水貂病毒性肠炎、犬瘟热、伪狂犬病三联弱毒苗的研究——Ⅱ.水貂病毒性肠炎弱毒基础苗的研制

用猫细小病毒(FPV)作为水貂病毒性肠炎(ME)弱毒基础苗种毒,接种猫肾传代细胞,制出六批弱毒基础苗。疫苗毒价为4.0～5.5TCID_(50),HA≥128。接种水貂安全,第7天即产生37倍以上可抵抗 ME 病毒的免疫抗体.采取试验貂粪便作 HA 检查,从接种后第3天开始至第7天止,有95%的貂粪便排出 FPV,但对同居饲养的未免疫貂无致病力。应用该弱毒基础苗进行田间试验,接种的977只貂、貉、犬无一只因注苗发病。试验证明、用 FPV弱毒株作种毒制作的 ME 弱毒基础苗,安全性可靠,免疫原性良好,可作为 ME、犬瘟热,伪狂犬病三联弱毒苗中基础苗之一。     

水貂犬瘟热,病毒性肠炎和肉毒梭菌中毒三联疫苗的研究

用FPV弱毒、CDV弱毒和肉毒梭菌C型疫苗研制成功了水貂三联疫苗。该苗安全可靠、免疫原性强、免疫期达6个月以上,同各单苗相比抗体产生期及效价相近,室温可保存3天;4℃可保存2周;-20℃可保存6个月。该苗的免疫剂量为2ml,其中CDV和FPV弱毒滴度分别为10~(3.0),肉菌梭菌C型疫苗为2mg/ml。现场应用26000余只水貂,抗体阳转率达90％以上。     

水貂肠炎弱毒及用于三联苗的安全性与抗原性研究

水貂肠炎弱毒疫苗是将强毒在猫肾细胞培养上连续传代后培育成功的。67代后测定证明弱毒是安全的。疫苗病毒毒力稳定,复归本动物连续6代,不反强。虽注苗后数天内可从貂体组织及粪便中分离出疫苗病毒,但在注苗水貂和未注苗水貂密切接触3个月的期间内,未发现注苗木貂将病毒散播给未注苗的水貂。此弱毒苗无论单独使用或与貂的犬瘟热疫苗、肉毒梭菌C型β类毒素联合免疫均可使水貂产生良好的兔疫应答。     

水貂病毒性肠炎、犬瘟热、伪狂犬病三联弱毒苗的研究——1.猫细小病毒弱毒株的分离与鉴定

将引进的猫细小病毒(FPV)、鼻气管炎病毒(FKV)和杯状病毒(FCV)三联弱毒疫苗经理化学方法处理,通过猫肾传代细胞系 FK_(81)细胞培养,分离到 FPV 弱毒株,分离毒株的血清学及理化学特性与猫细小病毒一致;在 FK_(81)细胞上出现的病变较规律,毒力稳定;与水貂病毒性肠炎病毒有密切的亲缘关系;对水貂、貉和猫具有良好的免疫原性与安全性,是较理想的弱毒株.     

FPV细胞灭活苗与MEV细胞灭活苗对水貂病毒性肠炎免疫效力的比较

水貂病毒性肠炎(MVE)是由水貂病毒性肠炎病毒(MEV)引起的一种急性传染病,具有较高的发病率和死亡率,是世界公认的水貂三大传染病之一。 猫泛白细胞减少症病毒(FPV)和MEV同属于细小炳毒科细小病毒属,血清学试验、限制性核酸酶切图谱分析和单克隆抗体均证明两者之间有密切的亲缘关系。因此,     

犬冠状病毒YS1株细胞培养弱毒的实验免疫研究

应用纯化后的犬冠状病毒 (CCV)YS1株细胞培养致弱的弱毒 (CCVYS1V60 ) ,经口鼻和肌肉接种CCV ,SN抗体小于 1∶2的易感犬作安全试验 ,结果未见任何CCV临床症状与病理解剖学改变 ;用不同剂量的该CCV弱毒分组免疫CCV易感犬 ,经SN抗体测定与用CCV强毒攻毒试验 ,结果SN抗体达 1∶60以上的犬 90 %以上可获得免疫保护 ;应用该CCV弱毒分别免疫母源抗体达 1∶4和 1∶8的 2组试验犬 ,第 1次免疫 1 4d后SN抗体均未见升高 ,追加 2～ 3次免疫后 ,SN抗体才逐渐上升至 1∶60以上的免疫保护水平 ;用CCV易感犬和猫肾传代细胞 (CRFK)分别将该CCV细胞培养弱毒连续传 5代和 2 0代 ,结果对犬仍然安全 ,免疫原性也未见下降 ;与犬瘟热病毒 (CDV)、犬传染性肝炎病毒 (ICHV)和犬细小病毒(CPV)弱毒作免疫互扰试验 ,结果与各弱毒的单独免疫结果未见差异 ;该毒的免疫期在 1年以上 ,- 2 0℃冻结保存的保存期为 9个月。     

