Exploiting Mycobacterium tuberculosis biofilm-derived phenotypes for transformative novel drug discovery

结核病（TB）在2018年造成130万人死亡。随着新冠肺炎对全球结核病控制计划的破坏，预计未来几年死亡人数将上升。结核病药物治疗正在失败，药物敏感性疾病的治疗需要4种药物联合治疗6个月，结核病占抗菌药物耐药性（AMR）感染的三分之一。目前迫切需要新的药物，但由于我们不了解作为化疗靶点的肺病变中杆菌生物膜样生长的生理学机制，这些努力受到了阻碍。该项目将应用尖端分子技术来模拟结核分枝杆菌（M.tb）的亚群，以模拟人类肺表型。该提案将结核分枝杆菌发病机理/转录组学（Waddell）、细菌生物膜（Webb）的专业知识与PHE-Porton的长期工业合作伙伴、药物发现专家（Bacon）合作。目标和方法：优化Waddell实验室（苏塞克斯）开发的现有分枝杆菌生物膜样生长模型，并利用Webb（南安普顿）的生物膜专业知识。微生物学培训（苏塞克斯），生物膜方法学培训（南安普顿）。描述由生物膜样生长产生的结核菌亚群。测量表型多样性对药物耐受性的影响。与Dolomite Bio一起开发单细胞/亚群水平的转录组学，Dolomite Bio是一家成熟的微流体公司，专门从事Dropseq应用的液滴细胞封装。分子生物学和细菌转录组学培训（苏塞克斯）：通过培根实验室（PHE-Porton）世界领先的CL 3-FCM设施，使用流式细胞术/流式细胞仪确定生物膜中结核的异质性。PHE-Porton的行业安置。建立微量滴定板结核生物膜测定法，以筛选新的范式转变杀菌化合物。来自所有合作伙伴和NBIC的早期药物发现培训。该项目将开发创新技术，以了解结核分枝杆菌的基本肺生物学，以及在生物膜中形成的各种细菌表型，确定新的药物靶点和治疗策略。

Tuberculosis (TB) killed 1.3 million people in 2018. Deaths are predicted to rise in the coming years as COVID has devastated TB control programmes worldwide.TB drug therapies are failing, treatment of drug-sensitive disease takes a combination of 4 drugs for 6 months, and TB accounts for one third of antimicrobial drug resistant (AMR) infections. New drugs are desperately required, however efforts are hampered as we do not understand the physiology of the biofilm-like growth of bacilli in pulmonary lung lesions that are the target of chemotherapy.This project will apply cutting-edge molecular techniques to characterise the sub-populations of Mycobacterium tuberculosis (M.tb) that exist when grown as a biofilm, to mimic the human lung phenotype. The proposal partners expertise in M.tb pathogenesis/transcriptomics (Waddell), bacterial biofilms (Webb), with a long-standing industrial partner at PHE-Porton, expert in drug discovery (Bacon).Objectives and Approaches:Optimise an existing model of mycobacterial biofilm-like growth developed in the Waddell lab (Sussex), with biofilm expertise from Webb (Southampton). Training in microbiology (Sussex), biofilm methodologies (Southampton).Characterise the tb sub-populations that are generated by biofilm- like growth. Measure the impact of phenotypic diversity on drug tolerance. Develop single cell/sub-population level transcriptomics with Dolomite Bio, an established microfluidics company specialising in the encapsulation of cells in droplets for Dropseq applications. Training in molecular biology and bacterial transcriptomics (Sussex).Define the heterogeneity of tb in biofilms using flow cytometry/FACS through a world-leading CL3-FCM facility in the Bacon lab (PHE-Porton). Industry placement(s) at PHE-Porton.Establish a microtitre plate tb biofilm assay to screen for new paradigm-shifting cidal compounds. Training in early drug discovery from all partners and NBIC. The project will develop innovative technologies to understand the fundamental lung biology of M.tb, and the diverse bacterial phenotypes that develop in biofilms, identifying novel drug targets and treatment strategies.