DEVELOPMENT OF AN AXENIC CULTURE FOR PNEUMOCYSTIS

描述：(改编自申请者的摘要)垫子博士最近 成功建立了支持有限的无菌文化体系 Pc的生长，类似于组织培养所实现的。它是 这个项目的总体目标是轴心培养PC。第一个具体 目的是试图确定产生 最适合培养的生物。为了加快文化发展， 研究人员还计划开发快速方法来评估 生物体数量和生存能力。然后，这些方法将应用于 实现第二个目标，对PC进行轴向培养。调查员 将尝试开发一种通过以下方式支持Pc增长的介质 遵循其他机构使用的营养评估和生化分析 成功地为其他人建立了这种文化的调查人员 原教旨主义者。通常需要的各种浓度的营养素块 目前将对微生物培养的Pc生长进行评估 无菌培养基或改良的粗培养基组分。每种植物的最佳浓度 区块将通过缩小稀释范围来识别。一次 确定后，区块的最佳浓度将保持不变 同时对另一块进行测试和优化。增长和生存能力将是 通过包括放射性标记前体在内的各种方法进行评估 生物的合并、计数和荧光分析。第三 该项目的目标是尝试描述生命周期阶段和 无菌培养中的生长动力学。超微结构和动力学 将使用分析来确定复制的生命周期阶段 培养，确定增长率，并监视 有机体。因为寄生原生生物通常会降低它们的毒力 在长时间培养的情况下，这种毒性通常是 与表面抗原相联系，培养生物的抗原性 将在超微结构上进行评估。这些项目的设计目的是 第一，提供新的或改进的评估有机体数量的方法 生存能力。第二，预计关于 从受感染的肺部分离出的微生物的生理状态将是 获得。最后，希望增加对 Pc的代谢能力，此外，无菌的发展 将获得Pc的培养。

DESCRIPTION: (Adapted from applicant's abstract) Dr. Cushion has recently succeeded in establishing an axenic culture system which supports limited growth of Pc, similar to that achieved with tissue cultures. It is the overall goal of this project to axenically culture Pc. The first specific aim is to attempt to identify isolation procedures that produce the healthiest organisms for culture. To expedite culture development, the investigator also plans to develop rapid methods for evaluation of organism numbers and viability. These methods will then be applied to accomplish the second goal, to axenically culture Pc. The investigator will attempt to develop a medium which supports the growth of Pc by following nutritional evaluations and biochemical analyses used by other investigators who have successfully established such cultures for other protists. Various concentrations of blocks of nutrients commonly required for culture of microbes will be evaluated for Pc growth in the present axenic medium or an improved crude medium. Optimal concentrations of each block will be identified by narrowing the range of dilutions. Once identified, the optimal concentration of a block will remain constant while another block is tested and optimized. Growth and viability will be assessed by a variety of methods including radiolabeled precursor incorporation, enumeration of organisms and fluorometric assay. The third goal of the project is to try to characterize the life cycle stages and kinetics of growth in axenic culture. Ultrastructural and kinetic analyses will be used to identify the life cycle stages replicating in cultures, determine the growth rate, and monitor the health of the organisms. Since parasitic protists commonly decrease their virulence when cultured over long periods and this virulence has oftentimes been linked to surface antigens, the antigenicity of the cultured organisms will be ultrastructurally evaluated. The projects have been designed to provide, first, new or improved methods of assessing organism number and viability. Secondly, it is anticipated that information on the physiologic state of organisms isolated from infected lungs will be obtained. Lastly, it is hoped that increased understanding of the metabolic capabilities of Pc and, in addition, the development of axenic culture for Pc, will be obtained.