免疫调节剂对小鼠实验性单纯疱疹性角膜炎发病的影响

实验组小鼠腹腔分别注射免疫调节剂胸腺五肽(TP5)或环孢霉素(CsA),对照组注射生理盐水(NS),尔后角膜感染单纯疱疹病毒(HSV),造成小鼠实验性单纯疱疹性角膜炎模型。用裂隙灯显微镜观察小鼠角膜上皮、角膜实质、角膜新生血管、结膜和眼睑的病变变化情况。结果:种毒唇4～6天,TP5组角膜上皮和角膜实质病变比NS组严重,差异有显著性,而CsA无此作用。三组小鼠新生血管形成程度差异无显著性。且均在第8天出现高峰。TP5组和CsA组的结膜和眼睑病变,比NS组严重。因此,在临床上,应根据不同病种和不同情况,慎重使用免疫调节剂。     

EBVTK激酶基因在原核细胞中的表达及其在NPC诊断中的意义

目的 探索以Epstein-Barr病毒（EBV）TK激酶为抗原，建立简便、快速、敏感和特异的鼻咽癌（NPC）诊断方法。方法 构建原核表达载体pRSET-TK，在原核细胞BL21（DF3）中获得高效表达的TK激酶，以Western blot检测证明其抗原的特异性和应用于NPC检测的可行性。以纯化的TK激酶为抗原初步建立ELISA检测方法，检测NPC患者血清中TK／IgG抗体。结果 在NPC患者血清中含有特异的针对TK激酶的IgG抗体，Western blot TK/IgG检测的敏感度和特异度均为100％。结论 初步建立了ELISA TK／IgG诊断NPC方法，具有较高的敏感性和特异性。     

Inactivation of viruses infecting ectothermic animals by amphibian and piscine antimicrobial peptides

The ability of five purified amphibian antimicrobial peptides (dermaseptin-1, temporin A, magainin I, and II, PGLa), crude peptide fractions isolated from the skin of Rana pipiens and R. catesbeiana, and four antimicrobial peptides (AMPs) from hybrid striped bass (piscidin-1N, -1H, -2, and -3) were examined for their ability to reduce the infectivity of channel catfish virus (CCV) and frog virus 3 (FV3). All compounds, with the exception of magainin I, markedly reduced the infectivity of CCV. In contrast to CCV, FV3 was 2- to 4-fold less sensitive to these agents. Similar to an earlier study employing two other amphibian peptides, the agents used here acted rapidly and over a wide, physiologically relevant, temperature range to reduce virus infectivity. These results extend our previous findings and strongly suggest that various amphibian and piscine AMPs may play important roles in protecting fish and amphibians from pathogenic viruses.     

DNA polymerase gene locus of <Emphasis Type="Italic">Cercopithecine herpesvirus 1</Emphasis> is a suitable target for specific and rapid identification of viral infection by PCR technology

The family Herpesviridae comprises at least 100 herpesviruses. Numerous human and animal pathogenic herpesviruses have been identified so far, including Cercopithecine herpesvirus 1 (CeHV-1). This virus is a member of the subfamily Alphaherpesvirinae and is the most hazardous herpesvirus to man. CeHV-1 is also known as B-virus or monkey B virus and as Herpesvirus simiae. In order to gain more genetic information, the viral DNA polymerase (DPOL) gene was identified using polymerase chain reaction (PCR) and DNA nucleotide sequence analysis. The deduced amino acid sequence contains the motifs and signatures that are typical for the B-family of DPOLs. The DPOL gene of CeHV-1 was found to be a suitable target for the specific and rapid identification of the Cercopithecine herpesvirus 1 infection by PCR technology. Comparative analysis of the DNA sequences of the DPOL gene loci of CeHV-1, Human herpesvirus 1 and 2 (HHV-1 and HHV-2), and other herpesviruses was carried out for determination of unique genomic regions of the individual DPOL genes. A primer set of 12 primers was used for screening the DNA of CeHV-1, HHV-1, and HHV-2 by detailed PCR. It was found that six out of twelve primer combinations are able to detect specifically the CeHV-1 genome without cross reactivity with the genome of HHV-1 and/or HHV-2. The specificity of the individual amplified DNA fragments was confirmed by DNA nucleotide sequence analysis. The results of these studies indicate that the six primer combinations of the specific CeHV-1 DPOL primer set is the method of choice for a rapid, precise and specific identification of a CeHV-1 infection by PCR. Due to the fact that this specific CeHV-1 DPOL primer set does not amplify any DNAs of HHV-1 or HHV-2 genome this technology is stressing and can be successfully used unlimited and more credible in all laboratories with PCR technical facility routinely for detection of a CeHV-1 infection in vivo or in vitro.The GenBank Accession No. of the sequence of DNA polymerase gene of Cercopithecine herpesvirus 1 (CeHV-1) reported in this study is AY568415, DPOL protein ID AAT67222; nuclear phosphoprotein ID AAT67223     

Epstein-Barr病毒与肿瘤

Epstein-Barr病毒(EBV)属于疱疹病毒科,在很多肿瘤中都能够检测到,如伯基特淋巴瘤、鼻咽癌、胃癌和淋巴瘤等.EBV致病谱广,在这些肿瘤中扮演的角色不同,其发病机制也不同,研究EBV感染在肿瘤发生发展中的作用有助于揭示疾病发生发展的机制.     

