日本脑炎病毒及其疫苗的研究

乙型脑炎病毒又称日本脑炎病毒(Japanese encephalitis virus，JEV)，简称乙脑病毒，属于黄病毒科(Flaviridae)黄病毒属。由乙脑病毒引起的疾病乙型脑炎(简称乙脑)是严重威胁人畜健康的一种中枢神经系统的急性传染病。以蚊为媒介而传播，以高热和狂暴或沉郁等神经症状为特征。具有明显的季节性和一定的地理分布区，多发生于蚊虫较多的夏秋季节，属于自然疫源性疾病，病毒通常在蚊-猪-蚊等动物间循环。猪被认为是JEV最重要的自然增殖动物。怀孕母猪感染后导致流产和死胎，公猪感染后睾丸有急性炎症反应。     

携带猪乙型脑炎DNA疫苗减毒沙门菌的构建及其免疫原性

将乙型脑炎病毒E蛋白主要抗原片段基因串联,构建真核表达载体pCI-EAB.将重组质粒pCI-EAB转染Vero细胞,表达产物通过RT-PCR和间接免疫荧光试验可检测出目的基因的转录与表达.质粒pCI-EAB电转化入减毒猪霍乱沙门菌,观察重组菌的安全性、体内外的稳定性与免疫原性.动物试验表明,重组菌是相对安全和稳定的;重组菌S.C500/pCI-EAB体外培养时,D600值在0.8左右质粒是比较稳定的;以1×108CFU剂量的重组菌S.C500/pCI-EAB口服免疫小鼠,在二免后1周和三免后1周,重组菌抗体水平与S.C500/pCI-neo空载体组差异显著(P＜0.01);脾脏淋巴细胞增殖情况显示,重组菌对ConA有显著的反应性,且与对照组差异明显,同时CD3+、CD4+和CD8+T细胞的数量也显著高于对照组(P＜0.01).上述结果初步表明,以减毒沙门菌为载体的猪乙型脑炎DNA疫苗具有良好的安全性、稳定性和免疫原性,为猪乙型脑炎口服活菌疫苗的研制奠定了基础.     

狐狸脑炎病毒细胞培养灭活疫苗的研究

采用分离到的狐狸脑炎病毒ＦＥＶ－Ｈ作种毒，经ＭＤＣＫ细胞培养，经最终含量为０．２％的甲醛灭活，制备狐狸脑炎病毒细胞培养灭活疫苗。皮下接种疫苗１０ｍｌ，试验狐，貉，犬均无临床特异反应，局部吸收良好。最小免疫剂量为１６倍稀释的疫苗原液１ｍｌ，使用剂量定为４倍稀释的疫苗原液１ｍｌ，试验狐狸免疫后３０天，血清中和抗体效价达到高峰值１：１４５，能耐受张毒攻击。免疫后６个月，抗体水平降至１：３２，但此时仍能耐     

Safety of multiple,submucosal inoculations of a live attenuated strangles vaccine in pregnant mares

A live attenuated vaccine against Streptococcus equi was administered submucosally in the upper lip to 224 pregnant and healthy mares to evaluate its safety. After a primary immunisation the mares were inoculated every 3 months until foaling. As control group, 206 mares of the same breeding farm were administered the solvent of the vaccine submucosally. None of the 430 mares presented any clinical evidence of strangles and neither local nor systemic reactions to vaccination were noticed. There was no association between abortion and vaccination. Furthermore no case of S. equi infection was revealed at post mortem examinations of aborted foals.     

Serologic responses of pregnant thoroughbred mares to vaccination with an inactivated equine herpesvirus 1 vaccine

The immunogenic potency and safety of a chemically inactivated equine herpesvirus 1 vaccine with added adjuvant was evaluated by testing serum-neutralizing and complement-fixation antibody responses of pregnant Thoroughbred mares. The vaccinated population comprised 321 pregnant mares on 7 farms; 3 in Normandy, France; 1 in Kildare, Ireland; and 3 in central Kentucky. The pattern of antibody response to vaccination was found qualitatively and quantitatively similar to that of pregnant mares previously vaccinated and determined by challenge exposure to be immune to abortigenic infection under experimentally controlled conditions. The safety of the vaccine was demonstrated by the occurrence of only 2 local untoward reactions to the administration of 941 IM injections.     

The effect of excess dietary iodine on pregnant mares and foals.

On a thoroughbred stud four foals were born with greatly enlarged thyroids and leg weakness. Two foals died within 18 hours of birth, the others subsequently recovered. An enlarged thyroid was also evident in one of the resident mares. The thyroids from the dead foals were hyperplastic. Feed analyses showed that the mares had an iodine intake of about 83 mg daily, 8-8 ppm of the dietary dry matter, due almost entirely to the high iodine content of a proprietary compound horse nut which had been fed at the daily rate of 12 lb per head. It was concluded from the histology of the thyroids, the high intake of iodine, the lack of response to treatment with potassium iodide and the elevated levels of serum protein bound iodine that the condition of the foals on the stud was caused by an excess of iodine fed to the mares during pregnancy.     

