关键词： heterogeneous-nuclear ribonucleoproteins   RNA   Polioviruses   Proteins   Complement 1  

摘要： We had previously demonstrated that a cellular protein specifically interacts with the 3' end of poliovirus negative-strand RNA. We now report the identity of this protein as heterogeneous nuclear ribonucleoprotein (hnRNP) C1/C2. Formation of an RNP complex with poliovirus RNA was severely impaired by substitution of a lysine, highly conserved among vertebrates, with glutamine in the RNA recognition motif (RRM) of recombinant hnRNP C1, suggesting that the binding is mediated by the RRM in the protein. We have also shown that in a glutathione S-transferase (GST) pull-down assay, GST/hnRNP C1 binds to poliovirus polypeptide 3CD, a precursor to the viral RNA-dependent RNA polymerase, 3D(pol), as well as to P2 and P3, precursors to the nonstructural proteins. Truncation of the auxiliary domain in hnRNP C1 (C1DeltaC) diminished these protein-protein interactions. When GST/hnRNP C1DeltaC was added to in vitro replication reactions, a significant reduction in RNA synthesis was observed in contrast to reactions supplemented with wild-type fusion protein. Indirect functional depletion of hnRNP C from in vitro replication reactions, using poliovirus negative-strand cloverleaf RNA, led to a decrease in RNA synthesis. The addition of GST/hnRNP C1 to the reactions rescued RNA synthesis to near mock-depleted levels. Furthermore, we demonstrated that poliovirus positive-strand and negative-strand RNA present in cytoplasmic extracts prepared from infected HeLa cells coimmunoprecipitated with hnRNP C1/C2. Our findings suggest that hnRNP C1 has a role in positive-strand RNA synthesis in poliovirus-infected cells, possibly at the level of initiation.

关键词： Animals   Antibodies, Viral   Antigens/chemistry   Antigens, Viral/chemistry   Biotin/chemistry   Biotinylation   Calibration   Cesium/pharmacology   Chlorides/pharmacology   Dose-Response Relationship, Drug   Enzyme-Linked Immunosorbent Assay/methods   Enzyme-Linked Immunosorbent Assay/methods   Immunoglobulin G/chemistry   Linear Models   Models, Theoretical   Peroxidase/metabolism   Poliovirus/metabolism   Poliovirus Vaccine, Inactivated/analysis   Poliovirus Vaccine, Oral/analysis   Rabbits   Temperature  

摘要： An improved ELISA test for determination of potency of Inactivated Poliovirus Vaccine (IPV) is proposed. The method is based on the use of IgG purified from immune rabbit serum conjugated with biotin. Optimized and validated materials for the test can be stored for a long time in the form of ready-to-use kits. Optimization included selection of anti-poliovirus rabbit antibody batches with the best specificity to D-antigen as well as finding the most efficient parameters for all steps of ELISA protocol. The assay is based on direct ("sandwich") ELISA scheme, in which antigens are captured on ELISA plates coated with purified rabbit polyclonal D-antigen specific IgG raised against wild polioviruses of three serotypes. D-antigen specificity of the IgG was at least 10 times higher than to H-antigen (heat-inactivated virus). The presence of antigen was detected using biotin-conjugated IgG from the same source. Eight-point dose-response curves were obtained for each sample and the reference vaccine. The protocol ensured low background (less than 0.2 OD), linear response over the entire range of optical density measurements (up to 3.0 OD), and high precision of data (assay variability was about 3%). The quantitative results and the validity of the test were determined by two numerical approaches, linear regression and a new analysis procedure called the local interpolation method. For the first approach we also proposed a new method for testing of parallelism of regression lines. The ELISA protocol for all three types of poliovirus is based on standard off-the-shelf reagents, and is highly reproducible and reliable. An in-house Reference Reagent was formulated and calibrated against the International Reference for IPV. Published by Elsevier Ltd on behalf of The International Association for Biologicals.

