关键词： 预防接种   脊髓灰质炎疫苗   异常反应   疫苗相关病   偶合  

摘要： 为探讨脊髓灰质炎减毒活疫苗预防接种的反应及处理原则,作者查阅了近年来脊髓灰质炎疫苗预防接种反应的文献报道,结果表明,接种脊髓灰质炎疫苗后,可发生皮肤过敏性反应、胃肠道异常反应、精神性反应、疫苗相关疾病等,还可以诱发和偶合多种疾病及婴儿猝死。异常反应的处理原则主要是抗过敏、对症治疗。诱发癔病要采取心理疏导和暗示治疗。偶合疾病,是由受种者患病或体质原因造成,与疫苗和接种技术无关,关键是要由有关部门及时做出权威的诊断,以免引起医患纠纷。

关键词： 环介导等温扩增   LAMP   检测   肠道病毒   脊髓灰质炎病毒   饮用水源水   粪便   食品包装  

摘要： 肠道病毒(Enteroviruses)隶属于小核糖核酸病毒科(Picornaviridae),是最常见的可引起全球范围内疾病流行的病毒之一,其中人类肠道病毒包括柯萨奇病毒、埃可病毒、脊髓灰质炎病毒及一些新型肠道病毒。肠道病毒感染主要通过粪-口和口-口传播。其中,接触感染者粪便污染的水、食物及土壤可以导致粪-口传播。每年全球感染肠道病毒人数可达数百万,可导致一系列临床疾病。肠道病毒的传统检测方法为病毒分离培养,然后进行血清中和试验定型分类,培养结果通常需要1周左右,操作耗时、费用高。因此,建立一种高效、灵敏、快速、简便的肠道病毒检测技术对患者的及时诊治具有重要的指导意义。 Notomi T等(2000年)发明了环介导等温扩增技术(loop-mediated isothermal amplification,LAMP),该技术具有更高的灵敏度及特异性,缩短了扩增时间,提高了扩增效率,并降低了仪器设备及检测条件的要求。 基于此,本研究利用LAMP技术建立了灵敏度高、特异性强的肠道病毒检测新方法,并成功应用于手足口病患儿粪便、饮用水源水及食品包装样品中肠道病毒的检测及饮用水源水样品中脊髓灰质炎病毒的检测。 目的: 建立肠道病毒通用LAMP检测体系及脊髓灰质炎病毒LAMP检测体系,并分别应用于实际样品中肠道病毒和脊髓灰质炎病毒的定性检测。应用实时RT-LAMP技术对天津市饮用水源水样品中的脊髓灰质炎病毒进行定量检测。 方法: 1从GenBank中收集肠道病毒基因序列,生物信息学分析得出特异性靶序列,设计出4条特异性引物。恒温条件下经链置换DNA聚合酶扩增。通过调整Mg2+浓度、甜菜碱浓度、dNTPs浓度、反应温度等优化反应体系。评价此体系的特异性及灵敏度。将已建立的肠道病毒通用LAMP方法应用于手足口病患儿粪便、饮用水源水及食品包装样品中肠道病毒的检测。LAMP扩增结果可通过肉眼观察白色沉淀或在产物中加入SYBR Green I判定,也可通过内切酶NlaIV酶切及测序确定。 2从GenBank中收集脊髓灰质炎病毒基因序列,分析其生物学信息,设计脊髓灰质炎病毒LAMP引物。优化LAMP反应体系,评价该体系的特异性及灵敏度。将已建立的脊髓灰质炎病毒LAMP方法应用于9例天津市饮用水源水样品中脊髓灰质炎病毒的检测。肉眼观察白色沉淀或在产物中加入SYBR Green I判断检测结果。LAMP产物也可通过内切酶HpyCH4III酶切鉴定。同时,随机选取2例样品扩增产物进行序列分析。 3应用实时RT-LAMP技术对18例天津市饮用水源水样品中的脊髓灰质炎病毒进行定量检测。 结果: 1本研究建立的肠道病毒通用LAMP检测方法1-1.5小时即可完成检测。该方法具有良好的特异性,可检测包括肠道病毒71型、柯萨奇A16、B3、B5以及埃可30在内的肠道病毒。其检测下限为101copies/μL,与PCR检测下限一致。样品检测结果表明,此方法对于水源水及粪便样品的检出率较高,分别为9/9(100%)、18/19(94.7%),而4份食品包装样品全部为阴性。产物经NlaIV酶消化后,主要产生103bp、87bp两种片段,与预期结果一致。随机选取的10例样品产物测序表明,2例样品与柯萨奇A16高度同源,8例样品与肠道病毒71型高度同源。 2本研究建立的脊髓灰质炎病毒LAMP检测方法1-1.5小时即可完成检测。该方法具有良好的特异性,除脊髓灰质炎病毒外,对其它类型肠道病毒及甲肝病毒均无扩增。其检测下限为101copies/μL,与传统PCR基本一致,但所需设备简单、快速,且结果观测方便。9例饮用水源水样品检测结果表明,8例样品中检测出脊髓灰质炎病毒。产物经HpyCH4III酶消化,主要产生大小为97bp的片段,与预期结果一致。2例样品产物经测序表明均与脊髓灰质炎病毒高度同源。 3超滤离心管浓缩方法中病毒的回收率为34.83%。实时RT-LAMP方法对18例水样中的脊髓灰质炎病毒定量检测表明,阳性结果中脊髓灰质炎病毒的浓度范围为7.5×105至1.1×108copies/L。 结论: 本研究成功建立了肠道病毒通用LAMP检测新方法及脊髓灰质炎病毒LAMP检测新方法,具有简便、快速、特异性好、灵敏度高的特点,为实际样品中肠道病毒和脊髓灰质炎病毒的检测提供了技术平台,在快速检测方面有很好的应用前景。