水貂犬瘟热Vero细胞弱毒、肉毒梭菌(C型)二联苗的研制及免疫研究

为适应我国毛皮动物养殖业发展的需要,本文进行了水貂犬瘟热Vero细胞弱毒与肉毒梭菌(C型)二联混合苗的研制和免疫研究。经3年多实验室和田间免疫试验,以及二联苗中试生产,水貂CDV—Vero细胞传代毒,肉毒梭菌(C型)二联苗已通过30多万只水貂的免疫观察;效果可靠、使用安全、并减少了接种次数、剂量。     

猪瘟兔化弱毒ⅢC_3克隆株的培育与研究——Ⅰ.应用终点稀释-荧光抗体技术克隆猪瘟兔化弱毒

应用终点稀释-荧光抗体技术获得了克隆化弱毒株(ⅢC_3株)。ⅢC_3克隆株在PK_(15)细胞上传至15代,用DIF试验测定各代次毒价,TCID_(50)可达到10~(5.0)以上。经生物学鉴定试验表明,该克隆株保留了猪瘟兔化弱毒淋脾毒感染家兔出现的特异性热反应和对猪的安全性及良好的免疫原性。对猪最小免疫量达到10~(-5)稀释1ml,与淋脾毒一致。DIF试验与猪体测毒,两者之间存在着密切的相关性。     

雏鸭病毒性肝炎弱毒疫苗的研究——Ⅰ.弱毒疫苗的培育及实验室试验

水试验通过将鸭肝炎病毒(DNV)在鸡胚传代,培育出雏鸭用鸭病毒性肝炎(DVH)弱毒疫苗——鸭病毒性肝炎81代鸡胚化弱毒疫苗(简称BAU-1),并对该苗进行了实验室试验。 试验表明,BAU-1对雏鸭安全无毒性,稳定性好,在敏感鸭体内连传5代无毒力返强现象。经测定,BAU-1的免疫剂量为5×10~(9·4)ELD50/只(皮下免疫)。通过对免疫雏鸭的血清抗体滴度进行测定,证实该BAU-1疫苗对雏鸭的免疫保护期在6周以上。     

蛋鸡首免乙型脑炎病毒后卵黄抗体变化规律的研究

本试验旨在研究疫苗、免疫剂量和注射方式对卵黄抗体效价规律的影响,探讨制备抗猪乙型脑炎病毒卵黄抗体蛋鸡的最佳免疫程序。选用无免疫褐羽蛋鸡180只,随机分成18组,每组10只。1、2组均为对照组,注射无菌生理盐水;3~10组采用皮下和肌肉两种注射方式,并依次注射灭活苗0.2、0.5、1.0和1.5 mL;11~18组同样采取两种注射方式,依次免疫相同剂量弱毒苗。各试验组分别于免疫前1 d、免疫后第3、7、10、14、18、21和28天采集当日鸡蛋6枚,提取卵黄抗体,测定效价。试验结果显示,1~6、11、12组卵黄抗体效价均为0,未产生明显免疫应答;7~10组注射灭活苗后7 d,平均卵黄抗体效价达到峰值,抗体持续时间为14 d;13~18组注射弱毒苗后14 d达到最高值,抗体持续时间为21 d;注射剂量相同但注射方式不同的两组之间比较,卵黄抗体效价差异不显著（P>0.05）;相同注射方式,相同疫苗的各试验组间,随着免疫剂量的增加卵黄抗体效价逐渐加强,且差异显著（P<0.05）。以上结果表明,肌肉或皮下两种注射方式,蛋鸡产生明显免疫应答至少需要免疫灭活苗1.0 mL或弱毒苗0.5 mL。比较弱毒苗与灭活苗,灭活苗刺激机体产生的抗体速度较快,但维持时间较短;较少剂量弱毒苗就可刺激机体产生抗体,但速度慢、维持时间长。     