水痘疫苗生产工艺变更前后Oka株病毒基因序列及疫苗质量分析

目的 通过对采用细胞工厂工艺进行细胞培养前后的Oka株病毒基因序列和水痘疫苗质量进行分析比较,评价生产工艺变更对水痘疫苗质量的影响.方法 从水痘疫苗原液提取病毒DNA,采用PCR法扩增片段,通过对PCR产物测序或克隆质粒测序并拼接,获得Oka株的病毒全基因序列,并与GenBank中相关序列比对.对工艺变更前后生产的疫苗进行病毒滴度、牛血清白蛋白残留量和抗生素残留量比较.结果 工艺变更前后的疫苗病毒基因序列一致;检测到33个单核苷酸多态性位点为亲本型和减毒型碱基共存,具有典型的Oka株特征;没有发现GenBank未登录或文献未报道的新突变.工艺变更前后生产疫苗平均病毒滴度分别为5.00和5.02 lg蚀斑形成单位/ml,两者间的差异无统计学意义(t=1.1,P=0.26),且各项关键指标均符合企业标准和药典要求.结论 将细胞工厂引入生产工艺后,水痘疫苗具有与原工艺生产的制品相似的安全性和质量一致性.     

杭州市一株单纯疱疹2型病毒株的分离鉴定

目的 建立一种能够对样本中的单纯疱疹病毒(Hsv)进行准确鉴定分型的方法,并从生殖器疱疹(GH)患者皮损的水疱液中分离出1株HSV-2.方法 从1例GH患者病变部位采集样本,样本接种Vero-E6细胞,待出现细胞病变后收获病毒感染物,提取病毒感染物的基因组.根据特异性PCR对分离出的病毒进行分型鉴定.结果 分离的毒株引起Vero-E6典型细胞病变,病变细胞圆缩、融合.型特异性PCR反应及l%琼脂糖凝胶电泳鉴定结果表明,扩增出的片段大小约为183 bp,与预期的HSV-2型特异性片段长度相符,且与数据库的比对结果显示相似性为100%.结论 通过细胞分离培养和特异性PCR分型鉴定,能够实现对样本中的HSV的准确分型鉴定.     

EB病毒感染对人胃上皮细胞GES-1增殖的影响

目的:探讨EB病毒感染人胃上皮细胞GES-1后对GES-1增殖的影响。方法:用携带新霉素抗性(NEOr)基因的重组EBV以密切接触法感染人胃上皮细胞系GES-1,同时以正常人胃上皮细胞GES-1为对照。经G418筛选出EBV感染阳性克隆,进行细胞形态学检测,并采用免疫组织化学染色法测定EBNA1的表达以鉴定EBV感染细胞克隆。对上述细胞用细胞计数法及MTT法检测细胞增殖情况;流式细胞术对细胞周期进行分析。结果:感染EBV的GES-1细胞大部分为EBNA1表达阳性,通过G418筛选,获得了稳定的EBV感染细胞克隆。EBV感染组细胞体积变大,核浆比增加。细胞计数法及MTT法绘制细胞生长曲线显示细胞增殖速度明显加快。流式细胞分析结果显示EBV感染组细胞S期延长。结论:EBV感染后使GES-1细胞稳定且高表达EBNA1,并且能够促进GES-1细胞的增殖。     

用显微注射法制备携带Epstein-Barr病毒膜抗原基因--BLLF1的转基因小鼠

目的 :用显微注射法制备携带EB病毒 (Epstein Barrvirus,EBV)膜抗原 (membraneantigen ,MA)基因 BLLF1的转基因小鼠。方法 :对 38只昆明种小鼠进行超排卵 ,收集受精卵 ,将EBVMA基因 BLLF1显微注射到受精卵的原核内。将注射后成活且健康的受精卵移植到假孕母鼠的输卵管内使其发育直至分娩。PCR分析检测仔鼠基因组中转基因的整合。结果 :显微注射 6 77个受精卵 ,其中成活且健康的 32 0个 ,卵的成活率为 47.2 %。将其移植到 12只假孕母鼠的输卵管内 ,其中 2只鼠怀孕并产仔 12只 ,出生率为 3 .75 %。PCR分析 5只仔鼠基因组整合有BLLF1基因 ,整合率为 41.7%。结论 :携带EBVMA基因 BLLF1的转基因小鼠已制备成功     

EB病毒VCA-IgM抗体阳性的Kikuchi-Fujimoto病

目的：分析并探讨KDD的发病机制、临床特点、诊断及治疗方法。方法：对5例于1992～1996年收治的EBV－VCA-IgM抗体阳性的KFD患者进行回顾性研究。结果：5例病人均通过肿大淋巴结活检确诊。临床都表现为颈淋巴结肿大、发热、白细胞减少、血沉增快及LDH增高。对强的松治疗较敏感。结论：尚不能确定EBV即为KFD的发病原因。淋巴结活检是唯一确诊手段。该病报告日渐增多，应引起临床医师足够重视。