胆作用,对奶牛细胞免疫和体液免疫机能有明显的提升作用。     

Effect of Herbal Compound Outikang on Antibody Titer of Foot and Mouth Disease Vaccine，Cytokines and Biochemical Indices in Cows

The experiment was aimed to study the effects of herbal compound Outikang on antibody titer of foot and mouth disease vaccine type O,type A and type Asia Ⅰ,serum levels of ten cytokines and biochemical indices. 200 healthy adult cows were averagely divided into drug group and blank control group. Vaccinating the cows with FMD triplex vaccine when Outikang was given to the cows of drug group for 7 d running and then got the blood of all the cows on 0, 14, 28 and 56 d, respectively. Then blood biochemical examination, vaccine effectiveness and serum levels of cytokines were performed. The results showed that there were no significant differences in antibody titer of FMDV between blank control group and drug group on 0, 28 and 56 d after vaccination (P>0.05), while on 14 d the effect of Outikang on antibody titer of FMDV reached extremely significant level (P<0.01). Compared with blank control group, the contents of T-Bil in drug group was significantly decreased on 0 and 28 d (P<0.05), while it was extremely significantly decreased on 56 d (P<0.01).Compared with blank control group, the contents of Glu had significant difference on 28 and 56 d (P<0.05).The contents of P had extremely significant difference on 0 d (P<0.01), which was within the normal range. On 14 d after vaccination, the content of CD4 significantly increased in the drug group comparing to those in blank control group (P<0.05). On 28 d after vaccination, the content of CD4, IL-2, IL-6, IL-10, IL-12, IL-18 and IFN-γ extremely significantly increased in the drug group comparing to those in blank control group (P<0.01). On 56 d after vaccination, the contents of IL-6 and IL-12 extremely significantly increased in the drug group comparing to those in blank control group (P<0.01), while the content of CD4, IL-2, IL-4 and IL-18 significantly increased (P<0.05). Therefore, Outikang could significantly rise the antibody titer of foot and mouth disease vaccine type O, type A and type Asia Ⅰ.While it had no damage to the liver and kidney functions and had cholagogue effect, and it could enhance cellular immunity and humoral immunity function obviously.     

内蒙古地区牛羊口蹄疫灭活疫苗的免疫效果比较研究

为准确掌握牛羊口蹄疫疫苗免疫效果,2012年对内蒙古自治区的牛羊口蹄疫O—AsiaI型双价灭活疫苗和牛羊口蹄疫A型灭活疫苗免疫效果进行了监测。采用液相阻断ELISA方法。共检测了8个旗县23018份免疫牛羊口蹄疫O—AsiaI型双价灭活疫苗和牛羊口蹄疫A型灭活疫苗15日、30日、60日的血清中口蹄疫免疫抗体水平。结果显示,牛羊口蹄疫疫苗免疫效果总体良好,免疫抗体合格率超过了70％。规模化奶牛养殖场免疫效果稳定,散养地区免疫效果稳定性差,且不同厂家的疫苗免疫效果不同,加强免疫后A、B、D三个厂家抗体合格率能达到100％,C厂家的免疫效果较差,抗体合格率为93-3％。采用3ABC—ELISA检测试剂盒对部分地区牛羊进行感染抗体检测,结果为阴性。     

Detection of foot-and-mouth disease antigen in bovine epithelial samples: comparison of sites of sample collection by an enzyme linked immunosorbent assay (ELISA) and complement fixation test

Serially collected epithelial samples from lesions in the mouth and on the feet of calves experimentally infected with foot-and-mouth disease (FMD) type O1 BFS 1860 were assayed for the presence of FMD viral antigen using a double antibody sandwich enzyme-linked immunosorbent assay (ELISA) and a complement fixation (CF) test. The amount of infectious virus in each sample was also determined. FMD viral antigen was detected by ELISA in 70 per cent of the mouth samples and 92 per cent of samples from the feet. The CF test was less sensitive; it detected antigen in 44 per cent of mouth and 85 per cent of foot samples. In mouth samples the amount of antigen decreased rapidly becoming undetectable by the fourth day of sampling whereas in foot samples the quantity of antigen declined more slowly, and could be detected until the seventh day of sampling. Therefore it was concluded that the age of lesion and the site from which epithelial samples are collected are both important determinants in the laboratory diagnosis of FMD. In cattle, foot lesions are more likely than mouth lesions to yield antigen and to remain positive for a longer period.     