关键词： 脊髓灰质炎疫苗   小儿麻痹症   脊髓灰质炎病毒   患儿   预防感染   肢体麻痹   生活质量   接种疫苗   脊髓神经   儿童   现阶段   医学   损伤   死亡   时间   家长   恢复  

摘要： 小儿麻痹症是一种感染损伤儿童脊髓神经的病毒,医学上称之为“脊髓灰质炎”.患儿一旦感染,大多数会出现肢体麻痹,行走不便,终身无法完全恢复正常,影响患儿日后的生活质量,严重者甚至导致患儿死亡.因此,预防小儿麻痹症尤为重要.现阶段完全可以通过服用“糖丸”、接种疫苗来预防感染脊髓灰质炎病毒.每年的12月初是强化接种儿童脊髓灰质炎疫苗的时间,有小宝宝的家长别忘记啦!

关键词： 脊髓灰质炎病毒   结构蛋白   Sf9昆虫细胞   杆状病毒表达系统  

摘要： 脊髓灰质炎是由脊髓灰质炎病毒感染引起的,没有特效治疗方法,只能接种疫苗预防。口服脊髓灰质炎疫苗已大幅度降低了全球脊髓灰质炎的发病率,然而,由于疫苗相关麻痹型脊灰和疫苗衍生脊髓灰质炎病毒风险,今后必须停止使用口服脊髓灰质炎疫苗,以彻底根除脊髓灰质炎。消灭脊髓灰质炎野病毒之后,将会严格管理来源于野毒株的传统脊髓灰质炎灭活疫苗生产,而用Sabin株研制的脊髓灰质炎灭活疫苗抗原性及免疫原性与传统脊髓灰质炎灭活疫苗不同。新型脊髓灰质炎疫苗形式有待开发。 本研究利用Bac-to-Bac杆状病毒表达系统表达脊髓灰质炎病毒Sabin2结构蛋白,由于脊髓灰质炎病毒的主要抗原位点位于结构蛋白VP1上,其功能蛋白3C及其前体3CD具有蛋白酶的功能,能将多聚蛋白P1剪切加工成结构蛋白VP0、VP3及VP1,因此,首先利用RT-PCR方法扩增得到脊髓灰质炎病毒Sabin2株基因VP1、P1、3C及3CD,分别构建以上基因的重组杆状病毒,然后将重组杆状病毒rBac-VP1、rBac-P1和rBac-3C、rBac-P1和rBac-3CD三组分别感染Sf9细胞进行表达。结果显示:分别克隆有Sabin2株基因VP1、P1、3C及3CD的重组杆状病毒表达质粒转染Sf9细胞后24小时出现细胞病变,72小时细胞病变明显;间接免疫荧光检测显示目的蛋白为细胞内表达;间接免疫荧光及Western-Blot结果显示三组均有VP1结构蛋白的生成,且均具有抗原性;rBac-P1和rBac-3C、rBac-P1和rBac-3CD分别共感染Sf9细胞两组有结构蛋白多聚体的生成,提示可能有病毒样颗粒的生成。 脊髓灰质炎病毒Sabin2株结构蛋白在昆虫杆状病毒表达系统中的成功表达,为脊髓灰灰质炎病毒分子生物学特性研究提供了新的科研思路,为新型脊髓灰质炎疫苗形式的开发奠定了基础,提供了新的技术路线。

