应用琼脂扩散试验检测新城疫抗原

应用ＰＥＧ平板进行琼脂扩散试验检测新城疫抗原,提高了对新城疫抗原的检出率。采用多器官检查和统一判定方法,即脾、胸腺、盲肠扁桃体、肠淋巴结和法氏囊等器官检出阳性时即可判为新城疫,对新城疫死鸡的检出率为１００％,解决了非典型新城疫诊断困难这一难题,方法简单准确,快速易行,值得推广和应用。     

小反刍兽疫疫苗毒株与强毒株的鉴别性荧光RT-PCR检测方法的建立

对GenBank已公布的小反刍兽疫病毒N基因序列进行分析,设计引物与探针,建立PPRV通用型与Ⅱ系疫苗毒特异型二重实时荧光RT-PCR方法,同时建立针对PPRVIV系强毒株的特异型荧光PCR方法。特异性试验和灵敏度试验表明：建立的二重荧光RT-PCR方法可特异性检测PPRV病毒,其HEX信号通道（通用型）和TAMRA信号通道（Ⅱ系疫苗毒特异型）的检测灵敏度分别可达10^1 TCID50/mL和10^2 TCID50／mL,完全满足PPRV的检测要求。用二重荧光RT-PCR方法对西藏采集的14份羊血清样品进行检测,并用建立的Ⅳ系强毒株特异型荧光PCR方法对二重荧光RT-PCR的检测效果进行评估,结果显示,该方法可有效甄别PPRV强毒株和疫苗毒株,避免假阳性结果的出现。     

山羊痘病毒ORF8～ORF18缺失重组毒株的构建和鉴定

本研究旨在通过敲除山羊痘病毒(GTPV)大段基因组片段尝试构建安全性更好的基因工程减毒株和病毒表达载体.采用PCR方法,克隆了GTPV AV41株基因组ORF7～ORF8间1 242 bp的基因片段,以及ORF18～ORF19间1 355 bp的片段.接着分别以上述2片段作为左、右侧同源重组臂,构建了包含报道基因EGFP和抗性基因gpt的GTPV重组转移载体pCF-Eg.然后采用脂质体介导法,将重组转移载体pCF-Eg与GTPV AV41株共转染原代羊睾丸细胞,通过空斑纯化筛选阳性重组病毒,并鉴定重组病毒的遗传稳定性和生长特性.结果成功获得1株敲除掉ORF8～ORF18间9个ORF、长达7 kb基因组片段的重组病毒vCF-Eg.该重组病毒能稳定表达绿色荧光蛋白至少10代,并且其增殖滴度与亲本毒株基本相同.表明上述区域是GTPV的复制非必需区.     

活猪携带猪瘟病毒检测方法比较

对2个猪场的母猪血清、扁桃体、全血样品和公猪精液分别应用荧光抗体染色法、RT-PCR、CS-FV抗原ELISA、CSFV糖蛋白Erns ELISA 4种方法进行CSFV对应检测和比较。结果显示,荧光抗体染色法检出率最高,但与其他3种方法存在较大差异,RT-PCR方法检出率和重复性次之;猪瘟抗原ELISA方法检出率第3,重复性第1;CSFV糖蛋白Erns检出率和重复性均最低,但可筛选是否感染猪瘟病毒野毒。在母猪3种样品中扁桃体的检出率最高,全血样品次之,血清检出率最低。4种样品中公猪精液重复性最好,其次是母猪全血样品,母猪扁桃体第3;母猪血清样品最差。     

The 16MΔvjbR as an ideal live attenuated vaccine candidate for differentiation between Brucella vaccination and infection