Dose response studies of acute feline immunodeficiency virus PPR strain infection in cats

The effects of virus dose on host response were evaluated for the PPR strain of feline immunodeficiency virus (FIV-PPR). Specific pathogen-free cats were inoculated intravenously with 50, 250 or 1250 TCID(50) of FIV-PPR. Two weeks after inoculation, virus was detected in 10(6) peripheral blood mononuclear cells (PBMCs) of all infected animals, and the CD4(+):CD8(+) T lymphocyte ratios fell from greater than 2 to approximately 1 in all infected animals within the first 8 weeks after infection. Provirus detected in all groups using PCR and 10(3) PBMC was biphasic. Nine of 15 animals were positive between weeks 2 and 4 p.i. and 14 of 15 were positive by week 8 p.i. Transient lymphadenopathy was detected in most cats receiving 1250 TCID(50) and the 250 TCID(50) of virus, whereas no lymphadenopathy was detected in the 50 TCID(50) group or the five uninfected cats. Animals that had received the largest dose seroconverted earliest (on average at week 4.0) and those receiving the least seroconverted last (on average at week 5.6). Neither neutropenia nor lymphopenia were detected. FIV-specific CTL responses of memory effector cells could be detected in animals receiving all three doses but was highly variable among individual animals. Neurological manifestations determined after 15 weeks p.i. were observed in most infected cats, including two of the three that had received 50 TCID(50) of virus. However, the observed neurologic abnormalities were markedly less severe in the animals receiving the least amount of virus. Therefore, lymphadenopathy and neurologic signs of illness were less severe and seroconversion was slower in the animals that received the lowest dose compared with those receiving the 250 and 1250 TCID(50) doses of the FIV-PPR strain.     

Vaccination of turkeys with an avian pneumovirus isolate from the United States

Four-week-old poults obtained from avian pneumovirus (APV) antibody-free parents were vaccinated with different serial 10-fold dilutions of cell culture-propagated APV vaccine. The birds were vaccinated with 50 microl into each conjunctival space and nostril (total of 200 microl). Each poult of each group was vaccinated in groups that received doses of 4 x 10(4), 4 x 10(3), 4 x 10(2), 4 x 10(1), or 4 x 10(0) 50% tissue culture infective dose (TCID50) of APV vaccine, respectively. Respiratory signs were seen between 3 and 12 days postvaccination (PV) in the poults that were vaccinated with 4 x 10(4), 4 x 10(3), and 4 x 10(2) TCID50, respectively. In these groups, APV was detected from swabs collected at 5 days PV and seroconversion was detected at 2 wk PV. The groups that were originally vaccinated with 4 x 10(1) and 4 x 10(0) TCID50 developed mild clinical signs after vaccination, but neither virus nor antibody was detected PV. At 2 wk PV (6 wk of age), birds from each group, along with five unvaccinated controls, were challenged with APV. Upon challenge, the 4 x 10(4) and 4 x 10(3) TCID50 groups were protected against development of clinical signs and were resistant to reinfection. The group previously vaccinated with 4 x 10(2) TCID50 developed clinical signs after challenge that were considerably milder than those seen in the groups that had previously been vaccinated with lower doses or no virus. Even though 4 x 10(2) TCID50 vaccine dose administered by intranasal ocular route resulted in infection, incomplete protection resulted with this pivotal dose. Upon challenge, the 4 x 10(1) and 4 x 10(0) TCID50 groups exhibited milder disease signs than those seen in the challenged unvaccinated controls. In these groups, APV was detected in preparations of swabs collected at 5 days postchallenge (PC) and seroconversion was detected at 2 wk PC. These results indicate that the dose of APV vaccine that causes protection is higher than that required to produce infection.     