关键词： 脊髓灰质炎病毒   非脊灰肠道病毒   病原学监测  

摘要： [目的]分析开展急性驰缓性麻痹(AFP)病例、健康人群粪便标本病原学监测结果,为保持无脊灰状态提供实验室依据。[方法]病毒分离及鉴定,脊灰病毒株型内鉴别,确定疫苗株或野病毒株。[结果]青海省2000～2009年AFP病例、健康人群粪便标本1613份,分离出脊髓灰质炎病毒22株,分离率为1.36%,其中脊灰I型疫苗株3株,脊灰II型疫苗株2株,脊灰III型疫苗株9株,脊灰混合型疫苗株8株,无脊灰野病毒株。分离到非脊灰肠道病毒(NPEV)187株,分离率为11.59%。[结论]该省2000～2009年无本地脊灰野毒株病例流行,阻断了脊灰本地野病毒的传播,保持了无脊灰状态。

关键词： replication   picornavirus  

摘要： Positive-strand RNA viruses include a large number of human and animal pathogens whose essential RNA-dependent RNA polymerases (RdRPs) share a structurally homologous core with an encircled active site. RdRPs are targets for antiviral drug development, but these efforts are hindered by limited structural information about the RdRP catalytic cycle. To further our understanding of RdRP function, we assembled, purified, and then crystallized poliovirus elongation complexes after multiple rounds of nucleotide incorporation. Here we present structures capturing the active polymerase and its nucleotide triphosphate complexes in four distinct states, leading us to propose a six-state catalytic cycle involving residues that are highly conserved among positive-strand RNA virus RdRPs. The structures indicate that RdRPs use a fully prepositioned templating base for nucleotide recognition and close their active sites for catalysis using a novel structural rearrangement in the palm domain. The data also suggest that translocation by RDRPs may not be directly linked to the conformational changes responsible for active site closure and reopening.

关键词： Antigens/metabolism   Base Sequence   Capsid Proteins/chemistry   China   Genome, Viral   Genome, Viral   Genotype   Humans   Molecular Sequence Data   Muscle Hypotonia/virology   Nucleotides/genetics   Paralysis/virology   Phenotype   Poliovirus Vaccine, Oral/metabolism   Sequence Homology, Nucleic Acid   Temperature  

摘要： Background: Ten uncommon natural type 3/type 2 intertypic poliovirus recombinants were isolated from stool specimens from nine acute flaccid paralysis case patients and one healthy vaccinee in China from 2001 to 2008. Principal Findings: Complete genomic sequences revealed their vaccine-related genomic features and showed that their first crossover sites were randomly distributed in the 39 end of the VP1 coding region. The length of donor Sabin 2 sequences ranged from 55 to 136 nucleotides, which is the longest donor sequence reported in the literature for this type of poliovirus recombination. The recombination resulted in the introduction of Sabin 2 neutralizing antigenic site 3a (NAg3a) into a Sabin 3 genomic background in the VP1 coding region, which may have been altered by some of the type 3-specific antigenic properties, but had not acquired any type 2-specific characterizations. NAg3a of the Sabin 3 strain seems atypical;other wild-type poliovirus isolates that have circulated in recent years have sequences of NAg3a more like the Sabin 2 strain. Conclusions: 10 natural type 3/type 2 intertypic VP1 capsid-recombinant polioviruses, in which the first crossover sites were found to be in the VP1 coding region, were isolated and characterized. In spite of the complete replacement of NAg3a by type 2-specific amino acids, the serotypes of the recombinants were not altered, and they were totally neutralized by polyclonal type 3 antisera but not at all by type 2 antisera. It is possible that recent type 3 wild poliovirus isolates may be a recombinant having NAg3a sequences derived from another strain during between 1967 and 1980, and the type 3/type 2 recombination events in the 39 end of the VP1 coding region may result in a higher fitness.

