丙型肝炎病毒E1E2蛋白原核表达载体的构建及表达产物的鉴定

目的 :构建丙型肝炎病毒 (HCV)包膜糖蛋白 (E1E2 )基因的原核表达载体 ,为研究HCVE1E2蛋白属性及基因免疫奠定基础。方法 :设计HCVE1E2区基因上、下游引物 ,分别引入BglⅡ及EcoRI酶切位点 ,以含有HCVH株基因序列的质粒pBRTM/HCV1- 30 11为模板 ,通过PCR扩增获得HCVE1E2区基因片段 ,将其克隆入原核表达载体pRSETA ,然后通过酶切和序列测定对其进行鉴定 ;将此表达载体转化入表达菌BL2 1(DE3)pLySs诱导表达 ,并采用SDS -PAGE对表达产物进行鉴定。结果 :经酶切及序列测定分析表明 ,成功地构建了重组原核表达载体pRSETA -HCVE1E2 ;SDS -PAGE显示目的蛋白大量表达 ,且呈非溶状态。结论 :HCVE1E2融合蛋白可以在大肠杆菌中大量表达 ,为HCVE1E2蛋白的制备及基因疫苗的研制奠定了基础。     

髓样相关蛋白14真核表达载体的构建及其细胞内定位研究

目的:构建小鼠髓样相关蛋白14(MRP14)的真核表达载体,观察其在NIH3T3细胞中的表达定位情况。方法:提取BALB/c小鼠肝脏组织的总RNA,通过逆转录-聚合酶链反应(RT-PCR)扩增得到MRP14编码序列。然后将该编码序列克隆到带有血凝素(HA)标记的载体pcDNA3-HA上,随后转染NIH3T3细胞,在荧光显微镜下观察结果。结果:重组质粒经聚合酶链反应(PCR)、酶切和测序鉴定证明构建正确。转染实验发现,该质粒能够在NIH3T3细胞中表达,表达产物主要定位在细胞质中。结论:成功构建带HA标签的MRP14真核表达载体。该载体能在哺乳动物细胞中有效表达并正确定位,为下一步深入研究MRP14作用细胞的信号通路提供了一个重要工具。     

佛山口岸2002～2005年进境集装箱卫生检疫工作质量分析与探讨

目的通过掌握佛山口岸进出境集装箱卫生检疫查验结果,对集装箱卫生监管情况进行分析,探讨管理新模式。方法对2002-2005年佛山口岸进境集装箱报检查验资料进行整理分析。结果2002-2005年佛山口岸共受理出入境集装箱报检1 550 565个标箱,集装箱检疫呈现出逐年稳定增长的良好势头,但在检疫查验过程中发现集装箱卫生状况还不够理想。结论今后在集装箱检验检疫工作中不断改进,采取有效措施,防止公共卫生危害通过集装箱传入。     

Importance of the major splice donor and redefinition of cis-acting sequences of gutless feline foamy virus vectors

Foamy virus vectors are potent alternatives to lenti- and gamma-retroviral vectors for gene therapy. To construct and optimize gutless feline foamy virus (FFV) replication-deficient (RD) vectors, viral elements essential for optimal efficient marker gene transduction were characterized and fine-mapped and packaging clones constructed. For these purposes, new Gag and Pol expression clones which allow efficient expression of packaging proteins and vectors carrying deletions in coding and non-coding regions of the genome were constructed and functionally evaluated. These studies demonstrate that the 5′ major splice donor (5′ SD) is indispensable for RD vectors while defined mutations introduced to inactivate the gag start codon improve transgene delivery efficiency. Based on these findings, new gutless FFV vectors were generated yielding un-concentrated vector titers above 10⁵ transducing units (TU)/ml. By minimizing the second cis-acting sequence in the pol gene, only 3.8 kb viral sequences are maintained in the novel gutless FFV RD vectors.     

Effects of genetic factors and infection status on wing morphology of Triatoma dimidiata species complex in the Yucatán peninsula, Mexico

Triatoma dimidiata is one of the main vectors of Chagas disease, and it has been shown to be a species complex. In the Yucatán peninsula, Mexico, vector populations are non-domiciliated, and the transmission of Trypanosoma cruzi thus critically relies on vector dispersal. This leads us to study the morphologic variations in T. dimidiata wings with respect to genetic factors (sex and genotype at the ITS-2 locus), geographic location, and T. cruzi-infection status. Females were found to have larger and more symmetrical wings than males. Wing shape was influenced by ITS-2 genotypes, although differences are unlikely sufficient to allow taxonomic discrimination of the sibling species. Hybrids were shown to have similar fluctuating asymmetries in wing size and shape as parental species, but the level of asymmetry in shape varied slightly between villages. The two later findings are consistent with a high level of gene flow between parental species, and the high dispersal potential of these non-domiciliated vectors. More surprisingly, individuals infected with T. cruzi were found to have larger wings than non-infected ones. This effect, which was consistently observed across sexes, genotypes and villages, is likely to be due to a direct impact of T. cruzi on insect development. Sex and infection status are thus likely to be key factors influencing vector dispersal with important impacts on disease transmission, since dispersal directly controls the domestic abundance of these vectors. These aspects should be investigated further to fully capture the ecology and evolution of Chagas disease transmission by non-domiciliated vectors.     