关键词： AOD use initiation   Polioviruses   replication   RNA   Elements  

摘要： The 5' cloverleaf in poliovirus RNA has a direct role in regulating the stability, translation, and replication of viral RNA. In this study, we investigated the role of stem a in the 5' cloverleaf in regulating the stability And replication of poliovirus RNA in HeLa S10 translation-replication reactions. Our results showed that disrupting the duplex structure of stem a destabilized viral RNA and inhibited efficient negative-strand synthesis. Surprisingly, the duplex structure of stem a was not required for positive-strand synthesis. In contrast, altering the primary sequence at the 5'-terminal end of stem a had little or no effect on negative-strand synthesis but dramatically reduced positive-strand initiation and the formation of infectious virus. The inhibition of positive-strand synthesis observed in these reactions was most likely a consequence of nucleotide alterations in the conserved sequence at the 3' ends of negative-strand RNA templates. Previous studies suggested that VPgpUpU synthesized on the cre(2C) hairpin was required for positive-strand synthesis. Therefore, these results are consistent with a model in which preformed VPgpUpU serves as the primer for positive-strand initiation on the 3'AAUUUUGUC5' sequence at the 3' ends of negative-strand templates. Our results suggest that this sequence is the primary cis-acting element that is required for efficient VPgpUpU-primed positive-strand initiation.

关键词： Child   Guinea-Bissau/epidemiology   Hospital Mortality   Humans   Poliomyelitis/mortality   Poliomyelitis/mortality   Poliomyelitis/prevention & control   Poliomyelitis/prevention & control   Poliovirus Vaccine, Oral/therapeutic use  

摘要： Children with multiple clinic visits have more opportunities to be vaccinated that children attending a single occasion. [...]DTP vaccination during a time of intermittent vaccine shortages in Bissau may simply be a marker for the failing health of a child who subsequently died. While the authors discount the importance of socio-economic status, a mother's work in agricultural labor, the location of residence, having money for transport and the availability of food, clean water, bed nets and other health factors would have influenced when and if a child was brought to the clinic.

关键词： oral poliovirus vaccine   CHAT   HIV/SIV   contamination   mitochondrial DNA   numts  

摘要： Inoculation of live experimental oral poliovirus vaccines (OPV CHAT) during the 1950s in central Africa has been proposed to account for the introduction of HIV into human populations. For this to have occurred, it would have been necessary for chimpanzee rather than macaque kidney epithelial cells to have been included in the preparation of early OPV materials. Theoretically, this could have led to contamination with a progenitor of HIV-1 derived front a related simian immunodeficiency virus of chimpanzees (SIVCPZ). In this article we present further detailed analyses of two samples of OPV, CHAT 10A-11 and CHAT 6039/Yugo, which were used in early human trials of poliovirus vaccination. Recovery of poliovirus by culture techniques confirmed the biological viability of the vaccines and sequence analysis of poliovirus RNA specifically identified the presence of the CHAT strain. Independent nested sets of oligonucleotide primers specific for HIV-1/SIVCPZ and HIV-2/SIVMAC/SIVSM phylogenetic lineages, respectively, indicated no evidence of HIV/SIV RNA in either vaccine preparation, at a sensitivity of 100 RNA equivalents/ml. Analysis of cellular substrate by the amplification of two distinct regions of mitochondrial DNA (D-loop control region and 12S ribosomal sequences) revealed no evidence of chimpanzee cellular sequences. However, this approach positively identified rhesus and cynomolgus macaque DNA for the CHAT 10A-11 and CHAT 6039/Yugo vaccine preparations, respectively. Analysis of multiple clones of mtDNA 12S rDNA indicated a relatively high number of nuclear mitochondrial DNA sequences (numts) in the CHAT 10A-11 material, but confirmed the macaque origin of cellular substrate used in vaccine preparation. These data re-inforce earlier findings on this topic providing no evidence to support the contention that poliovirus vaccination was responsible for the introduction of HIV into humans and sparking the AIDS pandemic. (C) 2004 Elsevier Ltd. All rights reserved.