猪细小病毒灭活疫苗安全性试验及佐剂的筛选

本研究以猪细小病毒(PPV)现地分离株(BQ)第30代细胞培养毒株作为灭活疫苗研究用种毒,通过优化病毒增殖条件和培养方法,获得较高滴度的病毒传代细胞培养物用于PPV灭活疫苗的制备.PPV细胞培养毒株分别用甲醛和β-丙内酯进行灭活,对2种灭活剂灭活的病毒液分别用国产铝胶佐剂、进口矿物质白油佐剂以及法国赛比克公司的MONTANIDETM ISA 206、ISA15AVG、IMS251CVG 3种佐剂制备10种灭活疫苗,然后分别进行10日龄乳鼠、60日龄仔猪、不同妊娠阶段母猪的安全性试验及成年豚鼠的免疫效果对比试验.通过比较不同灭活疫苗的安全性和对成年豚鼠的免疫效果,初步确认甲醛灭活的病毒液与佐剂ISA15AVG的组合为PPV灭活疫苗的最佳灭活剂和佐剂组合.     

猪流行性乙型脑炎病毒B细胞多表位抗原的表达及免疫原性分析

流行性乙型脑炎病毒(JEV)可引起猪的繁殖障碍性疾病。本文利用基因重组技术,选取JEV prM/M、E、NS1蛋白的6个B细胞抗原表位,设计B细胞多表位基因,构建原核表达质粒,在大肠杆菌BL21中表达融合重组蛋白rMEP。纯化的rMEP免疫小鼠,通过抗体、中和抗体及亚型检测研究其免疫原性。结果表明,rMEP在上清中高效表达,Western blot结果证实rMEP具有较好的抗原性。在小鼠免疫模型中,rMEP免疫组小鼠血清中产生较高水平的针对rMEP的抗体和特异性中和抗体,且rMEP可诱导产生IgG1和IgG2a抗体,但以IgG1为主。这些结果表明,构建的JEV B细胞多表位蛋白能够刺激小鼠产生较强的体液免疫应答,为研究JEV的多表位疫苗提供新的思路和参考。     

Safety and immunogenicity of a vaccine bait containing ERA strain of attenuated rabies virus.

Ninety percent of foxes fed commercial ERA vaccine in a specially designed bait developed rabies serum neutralizing antibodies. The vaccine bait did not cause clinical signs of rabies when consumed by foxes, raccoons, skunks, dogs, cats, cattle and monkeys. When presented, in the laboratory, to wild rodents of the species Microtus, Mus musculus and Peromyscus, the vaccine baits caused vaccine-induced rabies only in Mus musculus. Laboratory mice of the CD-1 and CLL strain were susceptible to vaccine-induced rabies; however, studies showed that transmission of virus to other animals did not occur. These studies suggest that the vaccine bait described could be useful in a rabies control program in areas where foxes and wild dogs are the principal vectors.     

Safety of a temperature-sensitive vaccine strain of bovine viral diarrhea virus in pregnant cows

Safety tests were conducted in 78 pregnant cows vaccinated with a commercial preparation of a temperature-sensitive vaccine strain of bovine viral diarrhea (BVD) virus. After vaccination, seroconversion was detected in 33 (97%) of 34 cattle that did not have antibodies against BVD virus. Overall, 43 (91%) of 47 cows with prevaccination titers less than or equal to 4 seroconverted. During the test period, cows did not become naturally infected with BVD virus, and BVD-associated reactions to the vaccine were not observed in vaccinated cows. Calves born to vaccinated cows did not have clinical signs of fetal BVD. Precolostral blood samples collected from the progeny of cows that were seronegative at vaccination were free of antibody against BVD virus. Bovine viral diarrhea virus was not isolated from the cattle evaluated in the present study.     

Improved immunogenicity of Newcastle disease virus inactivated vaccine following DNA vaccination using Newcastle disease virus hemagglutinin-neuraminidase and fusion protein genes

The present study describes the development of DNA vaccines using the hemagglutinin-neuraminidase (HN) and fusion (F) genes from AF2240 Newcastle disease virus strain, namely pIRES/HN, pIRES/F and pIRES-F/HN. Transient expression analysis of the constructs in Vero cells revealed the successful expression of gene inserts in vitro. Moreover, in vivo experiments showed that single vaccination with the constructed plasmid DNA (pDNA) followed by a boost with inactivated vaccine induced a significant difference in enzyme-linked immunosorbent assay antibody levels (p < 0.05) elicited by either pIRES/F, pIRES/F+ pIRES/HN or pIRES-F/HN at one week after the booster in specific pathogen free chickens when compared with the inactivated vaccine alone. Taken together, these results indicated that recombinant pDNA could be used to increase the efficacy of the inactivated vaccine immunization procedure.     

