犬细小病毒单克隆抗体的制备及其与免疫血清在血凝抑制中的比较

将用犬细小病毒免疫的ＢＡＬＢ／ｃ小鼠的脾细胞与Ｓｐ２／０骨髓瘤细胞在聚乙二醇作用下融合,经筛选、克隆,得到６株稳定分泌犬细小病毒单克隆抗体的杂交瘤。用杂交瘤腹水作血凝抑制试验,检测分别含有犬细小病毒、猫泛白细胞减少症病毒和水貂肠炎病毒的标本,及ＣＰＶ感染犬粪便标本,并与免疫血清作对比。结果表明,用单克隆抗体作血凝抑制试验,比用免疫血清作具有特异性高、操作简便省时等优点,而二者敏感性一致。     

真核藻类的病毒和病毒类粒子（VLPs）

真核藻类的病毒和病毒类粒子（ＶＬＰｓ）赵以军石正丽（中国科学院水生生物研究所,武汉４３００７２）（中国科学院武汉病毒研究所,武汉４３００７１）ＶｉｒｕｓｅｓａｎｄＶｉｒｕｓ－ｌｉｋｅＰｅａｒｔｉｃｌｅｓｏｆＥｕｋａｒｙｏｔｉｃＡｌｇａｅＺｈａｏＹｉｊ...     

中华鳖病毒的血清学检测

中华鳖病毒（ＴＳＶ）是从病鳖中分离到的一种病毒病原。经细胞培养和差异离心制备ＴＳＶ抗原,肌注家兔获ＴＳＶ抗体（ＴＳＶ－Ａｂ）,中和效价为１：２０,用ＴＳＶ－Ａｂ进行双向免疫扩散和间接ＥＬＩＳＡ检测,被检样品有健康和病鳖组织匀浆液、ＴＳＶ细胞培养液、提纯的ＴＳＶ,以及鱼传染性胰脏坏死病毒（ＩＰＮＶ）、草鱼呼肠孤病毒（ＧＣＶ）、鱼病毒性出血败血症病毒（ＶＨＳＶ）、鲁痘疮病毒（Ｃａｒｐ ｐｏｘ ｖｉｒｕ     

传染性软疣病毒（MCV）的电镜观察

传染性软疣病毒（ＭＣＶ）的电镜观察李德忠,肖同浩,武晓华（广州军区武汉总医院电镜室,武汉４３００７０）吴宁（广州军区武汉总医院皮肤科,武汉４３００７０）关键词传染性软疣病毒,病毒形态,电镜观察曾有研究描述ＭＣＶ的发育周期中有８种形态,也有按发育过程将...     

HIV感染的辅助因子—融合素

ＨＩＶ感染的辅助因子——融合素黄仕和秦椿华（卫生部武汉生物制品研究所,武汉４３００６０）（同济医科大学工业毒理研究室,武汉４３００３０）关键词融合素ＨＩＶ感染１９８４年,研究者发现,ＨＩＶ感染细胞是通过细胞表面受体ＣＤ４进行的。二年后又发现,仅有ＣＤ...     

沼泽绿牛蛙病毒的分离及其细胞感染的初步研究

沼泽绿牛蛙病毒的分离及其细胞感染的初步研究张奇亚,李正秋,江育林,梁绍昌,桂建芳（中国科学院水生生物研究所,武汉４３００７２）关键词沼泽绿牛蛙（美国青蛙）,病毒,细胞感染ＰＲＥＬＩＭＩＮＡＲＹＳＴＵＤＩＥＳＯＮＶＩＲＵＳＩＳＯＬＡＴＩＯＮＡＮＤＣＥＬ...     

