The Mycobacterium Tuberculosis Dormancy Program

描述（由申请人提供）：世界上三分之一的人口感染结核分枝杆菌（Mtb），其中大多数感染是潜伏的。结核杆菌可以在肺病变中保持不活跃状态，几十年后才出现，为结核病的新爆发播下种子。此外，结核病是最难治疗的细菌感染之一，造成的死亡人数比任何其他细菌感染都多。芽孢杆菌以复制和非复制状态存在于一系列不同氧浓度和营养物质可用性的微环境中。在潜伏感染期间存活的杆菌可能以非复制状态存在，抗微生物药物对活跃生长的细菌有效，但对非复制细菌往往无效。对于结核分枝杆菌在潜伏感染或坏死性干酪样结核病变的恶劣微环境中存活下来的代谢机制，人们知之甚少，那里氧气有限，营养来源欠佳。真菌不能生长，但在没有有氧呼吸的情况下存活。因此，非复制厌氧状态被认为是体内持久性杆菌的主要模式。在没有有氧呼吸的情况下，Mtb需要一个功能性的电子传递系统来驱动ATP合成。然而，杆菌维持氧化还原平衡的核心代谢机制尚不清楚。因此，我们的核心研究问题是“当有氧呼吸被抑制时，结核分枝杆菌采用什么代谢机制？”至少有三个因素限制结核分枝杆菌的有氧呼吸：巨噬细胞产生的一氧化氮和一氧化碳的抑制作用，以及成熟肉芽肿的结构。所有这三种条件都强烈诱导了DosR调节，这是厌氧生存所必需的调节。然而，该调控子并没有编码一个完整的可识别的中间代谢途径。我们的表达分析和生化数据强烈表明，Mtb -一种专性需氧菌-在没有有氧呼吸的情况下维持了独特的多方面代谢中间途径。因此，在结核分枝杆菌作为一种坦率的人类病原体的整个进化过程中，它一直保持着一系列广泛的酶，这些酶似乎专门用于厌氧代谢功能。该途径预测结核分枝杆菌具有代谢脂质和所有其他主要碳源厌氧的潜力。目前迫切需要设计用于杀死非复制性厌氧杆菌的抗菌剂。我们的研究直接支持这一目标。新的抗细菌药物和药物组合通常针对缺氧/厌氧杆菌进行测试，但关于适当方案的问题以及我们对相关中间代谢途径的有限理解限制了合理的药物设计方法。我们的初步数据表明，我们正处于对结核分枝杆菌厌氧代谢的基本理解的边缘。我们的工作假设是：结核分枝杆菌采用一种新的无氧代谢循环，与DosR调节相结合，在结核病变内的非呼吸条件下赋予生存。为了验证这一假设，我们将研究提出的厌氧代谢途径的关键方面。

DESCRIPTION (provided by applicant): A third of the world's population is infected with Mycobacterium tuberculosis (Mtb), and most of these infections are latent. Tubercle bacilli can remain inactive in lung lesions only to emerge decades later to seed new outbreaks of tuberculosis. In addition, tuberculosis is one of the most difficult bacterial infections to treat and continues to cause more deaths than any other bacterial infection. Bacilli exist in replicating and non-replicating states in a range of microenvironments that vary in oxygen concentration and nutrient availability. The bacilli that survive during latent infection likely exist in a non-replicating state and antimicrobials, effective against actively growing bacteria, are often not effective against non-replicating bacteria. Little is known about the metabolic mechanisms employed by Mtb to survive latent infection or the hostile microenvironment of necrotic caseous tubercle lesions where oxygen is limited and nutrient sources suboptimal. Mtb cannot grow but endures in the absence of aerobic respiration. Therefore, the non-replicating anaerobic state is considered a prime model for persistent bacilli in vivo. In the absence of aerobic respiration Mtb requires a functional electron transport system to drive ATP synthesis. However, the core metabolic mechanisms by which the bacilli maintain redox balance are unknown. Therefore, our central research question is "What metabolic mechanisms are employed by Mtb when aerobic respiration is inhibited?" At least three factors limit Mtb aerobic respiration: the inhibitory effect of macrophage-produced nitric oxide and carbon monoxide, and the structure of mature granulomas. All three of these conditions strongly induce the DosR regulon, a regulon essential for anaerobic survival. However, the regulon does not encode a complete recognizable intermediary metabolic pathway. Our expression analysis and biochemical data strongly indicate that Mtb - an obligate aerobe - maintains a unique multifaceted intermediary pathway for metabolism in the absence of aerobic respiration. Thus, throughout the evolution of Mtb as a frank human pathogen, it has maintained an extensive array of enzymes which appear to be geared specifically for anaerobic metabolic functions. The pathway predicts that Mtb has the potential to metabolize lipids and all other major carbon sources anaerobically. Antimicrobials designed to kill non-replicating anaerobic bacilli are sorely needed. Our research directly supports this goal. New antimycobacterial drugs and drug combinations are routinely tested against hypoxic/anaerobic bacilli, but questions about the proper protocol and our limited understanding of relevant intermediary metabolic pathways limit a rational approach to drug design. Our preliminary data suggests we are at the brink of a fundamental understanding of Mtb anaerobic metabolism. Our working hypothesis is: Mtb employs a novel anaerobic metabolic cycle in conjunction with the DosR regulon to confer survival during non- respiring conditions within TB lesions. To test this hypothesis we will investigate key aspects of the proposed anaerobic metabolic pathway. PUBLIC HEALTH RELEVANCE: A third of the world's population is latently infected with Mycobacterium tuberculosis. Tuberculosis is one of the most difficult bacterial infections to treat and continues to cause more deaths than any other bacterial infection. The bacteria that persist during latent infection and antibacterial treatment are likely in non-replicating states with low levels of metabolic activity. Our research is designed to demonstrate the unique metabolic pathway used by M. tuberculosis to survive in a non-replicating state in order to provide mechanisms that can be targeted by novel antimicrobials.