Quantification of water buffalo (Bubalus bubalis) cytokine expression in response to inactivated foot-and-mouth disease (FMD) vaccine

This study describes the quantification of cytokine expression of vaccinated water buffaloes with FMD inactivated vaccine. Using real-time PCR quantification assay, expression of Th1 (IL-2, IL-12p40, IFNγ); Th2 (IL-4, IL-10) and inflammatory (IL-6, TNFα) cytokines were quantified weekly for the entire three-week duration of the experiment. It was noted that IFNγ, IL-10 and TNFα had peaked on week three post-vaccination while the remaining cytokines peaked on the second week and decreased by the third week. The counteraction between IFNγ and IL-4 was noted as well as the possible suppressive action of IL-10 to that of IL-2 and IL-12, which is a common phenomenon between Th1 and Th2 cytokines. Synergy between TNFa and IL-6 was also observed. These findings suggest that within the immune system of water buffalo there is a dynamic cell-mediated and humoral interaction in response to immunogen. This assessment of the cytokine expressions is vital for the study of water buffalo disease progression and concurring protective immune responses.     

Risk factors for foot and mouth disease seroprevalence in indigenous cattle in Southern Ethiopia: the effect of production system

A serological survey to investigate risk factors for Foot and Mouth Disease (FMD) occurrence was conducted between October 2007 and March 2008 in Southern Ethiopia. Antibodies against non-structural protein of FMD virus (using 3abc ELISA) were measured as indicator of exposure to the virus. The seroprevalence of FMD was 9.5% (95%CI = 7.7 – 11.3, n = 1020) and 48.1% (95% CI = 36.8 – 59.4%, n = 79), respectively at animal and herd levels. Within herd seropositivity was ranged from 6.7 to 46.7% with 18.6% (95%CI = 14.6 – 22.5%) risk of being seropositive for an animal in positive herds. The most important herd level risk factors identified were pastoral system (OR = 16.3, 95% CI = 2.0 -133.7) compared to sedentary, low altitude (OR = 7.5, 95% CI 1.4 -40.7) compared to high altitude, keeping cattle with small ruminants (OR = 5.1, 95% CI 1.0 -25.2) when compared to one species or alone. Seroprevalence was significantly higher (P <0.05) in South Omo than Sidama and Gamo Gofa areas. The odds of seropositivity were 2.8 and 2.3 times higher in the adult (>4 years) and maturing animals (3–4 years) compared to young age category (<3 years). Both multivariable logistic and negative binomial regressions depicted that production system was the major risk factor for FMD seropositivity. Consequently, higher prevalence of FMD in pastoral system where animals are an integral part of life has substantial livelihood and economic implications, which signifies the need for devising control measures.     

携带抗口蹄疫病毒ScFv基因猪成纤维细胞系的建立

根据口蹄疫病毒的感染特征及其在临床发病过程中呈现的症状,筛选抗口蹄疫病毒单链抗体基因。通过EcoRⅠ/NotⅠ双酶切中间质粒pGEX-ScFv获得单链抗体基因片段,并克隆至反转录病毒载体质粒pFB-neo,构建反转录病毒穿梭载体质粒pFB-ScFv。将pFB-ScFv与辅助质粒pVPack-GP、pVPack-10A1共转染HEK293T细胞,包装重组反转录病毒MMLV-ScFv。同时,构建含增强型绿色荧光蛋白基因的重组反转录病毒MMLV-EGFP作为对照。利用所包装重组反转录病毒感染猪成纤维细胞,通过G418筛选获得携带目的基因的单克隆细胞集落。试验结果表明,本试验在成功构建反转录病毒穿梭载体pFB-ScFv和pFB-EGFP的基础上,获得了分别携带抗口蹄疫病毒单链抗体基因和增强型绿色荧光蛋白基因的反转录病毒,并最终筛选出分别携带上述基因的猪成纤维细胞克隆,为抗口蹄疫病毒转基因动物的研究奠定了基础。     

西藏牦牛O型、AsiaⅠ型口蹄疫疫苗免疫抗体效价分析

为调查口蹄疫疫苗对西藏牦牛的免疫保护情况,应用口蹄疫液相阻断ELISA方法对随机选取的西藏五区一市844头牦牛的血清进行O型、AsiaⅠ型口蹄疫抗体水平检测。结果显示,西藏地区牦牛口蹄疫O型、AsiaⅠ型疫苗抗体具有50%以上保护力的比例分别为83.44%、71%,口蹄疫疫苗免疫状况良好,但西藏同一地区不同县、市的牦牛口蹄疫疫苗免疫情况存在较大差异。     

Saponin Adjuvants. Vi. The Adjuvant Activity of Quil a in Trivalent Vaccination of Cattle and Guinea Pigs Against Foot-And-Mouth Disease

The saponin adjuvant Quil A was investigated in trivalent vaccination against foot-and-mouth disease with a concentrated vaccine based on BHK suspension cell virus of the serotypes O, A and G. The activity in cattle was estimated on the basis of seroneutra-lizing antibodies. Five and 10 ml doses with or without 1 mg of Quil A were each injected into 6 animals. Seroneutralizing antibodies were estimated at regular intervals during a period of 29 weeks. The activity in guinea pigs was estimated by experimental challenge. One ml doses of serial 4-fold dilutions of the vaccine with or without 50 µg of Quil A were injected into 24 groups of 20 guinea pigs. Challenge was given 3 weeks after vaccination.It was concluded that Quil A showed adjuvant activity in cattle and guinea pigs with all the serotypes used in the trivalent vaccination.     