犬瘟热病毒细胞膜受体的鉴定

犬瘟热病毒（ＣＤＶ）敏感细胞Ｖｅｒｏ用ＳＤＳ或ＲＩＰＡ溶解缓冲液溶解,利用病毒铺覆蛋白印迹技术（ＶＯＰＢＡ）鉴定犬瘟热病毒疫苗株（ＣＤＶ－ｏｎｄｅｓｔｅｐｏｏｒｔ）的细胞受体。结果发现,在Ｖｅｒｏ细胞上有两组ＣＤＶ结合蛋白质,即高分子量组蛋白质（１２７ｋＤ、１２０ｋＤ、１１０ｋＤ）与低分子量组蛋白质（２７ｋＤ和３０ｋＤ）。这些ＣＤＶ结合蛋白组分的性质及在ＣＤＶ致病中的作用有等进一步研究。     

炭凝集试验快速检测呼吸道合胞病毒的研究

用炭凝集试验（ＣＡＴ）检测呼吸道合胞病毒（ＲＳＶ）,结果表明该法是一种简便、快速、特异的诊断方法。用ＣＡＴ对１６株ＲＳＶ和８株其它病毒做试验,结果仅ＲＳＶ凝集,而其它病毒均阴性。用该法与细胞培养法检测８３份临床呼吸道感染幼儿鼻咽吸出物,结果ＣＡＴ法阳性率为６９．８８％（５３／８３）,细胞培养法为３９．７５％（３３／８３）,两者阳性检出率相差极显著。阻断试验证明ＣＡＴ是高度特异的。结果证明ＣＡＴ具有较高的敏感性与特异性,可用于临床ＲＳＶ标本的快速检测。     

真菌传杆状病毒组（Furovirus）内四种病毒的细胞病理学比较研究

本文研究丁真菌传杆状病毒组（Ｆｕｒｏｖｉｒｕｓ）四个病毒成员,即甜菜坏死黄脉病毒（ＢＮＹＶＶ）、甜菜土传病毒（ＢＳＢＶ）、花生丛生病毒印度株系（ＰＣＡ－Ｉ）以及非洲株系（ＰＣＡ－Ａ）和小麦土传花叶病毒（ＳＢＷＭＶ）所引起的细胞病理变化。这四种病毒在病毒粒子聚集的结构,是否引起细胞膜聚集及聚集的结构,过氧化物酶体泡状物的形成与否及结构均有差异。另外,同一种病毒的不同株系或分离物所引起的细胞病理变化也不相同。     

麻疹病毒血凝素基因H的点突变与血凝作用的转变

经Ｂ９５ａ细胞系纯化分离的三株麻疹病毒Ｆｕ、ＩＭＡ、ＳＭＤ的血凝试验（ＨＡＤ）为阴性,但将它们分别感染Ｖｅｒｏ细胞并传代培养后其血凝试验由阴性转变为阳性。实验证明这种血凝表型的转变与血凝素基因Ｈ的表达与吸附无关。以ＰＣＲ扩增血凝素基因Ｈ和融合基因Ｆ,并分别进行序列分析。表明三株病毒在两种培养细胞中传代后其融合基因Ｆ的序列未发现差异。但血凝素基因Ｈ的序列却有显著的点突变并伴随血凝表型的转变。其中Ｆｕ     

Antiviral Activity of Antimicrobial Lipopeptide from Bacillus subtilis fmbj Against Pseudorabies Virus, Porcine Parvovirus, Newcastle Disease Virus and Infectious Bursal Disease Virus in Vitro

Bacillus subtilis fmbj can produce lipopeptide antimicrobial substance, whose main components were surfactin and fengycin. In the study, the antiviral activity of antimicrobial lipopeptides (AMLs) from B. subtilis fmbj (CGMCC No. 0934) against Pseudorabies Virus (PRV), Porcine Parvovirus (PPV), Newcastle Disease Virus (NDV) and Infectious Bursal Disease Virus (IBDV) was evaluated in vitro. The AMLs represented a direct inactivation effect on cell-free virus stocks of PRV, PPV, NDV and IBDV, and it could effectively inhibit infection and replication of the NDV and IBDV, but failed to affect PRV and PPV. The AMLs were represented higher toxicity for the Porcine Kidney (PK-15) cells (50% cytotoxic concentration (CC₅₀) value was 32.87 μM) and lower for the Chicken Embryo Fibroblasts (CEF) cells (CC₅₀ value was 89.16 μM). The Selectivity index of AMLs on PRV, PPV, NDV and IBDV was 1.44, 2.23, 8.40 and 12.19, respectively.     

