FCV-VBS isolated from cats with typical symptoms caused VSD in experimental cats

Commercially available vaccines have been used widely to prevent feline calicivirus infection (FCI). However, with their widespread use, field strains, which are weakly cross-reactive with the live-virus vaccine strain F9, have posed the problem of vaccine breakdown. Recently the existence of FCV—associated virulent systemic disease (VSD) has been published. But their molecular diversity, antigenic mutations and physicochemical property have not been sufficiently clarified. Thus, we experimentally gave the vaccine breakdown strain (VBS) H10 to cats that had been inoculated with an F9 live vaccine. After the administration of strain H10, vaccinated cats (1 through 4) had no respiratory symptoms, whereas the non-vaccinated cat 5 showed clinical symptoms such as a fever of over 40°C, loss of vitality, decreased appetite, diarrhea, and nasal discharge after receiving strain H10, and died. Lethal FCV is rare, and may be a virulent systemic disease (VSD)—inducing strain. This is the initial report on VSD in Japan. It has been reported that symptoms of VSD were similar in vaccinated and nonvaccinated cats on experimental infection. However, no VSD-like symptoms developed, and the incidence of the disease varied depending on the presence or absence of vaccination, suggesting that there are two mechanisms of vaccine breakdown: one is associated with the vaccine immunity level, and the other is not. The characteristics of the VBS revealed were: (1) the duration of virus excretion was short when the originally carried antibody titer before virus challenge was high, (2) the excreted viral molecular species varied daily, not being limited to a specific species with time, and (3) the acquired physicochemical properties did not persist, and altered daily. FCV-VBS alters the molecular species and physicochemical properties daily due to the reduction of host immunity, which may lead to VSD.     

抗猫杯状病毒单克隆抗体的制备及生物学特性鉴定

为了制备抗猫杯状病毒的单克隆抗体,并对其基本生物学特性进行鉴定。分别采用饱和硫酸铵方法、差速离心、氯化铯密度梯度离心方法对猫杯状病毒(FCV)进行纯化,将纯化的猫杯状病毒作为抗原对BALB/c小鼠进行免疫,通过细胞融合和杂交瘤筛选技术建立抗猫杯状病毒单克隆抗体的杂交瘤系,鉴定单克隆抗体的亚型。结果表明：本实验获得了2株稳定分泌特异性抗猫杯状病毒单克隆抗体的杂交瘤细胞,分别命名为D8、E5,其亚型均为IgM。获得的单克隆抗体与犬细小病毒(CPV)、猫细小病毒(FPV)、犬瘟热病毒(CDV)均无交叉反应。成功制备了抗猫杯状病毒单克隆抗体,为建立相关诊断方法奠定了基础。     

烤烟成熟鲜烟叶生化组分高光谱估算方法筛选

为实现实时无损快速预测鲜烟叶的品质生化组分、筛选估算生化组分最佳的方法,使用ASD Fieldspec FR2500对烤烟成熟鲜烟叶进行了反射率、透射率光谱测定和常规生化组分分析。对可见近红外波段(350～1650 nm)单波段光谱和选用已有100种光谱指数共两类光谱参量进行了与生化组分之间线性函数、幂函数、指数函数共3种形式相关分析和基于决定系数的筛选。结果表明,对于叶绿素a、叶绿素b、类胡萝卜素、钾估算方法分别是Gitelson and Merzlyak2(GM₂) (R²=0.81)、光化学指数2(PRI₂) (R²=0.80)、Gitelson and Merzlyak2(GM₂) (R²=0.83)、1420 nm吸收峰开始位置(λs₁₄₂₀) (R²=0.67)的线性拟合最优。对于总糖、比叶重、氮、烟碱最优方法分别是在532 nm反射率一阶微分线性拟合(R²=0.54)、在1423 nm透射率线性拟合(R²=0.45)、在666 nm反射率倒数对数一阶微分的幂函数拟合(R²=0.44),在1135 nm反射率倒数对数二阶微分的线性拟合(R²=0.20)。通过筛选的光谱方法可以评估烟叶的品质状况。     

Sensitivity of FCV to recombinant feline interferon (rFeIFN)

Feline calicivirus cause feline respiratory diseases, and inactivated and attenuated vaccines are available for its prevention. Moreover, the presence of vaccine breakdown strains (VBS) is problematic. In Japan, feline recombinant interferon (rFeIFN) has been used for its treatment. However the method of compare with each strains has not established. To examine the relationship between the breakdown vaccine strain and rFeIFN sensitivity, the sensitivity of 47 field isolates to rFeIFN was determined. The Log PDD₅₀ values were normally distributed within the range 1.1–3.7, with a mean value of 2.3 ± 0.64. Since 68.3% of the PDD values fell in the range of the mean ± standard deviation, the values in the range 1.7–2.9, the lower values, and the higher values were defined as representing moderate, low, and high sensitivity, respectively. Among the 15 vaccine breakdown strains, strain Fukuoka9 showed a low sensitivity, but strains ML89, T58, and N74 were highly sensitive, showing no association with vaccine breakdown. The amino acid sequence changes specific to the low rFeIFN-sensitive Fukuoka-9 strain were found, suggesting that these sites are involved in rFeIFN sensitivity.     