Survival and immunization of raccoons after exposure to pseudorabies (Aujeszky's disease) virus gene-deleted vaccines

In a controlled experiment, 16 wild-trapped raccoons were exposed to 1 of 2 genetically modified live pseudorabies virus (PRV) vaccines used in swine. One vaccine had genes deleted for thymidine kinase (TK(-)) and glycoprotein G (gG(-)); the other had an additional deletion for glycoprotein E (gE(-)). These vaccines were administered orally and intranasally at four dose levels: 10(3), 10(4), 10(5), and 10(6) TCID(50). The 21 days survival rate was 37.5% for the gG(-)TK(-) vaccine; all of the survivors developed antibodies to PRV. All animals receiving the gG(-)gE(-)TK(-) vaccine survived; 75% (all except the lowest dose) developed anti-PRV antibodies. Survivors were challenged intranasally with a 3.2x10(3) TCID(50) dose of the virulent wildtype PRV Shope strain. Two of the remaining three gG(-)TK(-) vaccinated raccoons survived the challenge; for the gG(-)gE(-)TK(-) vaccine, the survival rate was 50% (4/8). The raccoons with higher vaccine-induced antibody titers were more likely to survive the challenge with the virulent PRV; there was a 100% mortality rate for raccoons lacking detectable anti-PRV antibodies. This experiment indicates that exposure of raccoons to modified live gene-deleted PRV vaccines may result in an immune response, and that this immunity provides some protection against exposure to virulent virus.     

Assessment of efficacy of a bivalent BTV-2 and BTV-4 inactivated vaccine by vaccination and challenge in cattle

The efficacy of a bivalent inactivated vaccine against bluetongue virus (BTV) serotypes 2 (BTV-2) and 4 (BTV-4) was evaluated in cattle by general and local examination, serological follow-up, and challenge. Thirty-two 4-month-old calves were randomly allocated into 2 groups of 16 animals each. One group was vaccinated subcutaneously (s/c) with two injections of bivalent inactivated vaccine at a 28-day interval, and the second group was left unvaccinated and used as control. Sixty-five days after first vaccination, 8 vaccinated and 8 unvaccinated calves were s/c challenged with 1 mL of 6.2 Log10 TCID50/mL of an Italian field isolate of BTV serotype 2, while the remaining 8 vaccinated and 8 unvaccinated animals were challenged by 1 mL of 6.2 Log10 TCID50/mL of an Italian field isolate of BTV serotype 4. Three additional calves were included in the study and used as sentinels to confirm that no BTV was circulating locally. At the time of the challenge, only one vaccinated animal did not have neutralizing antibodies against BTV-4, while the remaining 15 showed titres of at least 1:10 for either BTV-2 or BTV-4. However, the BTV-2 component of the inactivated vaccine elicited a stronger immune response in terms of both the number of virus neutralization (VN) positive animals and antibody titres. After challenge, no animal showed signs of disease. Similarly, none of the vaccinated animals developed detectable viraemia while bluetongue virus serotype 2 and 4 titres were detected in the circulating blood of all unvaccinated animals, commencing on day 3 post-challenge and lasting 16 days. It is concluded that administration of the bivalent BTV-2 and BTV-4 inactivated vaccine resulted in a complete prevention of detectable viraemia in all calves when challenged with high doses of BTV-2 or BTV-4.     

重组鸡痘病毒vFV282活疫苗最小免疫剂量及攻毒保护试验

将重组鸡痘病毒vFV282疫苗用生理盐水作10^-1，10^-2，10^-3，10^-4系列稀释，分别免疫7天龄鸡，于免疫后21d，分别用NDV、IBDV和FPV攻毒，观察其保护率，结果除NDV攻毒在10^-4组保护率为40％（4／10），其余各组均为100％（10／10）保护。表明该疫苗的最小免疫剂量≤10^-3TCID50/0.02mL。     

SHIV-KB9病毒中国恒河猴细胞适应株的制备

试验旨在研究体外制备SHIV-KB9病毒中国恒河猴细胞适应株,在细胞水平和中国恒河猴体内评价其生物学特性。 试验将SHIV-KB9半长质粒连接后转染CEMx174细胞,转染上清与正常恒河猴PBMCs共培养。定期测定培养液中的P27抗原水平。当病毒复制达高峰期时收集培养上清,分装并冻存,测定病毒RNA载量和TCID₅₀。静脉感染中国恒河猴,研究该批次SHIV-KB9在体内的病毒学、免疫学指标变化及变异情况,分析其基本的生物学特性。结果表明,本研究共制备了95 mL SHIV- KB9病毒原液,病毒载量为2.678×10⁵ 拷贝/mL,TZM-bl细胞测定病毒的TCID₅₀为3.16×10³/mL,gp120序列分析表明病毒未发生变异。10倍稀释的3个不同浓度的SHIV-KB9均静脉成功感染中国恒河猴,引起外周血CD4⁺/CD8⁺大幅下降。因此,此次制备的SHIV-KB9细胞适应株生物学特性稳定,适合作为毒种库建立SHIV-KB9/中国恒河猴模型。     