Inactivation of enveloped and non-enveloped viruses on seeded human tissues by gamma irradiation

Human tissue allografts are widely used in a variety of clinical applications with over 1.5 million implants annually in the US alone. Since the 1990s, most clinically available allografts have been disinfected to minimize risk of disease transmission. Additional safety assurance can be provided by terminal sterilization using low dose gamma irradiation. The impact of such irradiation processing at low temperatures on viruses was the subject of this study. In particular, both human tendon and cortical bone samples were seeded with a designed array of viruses and the ability of gamma irradiation to inactivate those viruses was tested. The irradiation exposures for the samples packed in dry ice were 11.6-12.9 kGy for tendon and 11.6-12.3 kGy for bone, respectively. The viruses, virus types, and log reductions on seeded tendon and bone tissue, respectively, were as follows: Human Immunodeficiency Virus (RNA, enveloped), >2.90 and >3.20; Porcine Parvovirus (DNA, non-enveloped), 1.90 and 1.58; Pseudorabies Virus (DNA, enveloped), 3.80 and 3.79; Bovine Viral Diarrhea Virus (RNA, enveloped), 2.57 and 4.56; and Hepatitis A Virus (RNA, non-enveloped), 2.54 and 2.49, respectively. While proper donor screening, aseptic technique, and current disinfection practices all help reduce the risk of viral transmission from human allograft tissues, data presented here indicate that terminal sterilization using a low temperature, low dose gamma irradiation process inactivates both enveloped and non-enveloped viruses containing either DNA or RNA, thus providing additional assurance of safety from viral transmission.     

Transfusion-transmitted infectious diseases

A spectrum of blood-borne infectious agents is transmitted through transfusion of infected blood donated by apparently healthy and asymptomatic blood donors. The diversity of infectious agents includes hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency viruses (HIV-1/2), human T-cell lymphotropic viruses (HTLV-I/II), Cytomegalovirus (CMV), Parvovirus B19, West Nile Virus (WNV), Dengue virus, trypanosomiasis, malaria, and variant CJD. Several strategies are implemented to reduce the risk of transmitting these infectious agents by donor exclusion for clinical history of risk factors, screening for the serological markers of infections, and nucleic acid testing (NAT) by viral gene amplification for direct and sensitive detection of the known infectious agents. Consequently, transfusions are safer now than ever before and we have learnt how to mitigate risks of emerging infectious diseases such as West Nile, Chikungunya, and Dengue viruses.     

Comprehensive mapping of common immunodominant epitopes in the West Nile virus nonstructural protein 1 recognized by avian antibody responses

West Nile virus (WNV) is a mosquito-borne flavivirus that primarily infects birds but occasionally infects humans and horses. Certain species of birds, including crows, house sparrows, geese, blue jays and ravens, are considered highly susceptible hosts to WNV. The nonstructural protein 1 (NS1) of WNV can elicit protective immune responses, including NS1-reactive antibodies, during infection of animals. The antigenicity of NS1 suggests that NS1-reactive antibodies could provide a basis for serological diagnostic reagents. To further define serological reagents for diagnostic use, the antigenic sites in NS1 that are targeted by host immune responses need to be identified and the potential diagnostic value of individual antigenic sites also needs to be defined. The present study describes comprehensive mapping of common immunodominant linear B-cell epitopes in the WNV NS1 using avian WNV NS1 antisera. We screened antisera from chickens, ducks and geese immunized with purified NS1 for reactivity against 35 partially overlapping peptides covering the entire WNV NS1. This study identified twelve, nine and six peptide epitopes recognized by chicken, duck and goose antibody responses, respectively. Three epitopes (NS1-3, 14 and 24) were recognized by antibodies elicited by immunization in all three avian species tested. We also found that NS1-3 and 24 were WNV-specific epitopes, whereas the NS1-14 epitope was conserved among the Japanese encephalitis virus (JEV) serocomplex viruses based on the reactivity of avian WNV NS1 antisera against polypeptides derived from the NS1 sequences of viruses of the JEV serocomplex. Further analysis showed that the three common polypeptide epitopes were not recognized by antibodies in Avian Influenza Virus (AIV), Newcastle Disease Virus (NDV), Duck Plague Virus (DPV) and Goose Parvovirus (GPV) antisera. The knowledge and reagents generated in this study have potential applications in differential diagnostic approaches and subunit vaccines development for WNV and other viruses of the JEV serocomplex.     