上海市宠物猫杯状病毒、疱疹病毒及流感病毒的分离与鉴定

为了解上海市猫上呼吸道疾病病例中猫杯状病毒(FCV)、猫疱疹病毒1型(FHV-1)和猫流感病毒(FIV)的感染比例及其遗传变化特点,对上海市冬季53份表现上呼吸道症状宠物猫的眼结膜、口咽和鼻黏膜拭子,进行FCV、FHV-1和FIV分离与鉴定,并对分离的病毒进行遗传进化分析.结果显示:53份样品中,FCV分离率为58.4...     

检测猫杯状病毒RT-qPCR方法的优化与应用

为建立一种通用性良好的SYBR GreenⅠ荧光定量RT-PCR方法,根据GenBank上猫杯状病毒(feline calicivirus, FCV) ORF1保守区域序列,设计合成1对特异性引物,用于FCV的快速检测。灵敏性试验结果显示,该方法的检测下限为4.93×10¹拷贝/μL,比普通RT-PCR检测下限高100倍;特异性试验结果显示,该检测方法与猫疱疹病毒、猫细小病毒、猫传染性腹膜炎病毒等无交叉反应;通用性试验结果显示,针对引物靶位点区间,该方法对所有已知变异位点的人工合成质粒和本实验室保存的不同变异位点的FCV毒株,均能够良好检出;重复性试验结果显示,组内的变异系数为0.09%～0.71%,组间变异系数为0.05%～0.82%,均小于1%。对23份临床样品进行检测,该方法的阳性检出率为60.9%,检出的14份阳性病料分别接种到CRFK细胞中,对细胞病变产物进行扩增并测序,BLAST对比后结果证实为FCV毒株。结果表明,本试验建立的荧光定量RT-PCR检测方法可用于FCV的快速诊断及流行病学的调查。     

21世纪的汽车动力

描述了传统车辆和先进车辆的现状和未来发展趋势。在21世纪,由于社会对低排放、低能耗和石油替代资源的需求,先进车辆的技术竞争趋势增强了,比如代用燃料汽车、混合电动汽车、电动汽车和燃料电池汽车等等,总之,混合技术将发挥很重要的作用,因为它们不仅可以和内燃机结合,也可以用于燃料电池车。     

Detection of Feline calicivirus (FCV) from Vaccinated Cats and Phylogenetic Analysis of its Capsid Genes

We analysed genogroups of four feline calcivirus (FCV) isolates (FCV-S, H10, Ao198-1 and ML89) obtained from cats that experienced FCV infection after having been vaccinated against FCV. New PCR primer sets (8F/8R, Ao-S/Ao-A, cp-S/cp-A) were also designed, since the conventional Seal primer failed to amplify the target sequences in two samples. The genogroups of the four isolates as well as eight global and 17 domestic strains were determined by phylogenetic analysis of their amino acid sequences. One out of the four strains (25%) isolated in this study, H10, was grouped into genogroup I, along with the vaccine strains F9 and FCV-255. The other three isolates (75%) belonged to genogroup II. Thus, there were more isolates in genogroup II than in genogroup I. However, the antibody values of the four isolates against cat anti-F9 antisera were significantly decreased. There may be no relationship between the neutralizing antibody titre and genogroup. Amino acid sequence alignment of the four isolates showed that only a single amino acid in region C, which is involved in neutralization epitopes, was different in ML89 strain from that of F9. The other three strains, H10, Ao198-1 and FCV-B, shared the same amino acid sequence with F9. Alignment of amino acids for linear epitopes in the F9 strain, which are located at regions D and E, showed variations in 5' hypervariable region (HVR) of E, whereas D and conE had only synonymous substitutions i.e. no change in the amino acid sequence. This mutation in 5' HVR of region E suggested a vaccine breakdown, as the region is known to be essential for antigenicity. The genogroup II FCV is likely to be the cause of the FCV infection in this study, while the vaccine strains belong to genogroup I. Thus, the existing vaccine may need reevaluation for its effectiveness.     

Feline calicivirus VP2 is involved in the self-assembly of the capsid protein into virus-like particles

Feline calicivirus (FCV) is considered the most common upper respiratory tract disease (URTD) associated pathogen in cats. We previously expressed FCV VP1 capsid protein in insect cells by baculovirus system and we observed that this protein self-assemble into virus-like particles (VLPs) different in size and lacking the typical cup-like depressions of caliciviruses. In the present study, VP1 and the small basic structural protein VP2 of FCV were individually expressed by baculovirus system. Coinfection of insect cells with both recombinant viruses resulted in VP1 and VP2 self-assembly to form depressions similar to native capsids in size and appearance, demonstrating that VP2 interacts with the VP1 protein in the formation of VLPs.