Role of OxyR in P. aeruginosa Biofilm Resistance to H202

描述（由申请人提供）：在囊性纤维化（CF）气道疾病中，有令人信服的证据表明存在两个不同的临床阶段，即氧化阶段（早期CF）和厌氧阶段（慢性晚期CF）。在CF气道内厚厚的粘液中，细菌作为“生物膜”生长，这是一种发展形式，为生物体提供了增强对抗生素和杀生物剂的抗性。早期氧化阶段是基于嗜中性粒细胞快速和急剧流入上呼吸道，这是由细菌感染引发的事件。当受到刺激时，这些专职吞噬细胞会产生一种强有力的“呼吸爆发”，一种抗微生物产品，其中是过氧化氢（H2O2）。事实上，嗜中性粒细胞可以在吞噬溶酶体空泡内产生毫摩尔水平的H2O2。相比之下，血液中的H2O2水平低近1000倍。令人惊讶的是，缺乏H2O2响应性反式激活因子OxyR的主要CF病原体铜绿假单胞菌的突变体对H2O2非常敏感，并且即使在血液H2O2水平（微摩尔范围）存在下细菌也会死亡。如果OxyR在人类感染期间受损，细菌将不能引起全身感染，因为它们将经由H2O2介导的杀伤而死亡。因此，本提案的目标是确定铜绿假单胞菌的OxyR是否可以在各种铜绿假单胞菌感染期间作为药物靶标。该提议的目标是（i）定义OxyR在动物毒力和对人嗜中性粒细胞的抗性中的作用，（ii）定义在OxyR突变体中引起对H2O2的极度敏感性的病变以及什么OxyR控制的基因产物有助于最大或最小的保护，以及（iii）确定OxyR对于生物膜细菌对H2O2和氨基糖苷类的存活是否是关键的。

DESCRIPTION (provided by applicant): In cystic fibrosis (CF) airway disease, there is compelling evidence for two distinct clinical stages, an oxidative phase (early-stage CF) and an anaerobic phase (chronic, late-stage CF). Within the thick mucus lining the CF airways, the bacteria grow as a "biofilm," a form of development that affords organisms the luxury of enhanced resistance to antibiotics and biocides. The early oxidative phase is based upon a rapid and dramatic influx of neutrophils to the upper airways, an event triggered by bacterial infection. When stimulated, these professional phagocytes mount a potent "respiratory burst," an antimicrobial product from which is hydrogen peroxide (H202). In fact, neutrophils can generate millimolar levels of H202 within the phagolysomal vacuole. In contrast, H202 levels in blood are nearly 1000-fold lower. Surprisingly, a mutant of the major CF pathogen, Pseudomonas aeruginosa, lacking the H202-responsive transactivator, OxyR, is exquisitely sensitive to H202 and the bacteria perish even in the presence of blood H202 levels (micromolar range). Were OxyR to be compromised during human infection, bacteria would be unable to elicit a systemic infection because they would die via H202-mediated killing. Therefore, the goal of this proposal is to determine if OxyR of P. aeruginosa could serve as a drug target during various P. aeruginosa infections. The goals of this proposal are to (i) define a role for OxyR in animal virulence and resistance to human neutrophils, (ii) define the lesions that evoke exquisite sensitivity to H202 in the OxyR mutant and what OxyR-controlled gene products contribute to maximal or minimal protection, and (iii) determine whether OxyR is critical for survival of biofilm bacteria to H202 and aminoglycosides.