日本脑炎病毒弱毒疫苗株全长cDNA的体外合成及恢复病毒的拯救

将病毒的全基因组分成3个重叠的区段分别扩增出来,把这3个片段连接到载体中。以这3个片段为模板,通过融合PCR方法,获得JEV的全长cDNA。以cDNA为体外转录的模板,体外转录获得病毒mRNA,转染BHK-21细胞,拯救JEV病毒。通过生物学特性、分子生物学、蛋白水平等几个方面对恢复病毒进行鉴定,并测定恢复病毒的生长曲线和LD50。结果显示,获得了全长cDNA,体外转录获得的病毒RNA转染BHK-21细胞后,二代恢复病毒可引起明显的细胞病变,间接免疫荧光试验和RT-PCR均为阳性。空斑试验表明,拯救病毒与原病毒空斑表型类似;恢复病毒与亲本毒相比在BHK-21细胞上生长更快;恢复病毒的LD50与亲本毒类似。     

Efficacy of Japanese encephalitis vaccine in horses.

The efficacy of Japanese encephalitis vaccine in horses has been described from the effect of mass vaccination on the local prevalence of the disease in horses in each district of Hokkaido, Japan.     

猪轮状病毒油乳剂灭活疫苗的制备及免疫原性试验

猪轮状病毒(porcine rotavirus)属呼肠孤病毒科(Reoviridae)、轮状病毒属(Rotavirus),是引起仔猪发生急性胃肠道传染病的重要病原,自1975年Woode和Bridge首次从猪体分离出猪轮状病毒以后,世界各国均有不同程度的猪轮状病毒感染报道.在我国猪轮状病毒感染亦十分普遍,李国平等在调查中发现仔猪1～10日龄阳性率42.4%～66%,10日龄到断奶阳性率83.2%～91.7%,断奶后阳性率为63.2%～72%[1].临床症状主要表现为恶心、呕吐、腹泻,成年猪一般呈隐性感染.鉴于猪轮状病毒感染普遍,危害大,对猪进行疫苗接种预防猪轮状病毒感染具有重要意义.     

Efficacy of a saponin-adjuvanted inactivated respiratory syncytial virus vaccine in calves

The objective of this study was to determine whether a commercially available, saponin-adjuvanted, inactivated bovine respiratory syncytial virus (BRSV) vaccine would protect calves from experimental infection with virulent BRSV. This was a randomized controlled trial comprising 14, 8- to 9-week-old calves seronegative for BRSV Group 1 calves (n = 8) were not vaccinated and group 2 calves (n = 6) were vaccinated on days 0 and 19 with an inactivated BRSV vaccine. All calves were challenged with virulent BRSV on day 46. Clinical signs, arterial PO2, and immune responses were monitored after challenge. Calves were euthanatized on day 54 (8 d after challenge) and lungs were examined for lesions. Vaccination elicited increases in BRSV-specific immunoglobulin (Ig) G and virus neutralizing antibody titers. Challenge with BRSV resulted in severe respiratory tract disease and extensive pulmonary lesions in control calves, but no signs of clinical disease and minimal or no pulmonary lesions in vaccinated calves. Arterial blood oxygen values on day 53 (7 d after challenge) in control calves were significantly lower than those in vaccinated calves, which remained within normal limits. Control calves shed BRSV for several days after challenge, whereas BRSV was not detected on deep nasal swabs from vaccinated calves. In summary, the results indicated that this inactivated BRSV vaccine provided clinical protection from experimental infection with virulent virus 27 d after vaccination and significantly decreased the prevalence and severity of pulmonary lesions. Efficacy was similar to that reported for other commercial inactivated and modified-live BRSV vaccines.     

猪链球菌多价灭活疫苗安全性和免疫原性试验

猪的链球菌感染已成为世界性的猪传染病。在国外 ,多由 D群链球菌的某些血清型特别是 2型猪链球菌 (Streptococci suis)引起猪的败血型和脑炎型链球菌病 [2、3、6、7、8]。而在国内对猪链球菌致病菌的研究证实 ,以 C群的兽疫链球菌为主 ,D群较少[1、4、5] ,E群链球菌则引起慢性链球菌病的关节炎和淋巴结炎 (脓肿 ) [2 ] 。近几年来我们发现 ,有些猪场对各类猪群均进行过链球菌苗的接种 ,但仍然大批发生链球菌病 ,造成大量死亡 ,后来经过对分离的致病菌作分群鉴定 ,同时对发病猪群采血检测群特异性抗体 ,结果发现免疫的疫苗抗原的菌群与当…