膜分离法纯化浓缩猪伪狂犬病疫苗毒的效果

本研究使用不同孔径的陶瓷(有机)膜过滤器,对不合格的猪伪狂犬病毒细胞收获液(病毒含量≤104TCID50/mL)滤除杂蛋白、超滤浓缩、除菌处理得到纯化浓缩的猪伪狂犬病疫苗病毒液;然后对纯化浓缩的猪伪狂犬病疫苗病毒液分别进行杂蛋白去除率检验与无菌检验、病毒含量测定、安全检验、效力检验;将检验合格的纯化浓缩的猪伪狂犬病疫苗病毒液添加保护剂冻干,并对纯化浓缩的猪伪狂犬病冻干活疫苗进行以上各项检验,以及进行免疫猪体内抗体消长变化的检测。结果表明:纯化的猪伪狂犬病疫苗杂蛋白去除率平均达到68.3%以上,病毒含量≥105TCID50/mL,效力检验合格;免疫猪体内抗猪伪狂犬病毒抗体增长幅度比同时期未纯化的常规疫苗显著,其中免疫至84 d时中和抗体效价平均高达40.35稀释倍数左右,比常规疫苗中和抗体效价平均高出14.22稀释倍数。此项研究为畜禽疫苗的纯化提供一定的参考。     

Preliminary development of a live attenuated canine parvovirus vaccine from an isolate of British origin

Canine parvovirus isolated from a case of haemorrhagic enteritis in a breeding kennel in England was passaged and cloned in cultured feline and canine cells. No significant evidence of pathogenicity was found during six serial passages of the modified virus back through young dogs. The attenuated virus was excreted by inoculated animals and spread rapidly to uninoculated animals held in contact. When high titre attenuated virus was given to the six-week-old offspring of a seropositive dam a prompt seroconversion was observed. When the attenuated virus was used as an experimental vaccine in 108 pups in an infected breeding colony a highly significant improvement was obtained in the accumulated morbidity and mortality compared with a parallel group vaccinated with modified live feline panleucopenia virus.     

Inhibition of feline infectious peritonitis virus replication by recombinant human leukocyte (alpha) interferon and feline fibroblastic (beta) interferon

Replication of feline infectious peritonitis virus (FIPV) in feline cell cultures was inhibited after incubation of cells with either human recombinant leukocyte (alpha) interferon (IFN) or feline fibroblastic (beta) IFN for 18 to 24 hours before viral challenge exposure. Compared with virus control cultures, FIPV yields were reduced by ranges of 0.1 to 2.7 log10 or 2 to 5.2 log10 TCID50 in cultures treated with human alpha- or feline beta-IFN, respectively; yield reductions were IFN dose dependent. Sensitivity to the antiviral activities of IFN varied with cell type; feline embryo cells had greater FIPV yield reductions than did similarly treated feline kidney or feline lung cells. Comparison of the virus growth curves in IFN-treated and virus control cultures indicated marked reduction in intracellular and extracellular FIPV in IFN-treated cultures. Compared with virus control cultures, intracellular and extracellular infectivity in IFN-treated cultures was delayed in onset by 12 and 30 hours, respectively, and FIPV titers subsequently were reduced by 3 to 3.5 and 5 log10 TCID50, respectively. Frequently, immunofluorescent and electron microscopy of IFN-treated cells or cell culture fluids did not reveal virus; however, even in cultures without viral cytopathic changes, small amounts of virus occasionally persisted in cells.     

自制貉源抗血清治疗犬瘟热病的研究

应用含有104.5TCID50/0.1 mL犬瘟热弱毒培养物4 mL免疫特种经济动物乌苏里貉,10 d后采集血液分离血清,病毒用200TCID50与等量连续倍比稀释的血清和经饱和硫酸铵粗提的免疫球蛋白做中和试验鉴定抗体的效价,并对上述2种抗体效价进行比较。结果表明,2917倍稀释全血清、2344倍稀释饱和硫酸铵粗提的免疫球蛋白能保护50%Vero细胞不出现CPE。临床应用全血清治疗24只患犬瘟热病的白貉,22只成活。由此说明可以应用狐貉等特种动物制备的抗血清治疗犬瘟热病。