Brucellosis brings great economic burdens for developing countries. Live attenuated vaccines are the most efficient means for prevention and control of animal Brucellosis. However, the difficulties of differentiating of infection from vaccine immunization, which is essential for eradication programs, limit their applications. Therefore, the development of a vaccine that could differentiate infection from immunization will overcome the limitations and get extensive application. VjbR is a quorum sensing regulator involving in Brucella's intracellular survival. The vjbR∷Tn5 mutants have been proven effective against wild type strain challenge, implying its possibility of use in vaccine candidate development. To further evaluate this candidate gene, in the present study, the antigenicity of purified recombinant VjbR protein was analyzed. Antibodies to Brucella melitensis VjbR could be detected in sera from patients and animals with brucellosis but not in control ones, implying the potential use of this protein as a diagnostic antigen. Then a vjbR mutant of B. melitensis 16M was constructed by replacing the vjbR with kanamycin gene. The mutant showed reduced survival in macrophage and mice. Vaccination of BALB/c mice with 16MΔvjbR conferred significant protective immunity against B. melitensis strain 16M challenges, being equivalent to which induced by the license vaccine Rev.1. The vjbR deletion mutant elicited an anti-Brucella-specific immunoglobulin G response and induced the secretion of gamma interferon and interleukin-10. The most importance is that, the use of vjbR mutants as vaccines in association with diagnostic tests based on the VjbR antigen would allow the serological differentiation between infected and vaccinated animals. These results suggest that 16MΔvjbR is an ideal live attenuated vaccine candidate against B. melitensis and deserves further evaluation for vaccine development.     

布鲁氏菌病研究进展

布鲁氏菌病的早期诊断及鉴定是该病防控的重要环节,其诊断技术主要包括细菌学诊断技术、血清学诊断技术以及分子生物学诊断技术。布鲁氏菌病防控应坚持预防为主的策略,其免疫主要以弱毒活疫苗为主,随着分子生物学及重组DNA技术的发展,基因工程疫苗成为近年研究热点,但目前尚未有鉴别疫苗免疫和自然感染的鉴别诊断技术及相应的疫苗制品。本文对布鲁氏菌病的病原学、流行病学、诊断技术和疫苗研究进展等方面进行概述,以期为建立灵敏性高、特异性强、高效便捷且易推广的布鲁氏菌病诊断技术和研发更加安全有效的布鲁氏菌病疫苗提供基础。     

庭院生态养猪新技术

本文介绍了庭院生态养殖的基本内涵，猪舍的构造特点，建筑布局及其应注意的问题，并预测了庭院生态养猪在我国的发展前景。     

诱导猪肺泡巨噬细胞CD163受体表达的细胞因子筛选

猪肺泡巨噬细胞(porcine alveolar macrophages,PAM)中的CD163受体作为明星分子不仅参与正常的生理活动,而且作为重要的病毒受体介导猪繁殖与呼吸综合征病毒(Porcine reproductive and respiratory syndrome virus,PRRSV)入侵宿主靶细胞,因...     

海南坡鹿迁地保护现状与建议

迁地保护的海南坡鹿种群可以分为野放、半野放和圈养等3种主要类型。野放种群个体数量最多，约占迁地保护种群个体总数的55．3％；其次为半野放种群，其个体数量占35．6％；圈养种群个体数量最少，只占9．1％。半野放是一种较为稳妥的坡鹿种群恢复方式。在海南坡鹿的迁地保护中，应该增加半野放种群个体数量。本文就海南坡鹿的迁地保护提出了7点保护管理及科学研究建议。     

适应无血清培养的HEK293细胞系的培育

为培育适合腺病毒增殖的无血清培养细胞系,本研究采用培养液渐进替换的方法获得了一株适应无血清培养的HEK293细胞(293SF),经检测293SF细胞具有稳定的支持腺病毒增殖与表达外源蛋白的能力;平均病变时间为97 h,比HEK293细胞缩短12.9%;毒价达到107.48TCID50/mL,比HEK293细胞提高43.3%;并且稳定性良好,连续传16代后293SF细胞状态与易感性未发生变化。该细胞系为腺病毒的无血清培养奠定基础。