犬细小病毒中国内蒙株VP2基因克隆及序列分析

从我国内蒙古地区流行的犬细小病毒病病犬的肠溶物中分离提纯犬细小病毒（ＣＰＶ）。提取病毒基因组ＤＮＡ,并以此ＤＮＡ为模板,采用人工合成的引物进行ＰＣＲ扩增,ＰＣＲ产物经ＢａｍＨＩ、ＳａｃＩ双酶切后,克隆于ｐＵＣ１９质粒的ＢａｍＨＩ／ＳａｃＩ位点。重组质粒ｐＵＣＶＰ２经ＰＣＲ鉴定、限制酶切分析和序列分析,结果表明：获得了犬细小病毒内蒙株（ＣＰＶ－ＩＭ）ＶＰ２基因的全长克隆,ＶＰ２基因全长１７５５ｎｔ,     

运用ELISA快速检测CPV抗体方法的建立与均衡性研究

用蔗糖密度梯度离心和凝胶层析纯化犬细小病毒 (CPV)作抗原 ,建立了检测CPV抗体的间接酶联免疫吸附试验 (ELISA)法 ,其特异性和稳定性良好 ,结果判定准确明显 ,易于把握。     

Assessment of the specificity of a commercial human parvovirus B19 IgM assay

Background: It is important to investigate a possible cross-reaction of anti-rubella IgM in the IDEIA^™ Parvovirus B19 IgM test because many B19 infections are either asymptomatic or have clinical symptoms similar to those of rubella virus infections. Epstein-Barr virus (EBV) IgM, cytomegalovirus (CMV) IgM, measles IgM and rheumatoid factor (RF) IgM cross-reactions were also studied.Objectives: In the period from February to September 1994 (including a parvovirus B19 epidemic) more than 10 000 serum samples were examined for parvovirus B19 IgM in Denmark. This gave an opportunity to evaluate the commercial IDEIA^™ Parvovirus B19 ELISA kit (DAKO A/S, Glostrup, Denmark), which was used routinely at Statens Serum Institut from the beginning of 1994 and onwards.Study design: A total of 123 parvovirus B19 IgM positive sera were tested for reactivity in rubella IgM EIA. A total of 78 rubella IgM positive sera, 60 EBV VCA-IgM positive sera, 30 CMV IgM positive sera and 24 measles virus IgM positive sera were tested for reaction in IDEIA^™ Parvovirus B19 IgM test. Finally, 25 parvovirus IgM positive sera were tested for specific IgM against measles virus, EBV (VCA), CMV and for RF.Results: One anti-B19 IgM positive serum sample reacted positively in the rubella IgM test. Of rubella IgM positive serum samples 4% cross-reacted in IDEIA^™ Parvovirus B19 IgM test, as did 17 and 20% of EBV VCA-IgM and CMV IgM positive serum samples respectively. None of measles virus IgM positive serum samples cross-reacted in the IDEIA^™ Parvovirus B19 IgM test. Of 25 initially parvovirus B19 IgM positive sera 20% cross-reacted in EBV VCA IgM test and 8% in the CMV IgM test. None reacted positively in measles virus IgM test; 28% showed weak reactivity in RF IgM test.Conclusions: Precautions must be taken when results of IgM assays are interpreted. Epidemiological and clinical observations must be considered.     

