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有两个不同的临床阶段，氧化期（早期CF）和厌氧期（慢性晚期CF）。在CF气道内的厚粘液中，细菌生长为一种“生物膜”，这种发育形式使生物体对抗生素和杀菌剂的抵抗力增强。早期氧化阶段是基于嗜中性粒细胞快速而剧烈地涌入上呼吸道，这是由细菌感染引发的事件。当受到刺激时，这些专业的吞噬细胞会产生一种强有力的“呼吸爆发”，一种由过氧化氢（H202）产生的抗菌产物。事实上，中性粒细胞可以在吞噬体液泡内产生毫摩尔水平的H202。相比之下，血液中的H202水平要低近1000倍。令人惊讶的是，主要CF病原体铜绿假单胞菌（Pseudomonas aeruginosa）的一种突变体，缺乏对H202反应的反激活因子OxyR，对H202非常敏感，即使在血液中存在H202水平（微摩尔范围），细菌也会死亡。如果OxyR在人类感染过程中被破坏，细菌将无法引发全身性感染，因为它们将通过h2介导的杀死而死亡。因此，本研究的目的是确定P. aeruginosa的OxyR是否可以作为各种P. aeruginosa感染的药物靶点。本提案的目标是：(i)确定OxyR在动物毒力和对人类中性粒细胞的抗性中的作用，（ii）确定在OxyR突变体中引起对H202敏感的病变，以及哪些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是否对生物膜细菌对H202和氨基糖苷的生存至关重要。

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.