P4: SPATIOTEMPORAL EXPRESSION OF BURKHOLDERIA PSEUDOMALLEI GENES

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 类鼻疽伯克霍尔德氏菌是一种潜在的B类生物恐怖制剂，可引起类鼻疽病这一全球性新发传染病。在整个热带地区发生了数千例类鼻疽病，在美国旅行者（平民和军人）中报告了几例病例。B。类鼻疽通过不同阶段感染宿主细胞：i）进入宿主细胞内吞囊泡，ii）囊泡逃逸进入细胞质，iii）细胞质复制，iv）利用宿主肌动蛋白进行细胞质移动，v）将自身弹射到相邻宿主细胞中，以通过膜突起传播感染。我们假设，不同的基因表达，以促进这种细菌通过这些不同阶段的细胞感染的过境。通过基因表达分析，长期目标是了解B发病的分子机制。在一些实施方案中，所述抗体相对于受感染细胞的细胞内环境而与类鼻疽病毒结合。两个目标将确定必要的基因和基因表达的时空尺度上的真核巨噬细胞模型。目的1将确定在不同的细胞感染过程中的时间尺度上表达的基因。这些基因的表达将进一步证实，通过基因融合和突变分析和视频延时记录。目标2将采用一种新的高通量负选择方法来鉴定感染过程所必需的基因，这将通过实时PCR、荧光基因融合和感染过程的延时记录来证实。我们预计，实现这一目标将产生关键的干预策略，新的药物靶点，以及治疗和预防类鼻疽的候选疫苗。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Burkholderia pseudomallei is a category B potential bioterrorism agent, which causes melioidosis, a global emerging infectious disease. Thousands of cases of melioidosis have occurred throughout the tropics, and several cases have been reported in American travelers (civilian and military). B. pseudomallei infects host-cells through various stages: i) entry into host cell endocytic vesicles, ii) vesicular escape into the cytoplasm, iii) cytoplasmic replication, iv) utilize host actin for cytoplasmic mobility to v) catapult themselves into neighboring host-cells to spread the infection through membrane protrusions. We hypothesized that various genes are expressed to facilitate the transit of this bacterium through these different stages of cellular infection. Through gene-expression analyses, the long-term objective is to understand the molecular mechanism of pathogenesis of B. pseudomallei with respect to the intracellular environment of an infected cell. Two aims will identify essential genes and genes expressed on a spatiotemporal scale in a eukaryotic macrophage model. Aim 1 will identify genes expressed on a temporal scale during the various cellular infection processes. The expression of these genes will further be confirmed by reporter-gene-fusion and mutational analyses and video time-lapse recording. Aim 2 will employ a novel high-throughput negative-selection approach to identify genes essential for the infection process, which will be confirmed by real-time PCR, reporter-gene-fusion, and time-lapse recording of the infection process. We expect that achieving the objective will yield critical intervention strategies, novel drug targets, and vaccine candidates for the treatment and prevention of melioidosis.