关键词： POLIO   VACCINATION   FLACCID dysarthria   IMMUNIZATION   COMMUNICABLE diseases -- Transmission   PREVENTION   POLIOMYELITIS vaccine   VIRUS-induced immunosuppression  

摘要： In China, 5 patients with acute flaccid paralysis (AFP) associated with type 2 vaccine-derived poliovirus (VDPV) were identified by an AFP surveillance system from 1996 through 2009. A maximum-likelihood tree shows that all 5 Chinese VDPVs were independent. These 5 VDPVs were 100-216 d old according to the number of synonymous substitutions per synonymous site and 176-292 d old according to the number of substitutions per site. This result indicates limited virus replication since the administration of the initiating oral polio vaccine (OPV) dose, which is consistent with the rapid evolution rate of poliovirus genomes. The above-mentioned VDPVs have important implications in the global polio eradication initiative. Localized, limited, and transient circulation may be typical of OPVs;hence, independent VDPVs could be found because of the large population and excellent surveillance system, which permitted early detection and response, but sustained transmission was limited because of high population immunity.

关键词： OPV derivatives   Antigenic properties   Immunity  

摘要： In this study, the immunity level of the southern Greek population in the 1-10-year, 11-20-year, 21-30-year and 31-40-year age groups with regard to Sabin vaccine strains and a collection of 11 recombinant and three non-recombinant poliovirus vaccine strains was determined. The results showed the lowest neutralization titre in the 21-30-year-age group against poliovirus type 3. Moreover, the capsid coding region of OPV (oral poliovirus vaccine) derivatives was sequenced in order to identify mutations that might lead to antigenic changes. In Sabin-1 derivatives a tendency of accumulation of mutations was observed in or near antigenic sites while in Sabin-2 and Sabin-3 derivatives in sites known to be involved in restoring neurovirulence or eliminating their temperature-sensitive phenotype. It was concluded that the combination of mutations in the capsid coding region and not the number of specific mutations in antigenic sites determines the antigenic properties of OPV derivatives and their reactivity with human sera. (C) 2010 Elsevier Ltd. All rights reserved.

关键词： Vaccine Strain   Oral Poliovirus Vaccine   Neutralise Antibody Titre   Oral Polio Vaccination   Geometric Mean Titre  

摘要： Background: In October 2007, the working group CEN/TC 216 of the European Committee for standardisation suggested that the Sabin oral poliovirus vaccine type 1 strain (LSc-2ab) presently used for virucidal tests should be replaced by another attenuated vaccine poliovirus type 1 strain, CHAT. Both strains were historically used as oral vaccines, but the Sabin type 1 strain was acknowledged to be more attenuated. In Germany, vaccination against poliomyelitis was introduced in 1962 using the oral polio vaccine (OPV) containing Sabin strain LSc-2ab. The vaccination schedule was changed from OPV to an inactivated polio vaccine (IPV) containing wild polio virus type 1 strain Mahoney in 1998. In the present study, we assessed potential differences in neutralising antibody titres to Sabin and CHAT in persons with a history of either OPV, IPV, or OPV with IPV booster. Methods: Neutralisation poliovirus antibodies against CHAT and Sabin 1 were measured in sera of 41 adults vaccinated with OPV. Additionally, sera from 28 children less than 10 years of age and immunised with IPV only were analysed. The neutralisation assay against poliovirus was performed according to WHO guidelines. Results: The neutralisation activity against CHAT in adults with OPV vaccination history was significantly lower than against Sabin poliovirus type 1 strains (Wilcoxon signed-rank test P < 0.025). In eight sera, the antibody titres measured against CHAT were less than 8, although the titre against Sabin 1 varied between 8 and 64. Following IPV booster, anti-CHAT antibodies increased rapidly in sera of CHAT-negative adults with OPV history. Sera from children with IPV history neutralised CHAT and Sabin 1 strains equally. Conclusion: The lack of neutralising antibodies against the CHAT strain in persons vaccinated with OPV might be associated with an increased risk of reinfection with the CHAT polio virus type 1, and this implies a putative risk of transmission of the virus to polio-free communities.