Creation of a tissue culture model for Toxoplasma gondii sexual development

描述(申请人提供)：弓形虫的性周期仅限于猫的肠道，在那里形成数百万个卵囊，然后从它们的粪便中排出。这项建议的目的是重述组织培养中弓形虫的有性周期。创造体外的性发育条件将使特定基因对生物过程的贡献和多余的途径得到快速分析。它还将允许对原生寄生虫的整个生命周期进行分子分析，因为弓形虫的无性发育已经可以在组织培养中进行。弓形虫组织培养有性发育的另一个好处是针对自身和其他病原体(如疟原虫)的弓形虫疫苗载体的稳定生产。表达约氏疟原虫或诺氏疟原虫环子孢子蛋白的重组弓形虫分别在小鼠和恒河猴中提供对疟疾感染的保护性免疫。作为卵囊生产的弓形虫疫苗载体将是理想的，因为它在几乎任何环境条件下都将是稳定的，而且它将是一种口服接种。我的实验室还在鉴定对弓形虫确定慢性感染至关重要的毒力基因。将这些提议的工作协同起来，将使我们能够创造一种减毒疫苗株，可以从细胞培养中生产出稳定的卵囊。与公共卫生相关：弓形虫的性周期仅限于猫的肠道，在那里形成数百万个卵囊，然后从它们的粪便中排出。这项建议的目的是重述组织培养中弓形虫的有性周期。弓形虫组织培养有性发育的另一个好处是针对自身和其他病原体(如疟原虫)的弓形虫疫苗载体的稳定生产。

DESCRIPTION (provided by applicant): The sexual cycle of Toxoplasma gondii is limited to the feline intestine where millions of oocysts are formed and subsequently excreted in their feces. The goal of this proposal is to recapitulate the T. gondii sexual cycle in tissue culture. Creation of ex vivo sexual development conditions will allow the contribution of specific genes to biological processes and redundant pathways to be rapidly analyzed. It will also allow for a molecular analysis of the complete lifecycle of a protistan parasite, as T. gondii asexual development can already be performed in tissue culture. Another benefit of tissue culture sexual development of T. gondii will be the stable production of a T. gondii vaccine vector against itself and other pathogens, such as Plasmodium. Recombinant T. gondii expressing circumsporozoite protein from Plasmodium yoelii or P. knowlesi provides protective immunity against malaria infection in mice and rhesus monkeys, respectively. A T. gondii vaccine vector produced as an oocyst will be ideal because it will be stable in virtually any environmental condition and it will be an oral inoculation. My laboratory is also identifying the virulence genes critical for T. gondii to establish a chronic infection. Synergizing the work from these proposals will allow us to create an attenuated vaccine strain that can be produced as a stable oocyst from cell culture. PUBLIC HEALTH RELEVANCE: The sexual cycle of Toxoplasma gondii is limited to the feline intestine where millions of oocysts are formed and subsequently excreted in their feces. The goal of this proposal is to recapitulate the T. gondii sexual cycle in tissue culture. Another benefit of tissue culture sexual development of T. gondii will be the stable production of a T. gondii vaccine vector against itself and other pathogens, such as Plasmodium.