牛口蹄疫Asia-Ⅰ型灭活疫苗LPB-E抗体滴度与免疫保护相关性研究

抽取7批牛口蹄疫Asia—Ⅰ型灭活疫苗，用OIE推荐的50％保护（PD50）试验方法进行免疫保护试验；同时，用OIE推荐的液相阻断酶联免疫吸附试验（LPB—E）检验程序测定了7批疫苗免疫保护试验的103头牛的血清抗体滴度。经过对获得数据的统计和处理分析证明，LPB—E抗体滴度与牛免疫保护率存在着显著的正相关性（P〈0．05），相关系数为0．91，概率分析表明，LPB—E抗体滴度≥2．1（1：126）时，95％的免疫牛能够抵抗与疫苗毒株同源的10^4ID50强毒的攻击。     

应用固相竞争ELISA、液相阻断ELISA和中和试验检测口蹄疫抗体的比较研究

为了评价建立的口蹄疫固相竞争酶联免疫吸附试验(solid phase competition ELISA,SPC-ELISA)方法与传统的中和试验(VNT)和液相阻断酶联免疫吸附试验(liquid phase block ELISA,LPB-ELISA)检测方法之间的关系,本研究对野外采集的不同种类的猪、牛和牦牛共94份血清样品分别进行O型口蹄疫病毒SPC-ELISA、LPB-ELISA和VNT检测,并分别比较3种方法的符合率和阳性检出率。结果表明,建立的O型口蹄疫病毒SPC-ELISA方法与VNT 具有很好的符合率,达88.3%,其中,猪和牦牛血清符合率高达90%和89.5%,阳性检出率为91.2%比LPB-ELISA更符合VNT,因此建立的SPC-ELISA方法比LPB-ELISA方法更符合VNT。     

实验性猪口蹄疫免疫病理形态学研究

选用健康仔猪13头,分成4组:健康对照组2头,健康攻毒组5头,免疫组和免疫攻毒级各3头。健康攻毒组经皮下注射猪口蹄疫病毒(FMDV);免疫组和免疫攻毒组均经皮下接种猪口蹄疫牛皮肤细胞弱毒苗(FMD-BC),观察1个月后,前者扑杀,后者再攻FMDV,1个月后扑杀。结果:健康攻毒组5头仔猪均出现口蹄疫(FMD)典型的临床症状和病现学变化,其淋巴结、脾脏不同程度的变性、坏死和出血,淋巴小结减少和缩小,淋巴细胞稀少,浆细胞呈坏死性变化,酯酶阳性(ANAE~ )细胞和次级淋巴小结均减少;免疫组未见任何FMD病理学变化,主要见全身各部位淋巴结及脾脏不同程度地增大,尤以肩前、髂下和脾门淋巴结更为显著,其淋巴小结显著增多、增大,淋巴细胞活化,大、中淋巴细胞增多,以过渡型和未成熟型浆细胞为主,ANAE~ 细胞和次级淋巴小结均增多;免疫攻毒组的变化与免疫组相似。     

口蹄疫研究进展

从口蹄疫的病原学、流行病学、临床症状、诊断、疫苗及免疫预防等多方面全面阐述口蹄疫的特点,特别突出地介绍了国际上(或国际组织)对该病的认识、流行情况以及在诊断和预防等多方面的相关规定,对于全面了解该病有参考价值.     

口蹄疫病毒重组鸡痘病毒活载体疫苗和DNA疫苗免疫牛的试验研究

将本课题组构建的共表达FMDV衣壳蛋白前体P1—2A基因以及蛋白酶3C基因的重组鸡痘病毒活载体疫苗vUTAL3CP1以及共表达P1—2A和猪白介索18基因的重组DNA疫苗（pVIRIL18P1），分别以单独及混合的共3种方式接种牛，然后通过间接ELISA、中和试验和T淋巴细胞增殖试验评价其诱导的特异性体液和细胞免疫水平。结果表明这2种基因工程疫苗均能诱导牛产生特异性的体液及细胞免疫应答。其中重组鸡痘病毒vUTAL3CP1免疫组以及联合免疫组（vUTAL3CP1／pVIRIL18P1，pVIRIL18P1／vUTAL3CP1）诱导的中和抗体滴度分别达到1：64、1：64和1：54，已接近于常规灭活疫苗水平（1：90），而特异性的T淋巴细胞增殖反应则比后者高得多（P〈0．01）。该研究结果为进一步进行免疫攻毒试验，并最终筛选出最佳疫苗和免疫程序奠定了基础。