Polymicrobial interactions between commensal obligate anaerobic bacteria and cystic fibrosis pathogen P. aeruginosa

摘要 很明显，人类微生物区系几乎影响到人类健康的方方面面，包括 直接影响某些疾病的临床结果。调节这些影响的机制是 通常是传入病原体和共生微生物之间的多菌相互作用。在囊性纤维化中 (Cf)，一种遗传性疾病，患者经常受到慢性细菌感染的困扰，最明显的是 铜绿假单胞菌、常住厌氧菌对疾病转归的影响相互矛盾且效果不佳 学习。然而，基于微生物区系的研究表明，特定的居民必须进行厌氧呼吸 细菌与改善CF结局有关，有限的证据表明，特定的专性 厌氧菌对铜绿假单胞菌的定植有抑制作用。事实上，我最近发现专性厌氧症 呼吸道细菌猫卟啉单胞菌通过非接触抑制铜绿假单胞菌的生长 机制。对猫疫霉基因组的基因组挖掘显示，没有已知与之相关的基因簇。 因此，通过抗菌合成，我们假设卡介苗肺炎杆菌通过一种新的方式抑制铜绿假单胞菌。 机制。该提案将采取多种方法来研究卡介苗疫霉对P.catoniae的抑制。 铜绿假单胞菌。为此，我们将构建和筛选猫疫霉转座子突变株文库，以 确定负责抑制活性的基因(S)。将利用质谱学对P。 卡介菌“分泌组”鉴定潜在的抗P.铜绿假单胞菌化合物(S)。全球铜绿假单胞菌 将通过RNA-seq检测和测量对P.catoniae的转录反应。此外，铜绿假单胞菌 TN-SEQ将被用来阐明哪些基因与在存在卡他肺炎杆菌的情况下存活有关。 此外，我们将评估细菌生活方式(浮游生物膜与生物膜)的影响，CF相关的环境 刺激，以及添加其他专性厌氧呼吸细菌对这种相互作用。中的更改 在CF相关条件下的生物膜生物地理学，包括其他共生体的存在 用荧光共聚焦显微镜进行评估。综上所述，这项提议试图为 最终可能揭示新的抗菌剂和/或分子靶标的细菌间行为和生活方式(S) 目的：治疗慢性铜绿假单胞菌感染，改善CF患者的临床预后。

ABSTRACT It has become apparent that the human microbiota impacts virtually every facet of human health, including directly influencing clinical outcomes for certain diseases. The mechanisms that mediate these effects are often the polymicrobial interactions between incoming pathogens and commensal microbes. In Cystic Fibrosis (CF), a genetic disease in which patients are often afflicted with chronic bacterial infections, most notably Pseudomonas aeruginosa, the effect of the resident anaerobes in disease outcome is conflicting and poorly studied. However, microbiota-based studies have revealed that specific resident obligate anaerobic respiratory bacteria are associated with improved CF outcomes, and limited evidence suggests that specific obligate anaerobic bacteria impair P. aeruginosa colonization. Indeed, I recently discovered that the obligate anaerobic respiratory bacterial species Porphyromonas catoniae inhibits P. aeruginosa growth by a contact-independent mechanism. Genomic mining of the P. catoniae genome revealed no gene clusters known to be associated with antimicrobial synthesis, we therefore hypothesize that P. catoniae inhibits P. aeruginosa by a novel mechanism. The proposal will take a multifarious approach to investigate P. catoniae inhibition of P. aeruginosa. To this end, a P. catoniae transposon mutant strain library will be constructed and screened to identify the gene(s) responsible for inhibitory activity. Mass spectrometry will be utilized to characterize the P. catoniae “secretome” to identify potential anti-P. aeruginosa compound(s). The P. aeruginosa global transcriptional response to P. catoniae will be examined and measured by RNA-seq. Moreover, P. aeruginosa Tn-seq will be used to illuminate which genes are associated with survival in the presence of P. catoniae. Furthermore, we will assess the effects of bacterial lifestyle (planktonic vs. biofilm), CF relevant environmental stimuli, and the addition of other obligate anaerobic respiratory bacteria on this interaction. Alternations in biofilm biogeography under CF relevant conditions including the presence of other commensals will be assessed with fluorescence confocal microscopy. En masse, this proposal seeks to provide novel insight into interbacterial behavior and lifestyle that ultimately could reveal novel antimicrobials and/or molecular target(s) to treat chronic P. aeruginosa infections and improve the clinical outcomes for CF patients.