天峻县牛羊AsiaⅠ-O型口蹄疫免疫抗体检测

为了探讨口蹄疫O型-亚洲Ⅰ型二价灭活苗免疫接种牦牛、藏羊后产生抗体效价,项目对天峻县阳康乡的牦牛、藏羊开展AsiaⅠ-O型口蹄疫疫苗的抗体效价检测,试验结果表明,口蹄疫AsiaⅠ-O型二价灭活疫苗免疫效果依次为成年牦牛、成年藏羊、犊牛、羔羊;免疫后Ⅰ型抗体分析结果与O型免疫抗体的免疫效果一致,免疫合格率在80%以上,免疫牛羊产生的整体抗体水平能够符合国家规定的要求。     

牛O型、亚洲Ⅰ型口蹄疫疫苗不同免疫次数免疫后群体抗体合格率比较

本研究选取沈阳地区饲养管理正规的牛规模场,采用液相阻断ELISA试验方法测定牛血清样品中O型-亚洲Ⅰ型口蹄疫抗体水平,并计算免疫抗体的群体合格率。用O型-亚洲Ⅰ型二价苗一免、二免、三免的牛在免后28d时牛O型-亚洲Ⅰ型口蹄疫免疫抗体群体合格率研究结果表明,O型口蹄疫免疫抗体群体合格率分别为82.5%、100%、100%;亚洲Ⅰ型口蹄疫免疫抗体群体合格率分别为57.5%、97.5%、97.5%。一免后O型和亚洲Ⅰ型口蹄疫免疫抗体群体合格率下降速度均较快,一免后60dO型免疫抗体群体合格率降到72.5%,免后80d时O型、亚洲Ⅰ型口蹄疫免疫抗体群体合格率分别降为47.5%、12.5%。二免以上的牛口蹄免疫抗体群体合格率在28～120d内均较一免组高,而且抗体持续时间也长,三免后120d时O型、亚洲Ⅰ型口蹄疫免疫抗体群体合格率仍高达77.5%、75%。     

Antibody responses of sheep vaccinated with foot rot vaccines

Enzyme-linked immunosorbent assay (ELISA) and crossed immunoelectrophoresis (IEP) were used to investigate antibody responses of sheep vaccinated with a double adjuvanted or single adjuvanted commercial foot rot vaccine. ELISA detected an antibody response of greater magnitude to the double adjuvant vaccine compared with the single adjuvant vaccine. Sera from sheep vaccinated with double adjuvant vaccine recognised at least six antigens of Bacteroides nodosus in crossed IEP while sera from the single adjuvant vaccinated sheep recognised one antigen. The use of non-denatured antigens of B nodosus in ELISA and crossed IEP enabled quantitative comparisons of antibody responses to the different foot rot vaccines to be made.     

三种试剂盒检测猪口蹄疫O型抗体的对比试验

使用三种猪口蹄疫O型抗体检测试剂盒分别检测接种猪口蹄疫O型合成肽疫苗和猪口蹄疫O型灭活疫苗的免疫抗体,以探讨三种抗体检测试剂盒的相关性。结果发现,猪口蹄疫病毒VP1结构蛋白抗体ELISA试剂盒和猪口蹄疫O型液相阻断ELISA试剂盒都可检测猪口蹄疫O型合成肽疫苗,两种试剂盒的相关性达90.8%,而间接血凝试剂盒不能用来检测猪口蹄疫O型合成肽疫苗;猪口蹄疫O型液相阻断ELISA试剂盒和间接血凝试剂盒可用来检测猪口蹄疫O型灭活疫苗,两种试剂盒的相关性达93.7%,而猪口蹄疫病毒VP1结构蛋白抗体ELISA试剂盒不能用来检测猪口蹄疫O型灭活疫苗。