CRISPR and Virulence Core

CRISPR 和毒力核心

• 批准号: 10540799
• 负责人: Bridget Marie Barker
• 金额: $ 21.39万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10540799
• 关键词: Affect; Algorithms; Antigens; Applied Genetics; Bioinformatics; Biological Assay; CRISPR/Cas technology; Candidate Disease Gene; Cell Culture Techniques; Cells; Centers for Disease Control and Prevention (U.S.); Clustered Regularly Interspaced Short Palindromic Repeats; Coccidioides; Coccidioides immitis; Coccidioides posadasii; Coccidioidomycosis; Coculture Techniques; Complex; Data; Data Analyses; Defect; Development; Diagnostic; Disease; Disease Outcome; Engineering; Fungal Drug Resistance; Gene Deletion; Gene Targeting; Genes; Genetic; Genetic Transcription; Goals; Human; Immune; In Vitro; Incidence; Infection; Label; Laboratories; Lead; Length of Stay; Macrophage; Marketing; Methodology; Modeling; Morphology; Mus; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Pattern; Phase; Phenotype; Plasmids; Play; Proteins; Reporting; Research; Research Proposals; Role; Sister; Technology; Testing; Therapeutic; Time; Transcript; Vaccines; Virulence; Virulence Factors; Visualization software; Work; candidate identification; cost; desert fever; experience; experimental study; gene function; genetic approach; genome analysis; genome wide association study; improved; in vitro Assay; insight; interest; mouse model; mutant; novel; pathogen; pathogenic fungus; programs; promoter; reverse genetics; tool; trait; vaccine development

PROJECT SUMMARY/ABSTRACT Little is known about virulence mechanisms for most emerging fungal pathogens. Coccidioides immitis and C. posadasii, are of specific concern due to rapid increase in disease (coccidioidomycosis, aka Valley fever) incidence in the US, with a more than 10-fold increase in reported disease over the last 20 years. The Centers for Disease Control and Prevention (CDC) report that average cost for treating Valley fever is $50,000 per patient, including extensive hospital stays and treatment regimes. Use of CRISPR/Cas9 and functional assessment of targeted gene deletion on virulence can help improve our understanding of mechanisms of pathogenicity and represent the foundational framework that is necessary for development of these approaches. We propose to functionally delete 10 gene targets in Coccidioides posadasii strain Silveira and assay these deletions for loss of virulence. We will further investigate pathogenicity in C. immitis strain RS for those genes that show a loss of virulence. Currently, few specific mechanisms of virulence have been investigated in Coccidioides. Analysis of these data will provide new insights into functions of Coccidioides genes, which will increase our understanding of fungal pathogenesis mechanisms. Finally, we will work to develop tools to engineer fluorescently labeled live strains for Coccidioides using mCherry expression that will increase tools for interrogating host:pathogen interactions in an in vitro cell culture assay. In total, this proposal represents needed improvements in functional assessment of virulence in an important, yet understudied, fungal pathogen.

项目总结/摘要 对于大多数新出现的真菌病原体的毒力机制知之甚少。粗球孢子菌（Coccidioides immitis）和C. 由于疾病（球孢子菌病，又名山谷热）的迅速增加， 美国的发病率，在过去20年中报告的疾病增加了10倍以上。中心 美国疾病控制和预防中心（CDC）报告说，治疗谷热的平均费用为每名患者50,000美元， 包括长期住院和治疗方案。CRISPR/Cas9的使用和CRISPR/Cas9的功能评估 靶向基因缺失可以帮助我们更好地理解致病机制， 代表了发展这些方法所必需的基本框架。我们建议 在Posadasii球孢子菌菌株Silveira中功能性缺失10个基因靶，并测定这些缺失的缺失， 毒性。我们将进一步研究C.犬恶螨菌株RS的那些基因显示出缺失， 毒性目前，很少有具体的毒力机制已被调查的球孢子菌。分析 这些数据将为球孢子菌属基因的功能提供新的见解，这将增加我们对球孢子菌属基因的了解。 真菌致病机制的研究最后，我们将致力于开发工具， 使用mCherry表达的球孢子菌菌株将增加询问宿主：病原体的工具 在体外细胞培养测定中的相互作用。总的来说，这一建议表明需要改进职能， 评估一种重要但尚未充分研究的真菌病原体的毒力。