Beyond the ‘back yard’: Lay knowledge about Aedes aegypti in northern Australia and its implications for policy and practice

Controlling dengue fever in Australia and internationally, relies heavily upon the actions of residents as well as community education and awareness of the risks. Although it has been well established in medical anthropology that the success of health interventions is highly dependent upon a clear grasp of lay knowledge of disease, limited attention has been given to lay understandings of dengue fever and its vectors in the extant literature. We begin addressing this hiatus through an examination of north Queensland residents’ knowledge of the breeding habitats of Aedes aegypti mosquitoes. Building on the insights of earlier social research, we use factor analysis to examine the results of a series of randomly selected telephone surveys and compare responses over time and between cities.Our analysis confirms that many people assume that Ae. aegypti is ubiquitous in the landscape, that it lives and breeds not only around the home, but also in a variety of geographical spaces located beyond the suburban ‘backyard’, and beyond the control of local residents. Lay understandings appear to be placing people at risk from dengue, influencing the mosquito management practices of local residents and acting as a source of resistance to public health messages that focus on individual responsibility. A way forward through the provision of new information that challenges key assumptions is provided in the discussion. We argue that rather than dismissing lay understandings as ignorance, strategies, practices and policy based on a detailed understanding of this knowledge will mean that practitioners are better able to address these assumptions and will likely be more effective at educating the public of the risks posed by dengue.     

河南省淮滨县实施全球基金疟疾项目效果的评价

[目的]评价河南省淮滨县2003～2005年实施全球基金疟疾项目期间的疟疾疫情、防治措施及防治效果。[方法]收集淮滨县2003～2005年疟疾流行、防治措施、防治效果及媒介监测资料,分析流行情况,评价防治措施及效果。[结果]2003～2005年淮滨县共报告疟疾病例1765例,平均年发病率为8.93/万。2004年和2005年疟疾发病分别比上年下降53.58%和72.04%。建立发热病人镜检站18个,血检发热病人39070人,发现650例阳性病人。捕获中华按蚊4744只,未发现嗜人按蚊,疟疾暴发发点氟氯氰菊酯药物浸泡蚊帐340顶。氯、伯喹治疗现症病人1765人,休止期服药7571人,流行季节预防服药2471人。培训专业技术116人次,疟防宣传6259份,督导检查108次。居民和中小学生疟防知识知晓率由2003年的14.36%和9.36%提高到2004年的54.21%和47.36%。2005年144例疟疾病人平均确诊时间(2.82±2.89d)显著低于河南省的平均确诊时间(6.64±10.49d)(单样本t检验,t=-15.844,P=0.000)。[结论]在全球基金疟疾项目的支持下,河南省淮滨县疟防工作成绩显著,以项目管理促进疟防的方式值得推广。     

纳米粒子介导特异基因转染的实验研究

目的：观察纳米粒子携带特异性基因转染的可行性及其效率,方法：应用聚乳酸聚乙醇酸共聚物和聚乙烯醇包载特异性反义单核细胞趋化蛋白－1基因,制备纳米级粒子混合物。检测其包埋率,体外释放情况及粒度。以阳离子脂质体为对照,利用培养的平滑肌细胞,应用聚合酶链反应（PCR）观察其为真核细胞传递基因的可能性。采用移植静脉内膜增生动物模型,应用RNA斑点杂交和病理形态学分析。观察其在体内的基因转染、表达及生物学效应。结果：制备的纳米粒子－DNA粒度为150－300nm,包埋率为：0．9％,体外释放时间为2周左右。PCR结果提示：纳米粒子可将目的基因转移至平滑肌细胞中,体内实验显示：纳米粒子可实现体内局部基因转染,表达,发挥相应的效应。结论：纳米粒子可用于特异性基因的转染。     

湖北省媒介按蚊生态与疟疾发病关系的研究

目的 了解湖北省嗜人按蚊和中华按蚊的生态习性以及与疟疾发病的关系。方法调查各县（市）疟疾发病情况及当地传播媒介的种类和生态习性。结果湖北省疟疾流行与媒介的种类、栖息场所、吸血习性等有密切关系，通过调查发现嗜人按蚊人房密度为84．04％，中华按蚊密度为15，96％。嗜人按蚊的媒介能量平均高出中华按蚊44．47倍，嗜人按蚊区疟疾发病数占全省发病数的81．38%-93．84％。结论在嗜人按蚊存在的地区，疟疾发病较高。应根据媒介生态习性、分布特点及发病规律采取不同的抗疟措施。     

建立国际航行船舶媒介生物监测预警系统的思路

随着全球经济一体化和海运业的蓬勃发展，媒介生物以国际航行船舶、集装箱、货物等为载体侵入我国，对我国人民和动物的健康构成极大威胁。为了保障我国的卫生安全和经济安全、应对国外技术性贸易措施和履行即将实施的新《国际卫生条例》，在科学性和实用性原则的基础上，提出建立国际航行船舶媒介生物监测预警系统。该系统框架包括4部分：媒介生物信息模块、国际航行船舶媒介生物监测模块、媒介生物风险评估模块和国际航行船舶媒介生物风险预测管理模块。实现这一系统，我们必须：采用相同的方法和标准、长期坚持媒介生物监测工作、开发应用计算机软件、加大财力投入和培养人才。