关键词： 肠道病毒   无症状   脊髓灰质炎病毒   柯萨奇病毒   临床   PV   血液   类病毒   细胞繁殖   器官  

摘要： 人肠道病毒(HEV)隶属于微小核糖核酸病毒科，主要包括脊髓灰质炎病毒(PV)、柯萨奇病毒(Cox)A群和B群、艾柯病毒(Echo)，以后又发现了5个新血清型并直接命名为新肠道病毒68—72型。这类病毒通常以无症状状态在人类消化道细胞繁殖，然后通过血液侵犯其他器官，引起各种临床综合症状。

关键词： 脊髓灰质炎疫苗   牛痘   预防和控制   儿童计划免疫   研究进展   天花   百日咳   问题   经济   现状  

摘要： 自古以来,免疫预防就是预防和控制传染病最经济、最简便、最有效的手段.而疫苗是免疫预防工作的最重要的工具.18世纪英国医生琴纳发明种牛痘预防天花,由于这种方法更为安全可靠而得到广泛应用.目前接种疫苗每年可以使全世界数百万人免于死亡,我国已经纳入儿童计划免疫的疫苗有卡介苗、脊髓灰质炎疫苗、百日咳白喉-破伤风联合疫苗、麻疹疫苗、乙肝疫苗5种,可以预防7种疾病.10多年计划免疫的开展使传染病得到了有效控制.继20世纪80年代成功消灭天花后,2000年10月29日中国的无脊髓灰质炎状态也得到证实.

关键词： 脊髓灰质炎病毒   试验方法   细胞病理变化   MTT法   活细胞染色法   中和产物   次氯酸钠   CPE法   灭活指数   细胞感染法   常用消毒剂   有效氯含量   悬液定量   过氧乙酸   观察结果   测定结果   正式试验   中和剂   存活性   毒试验   酸含量  

摘要： 目的:研究消毒剂灭活脊髓灰质炎病毒的试验方法。方法:试验采用细胞感染法,以活细胞染色法(MTT法)和观察细胞病理变化(CPE)法测定中和剂、消毒剂及中和产物对细胞存活性的影响。选择2种常用消毒剂:次氯酸钠和过氧乙酸分别进行悬液定量灭活病毒试验。结果:MTT法测定结果与CPE法观察结果基本一致;过氧乙酸含量为500mg/L作用10min, 对脊髓灰质炎病毒的灭活指数大于4,次氯酸钠有效氯含量300mg/L作用10min对脊髓灰质炎病毒的灭活指数大于4。结论:在预备实验中,MTT法测定中和剂、消毒剂及中和产物对细胞存活性的影响准确可靠;正式试验时,以CPE法观察细胞病理变化更简便易行。

关键词： 有效氯   次氯酸钠   脊髓灰质炎病毒   灭活效果  

摘要： 观察次氯酸钠消毒剂对脊髓灰质炎病毒灭活效果,采用悬液法进行检测,结果在20～22℃,相对湿度为54%～65%的条件下,次氯酸钠消毒剂有效氯含量为250.0mg/L,加入病毒悬液作用1min后,对脊髓灰质炎病毒平均灭活对数值为2.89;加入病毒悬液作用5min、10min后对脊髓灰质炎病毒平均灭活对数值≥4.00。

关键词： 下肢   脊髓灰质炎后综合征   矫形装置  

摘要： 目的:探求矫形器在康复工程上的治疗作用,分析下肢矫形器(髋膝踝足矫形器膝踝足矫形器,踝足矫形器)在脊髓灰质炎后遗症(简称儿麻)矫治术后康复上的应用。方法:2002-10/2004-06,在福建省石狮市,通过对19例术后儿麻患者佩带矫形器的治疗,分析下肢矫形器的治疗与矫治效果。结果:19例术后儿麻患者均达到了控制髋屈曲、膝反张、膝屈曲、马蹄足、高弓足,纠正畸形步态等,保持正确站立姿势及改善步态的效果,显效率为79%、有效率为21%、总显效率为80%。结论:下肢矫形器在巩固儿麻患者术后的下肢稳定功能、运动功能方面起到积极的作用。