传染性喉气管炎病毒gB基因和新城疫病毒F基因在重组禽痘病毒中共表达

在重组禽痘病毒中表达多个禽类病原的主要免疫原基因是构建多价基因工程疫苗的前提,但相关研究很少。在表达传染性喉气管炎病毒(ILTV)gB基因重组禽痘病毒的转移载体的基础上,构建了含有ILTV gB基因和新城疫病毒(NDV)F基因的重组禽痘病毒转移载体pSY-gB-F,采用脂质体转染禽痘病毒感染的鸡胚成纤维(CEF)细胞后,通过蓝斑试验筛选出重组禽痘病毒(rFPv-gB-F),并进行了6轮蚀斑纯化。Western-blot试验和间接免疫荧光试验证明ILTV gB基因和NBVF基因在rFPV-gB-F感染的CEF细胞中获得表达。为传染性喉气管炎、新城疫与鸡痘活载体多价疫苗的研制奠定基础。     

Rederivation of Transgenic and Gene-Targeted Mice by Embryo Transfer

Research on genetically engineered mice provides insights into the etiology, therapy, and genetic basis of human diseases. An important variable that affects the results of mouse studies is the health status of the animals. Pathogen burdens may confound observations and obscure underlying mechanisms. Mouse resource centers frequently rederive infected mouse strains. We review our experience on the use of a well-established technique, embryo transfer to rederive infected mouse strains. The following mouse pathogens were eliminated by embryo transfer: Mouse Parvovirus, Mouse Hepatitis Virus, Mouse Rotavirus, Mouse Encephalomyelitis Virus, Mouse Adenovirus, Helicobacter species, endoparasites, and ectoparasites. We rederived transgenic mouse lines, gene-targeted mouse lines, and lines with spontaneous mutations. In the majority of strains, fertilized eggs for embryo transfer were obtained by mating superovulated egg donors with males of the desired genotype. A total of 309 embryo transfers were performed to rederive 96 mouse strains. The pregnancy rate was 76%; 1996 pups were born, of which 43% carried the desired genotype. We performed 44 additional embryo transfers to rederive 15 other strains. The pregnancy rate was lower (45%) and none of the 135 pups carried the desired genotype. Although we successfully eliminated the pathogens in all transfers, we were unable to obtain pups with the desired genotype in 15 of 111 mouse lines. Multiple factors affect the efficiency of rederivation by embryo transfer. They include the response to superovulation by embryo donors, the number and age of stud males, the yield of fertilized eggs, the number of embryo transfers, and genotyping.     

Segregation of a single outboard left-end origin is essential for the viability of parvovirus minute virus of mice

During DNA replication, the hairpin telomeres of Minute Virus of Mice (MVM) are extended and copied to create imperfectly palindromic duplex junction sequences that bridge adjacent genomes in concatameric replicative-form DNA. These are resolved by the viral initiator protein, NS1, but mechanisms employed at the two telomeres differ. Left-end:left-end junctions are resolved asymmetrically at a single site, OriLTC, by NS1 acting in concert with a host factor, parvovirus initiation factor (PIF). Replication segregates doublet and triplet sequences, initially present as unpaired nucleotides in the bubble region of the left-end hairpin stem, to either side of the junction. These act as spacers between the NS1 and PIF binding sites, and their asymmetric distribution sets up active (OriLTC) and inactive (OriLGAA) forms of OriL. We used a reverse genetic approach to disrupt this asymmetry and found that neither opposing doublets nor triplets in the hairpin bubble were tolerated. Viable mutants were isolated at low frequency and found to contain second-site mutations that either restored the asymmetry or crippled one PIF binding site. These mutations either inactivated the inboard or activated the outboard form of OriL, a polarity that strongly suggests that, in the genus Parvovirus, an active inboard OriL is lethal.