传染性法氏囊病病毒VP2基因原核表达及抗原性分析

从辽宁省某鸡场分离到一株传染性法氏囊病病毒(IBDV)。以该毒株基因组核酸为模板,应用RT-PCR扩增得到VP2基因,构建表达质粒pET28a-VP2,再将其转化至大肠埃希菌BL21(DE3)用IPTG进行诱导表达。表达产物经SDS-PAGE分析在48ku处出现特异性蛋白条带;经Western blot分析VP2蛋白可以与IBDV阳性血清发生特异性反应。用VP2蛋白制备的油乳剂疫苗免疫接种SPF鸡,2周后体内可以检测到特异性抗体。证明VP2蛋白具有重要的应用价值,为IBDV基因工程亚单位疫苗的研制奠定了基础。     

Network simulation modeling of equine infectious anemia in the non-racehorse population in Japan

An equine infectious anemia (EIA) transmission model was developed by constructing a network structure of horse movement patterns in a non-racehorse population. This model was then used to evaluate the effectiveness and efficiency of several EIA surveillance strategies. Because EIA had not been detected in Japan since 1993, it was appropriate to review the current surveillance strategy, which aims to eradicate EIA by intensive testing, and to consider alternative strategies suitable for the current EIA status in Japan.The non-racehorse population was divided into four sectors based on horse usage: the equestrian sector, private owner sector, exhibition sector, and fattening sector. To evaluate the risk of disease spread within and between sectors accompanied by horse movements, a stochastic individual-based network model was developed based on a previous survey of horse movement patterns. Surveillance parameters such as targeting sectors and frequency of testing were added into the model to compare surveillance strategies.The disease spread heterogeneously among sectors. Infection occurred mainly in the equestrian sector; the infection was less disseminated in other sectors. Therefore, we considered that the equestrian sector posed a higher risk of disease dissemination within and between sectors through horse movements. However, surveillance strategies targeting only the equestrian sector were not effective enough for early detection of the disease. Alternatively, targeting horses that moved permanently and those in the private owner sector in addition to the equestrian sector is recommended to achieve effectiveness equivalent to that of the current surveillance. In terms of surveillance efficacy, by increasing the testing interval (once yearly to once every 3 years), this testing scheme could reduce the number of tested horses to 44% of the current surveillance, while maintaining almost equivalent effectiveness. Intensive strategies targeting high-risk populations are considered to enhance effectiveness and efficiency of surveillance. The approach in this study may be helpful in the decision-making process that is involved in setting up strategies for risk-based surveillance.     

传染性支气管炎病毒非结构蛋白nsp5在真核细胞中的重组表达及鉴定

本文以嗜肾型鸡传染性支气管炎病毒(Infectious bronchitis virus,IBV)毒株的RNA为模板,通过RT-PCR扩增获得IBV非结构蛋白5(non-structural protein,nsp5)基因片段后,构建了杆状病毒重组质粒IBVnsp5-Bacmid。IBVnsp5-Bacmid转染Sf9细胞,获得nsp5重组杆状病毒,经(immunofluorescence assay,IFA)和Western blot检测到转染细胞表达的nsp5蛋白。进一步从感染的细胞中纯化重组蛋白,并用纯化的蛋白免疫小鼠制备了抗nsp5的多抗血清,该多抗血清可检测到IBV四川株SC021202感染的DF-1细胞中特异性的nsp5蛋白。结果表明IBV nsp5在Bac-to-Bac真核表达系统中获得了成功表达,而且具有良好的免疫原性和反应原性。     

鸡传染性支气管炎病毒分离株与疫苗株抗原相关性和S1基因序列分析

本实验挑选8株近年来山东省IBV分离株与疫苗株H120和491,利用SPF鸡分别制备了高免阳性血清,进行鸡胚交叉中和试验,计算其抗原相关系数,鉴定其血清型,并结合S1基因序列分析,研究其血清型和基因型之间的关系.结果显示:SDWF0608、SDLY0612、SDLY0701与疫苗株H120同属Mass血清型的不同亚型,其它5个分离株与H120和491不属于同一个血清型.结合S1基因序列分析发现,S1基因序列分析与血清中和结果二者不具有明显的相关性,8个分离株分属三个基因群,其中SDTA06111与H120等同属一个基因群,但血清中和实验显示它们不属于一个血清型;CK/CH/SD09/005与一个参考株独自构成一个基因群,其S1基因变异程度较大,并且能够与两个疫苗株发生交叉中和实验,但不属于一个血清型,它可能是一个基因重组后病毒.     

传染性法氏囊病疫苗免疫雏鸡局部体液的免疫学变化

应用间接ELISA法,检测了传染性法氏囊病疫苗免疫雏鸡4种局部体液(泪液、气管液、胆汁、肠液)免疫球蛋白IgG、IgM、IgA含量的动态变化。结果发现免疫雏鸡局部体液3种免疫球蛋白含量较未免疫对照雏鸡均有不同程度地升高;IBD超强毒株攻击后,对照雏鸡上述指标均明显低于疫苗免疫雏鸡,发生典型IBD,并全部死亡,而疫苗免疫雏鸡免疫保护率达60%,表明疫苗免疫后,可增强雏鸡的局部体液免疫功能。     

Causes of Encephalitis and Encephalopathy in Brazilian Equids

The etiology of diseases that affect the central nervous system (CNS) of equids was investigated. Samples (n = 218) collected from equids showing clinical signs of nervous or behavioral changes were analyzed, of which 37 (17.0%) were positive for rabies, 13 (6.0%) for the presence of protozoans (one Sarcocystis neurona, 12 Toxoplasma gondii), three (1.4%) for equine herpesvirus type 1 myeloencephalopathy, and 24 (11%) for bacterial encephalitis. Histopathology of the CNS revealed one (0.4%) case of cryptococcal myelomeningoencephalitis and 20 (9.2%) cases of equine leukoencephalomalacia. Central nervous system samples were positive for Sarcocystis neurona and Toxoplasma gondii by nested PCR-ITS1 followed by nucleotide sequencing. Diagnosis of equine herpesvirus 1 was confirmed by cell isolation and polymerase chain reaction followed by sequencing of the GD and polymerase (ORF 30) genes in three samples. No case of equine encephalomyelitis was diagnosed in samples analyzed by isolation in mice, VERO cell cultures, and RT-PCR for the nsP1 gene. Bacterial agents (Staphylococcus spp., Streptococcus spp., Bacillus spp., Enterobacteriaceae spp., Corynebacterium spp., and nonfermenting gram-negative bacillus) were detected in pure or preponderant cultures. Diagnosis was conclusive in 45% of samples, indicating that other infectious and noninfectious etiologies of encephalitis and encephalopathy should be considered for investigation.