Polymicrobial interactions in Crohn's Disease

克罗恩病中的多种微生物相互作用

• 批准号: 10223109
• 负责人: Mahmoud A Ghannoum
• 金额: $ 61.74万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-08 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10223109
• 关键词: Address; Adhesions; Affect; Anaerobic Bacteria; Anti-Bacterial Agents; Antifungal Agents; Bacteria; Candida; Candida albicans; Candida tropicalis; Chronic; Clinical Research; Coculture Techniques; Colitis; Colon; Communities; Crohn' s disease; Data; Diagnostic; Environment; Escherichia coli; Exhibits; Exposure to; Family; Feces; Filament; Flare; Gastrointestinal tract structure; Gene Expression; Generations; Genes; Goals; Habits; Hyphae; Immune response; In Vitro; Indoles; Infection; Inflammation; Inflammatory; Inflammatory Bowel Diseases; Interruption; Intestinal Mucosa; Intestines; Knock-out; Lactobacillus; Letters; Lymphoid Cell; Mediating; Microbe; Microbial Biofilms; Modeling; Modification; Mucous Membrane; Mus; Oral; Organism; Outcome; Pathogenicity; Patients; Phase; Phenotype; Play; Prevalence; Process; Production; Pyrrolidonecarboxylic Acid; Recovery; Relapse; Reporting; Resolution; Role; Saccharomyces; Sampling; Serratia marcescens; Severities; Sodium Dextran Sulfate; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Symptoms; Testing; Therapeutic; Thick; Tissues; Trichosporon; Tryptophan; Ulcerative Colitis; Virulence Factors; Yeasts; aryl hydrocarbon receptor ligand; bacteriome; design; dextran sulfate sodium induced colitis; experimental study; fungus; gut microbiota; healing; host microbiota; in vivo; in vivo evaluation; in vivo imaging; indoleacetic acid; inflammatory disease of the intestine; inhibitor/antagonist; insight; interleukin-22; intestinal injury; metabolomics; microbial; microbiome; microorganism; microorganism interaction; mouse model; mutant; mycobiome; novel therapeutics; pathogen; polymicrobial biofilm; preclinical study; response; transcriptome sequencing

The role of gut fungal community (“mycobiome”, MYC) and interactions between the bacteriome (BM) and fungal communities in Crohn’s Disease (CD) have been relatively ignored. Recently, we compared the gut BM and MYC of CD patient’s to their healthy relatives and showed that abundance of the fungus Candida tropicalis (CT) was positively correlated with the bacteria Serratia marcescens (SM) and Escherichia coli (EC). We analyzed the number of CD patients that had all 3 species; the number CD patients exhibiting all 3 organisms was 30%, compared to 9% in healthy controls. Critically, abundance of all three organisms is 6.2 fold higher in CD patients vs. controls. Thus, the prevalence of this combination of microorganisms in the GI tracts of CD patients is significant. We further showed that ex-vivo, these polymicrobial triple species cooperated to form robust biofilms (TSBs) that were significantly increased compared to those formed by single-species (SSBs) or double-species (CT+EC or CT+SM, DSBs). Supporting data showed the interaction between CT, SM, and EC was specific; substitution of Trichosporon spp. (TC) or Saccharomyces fibuligera (SF) did not increase biofilm formation. However, preliminary data using Candida albicans (CA) as a control comparator reported by others to be elevated in CD patients, did cause increased TSB biofilm formation, indicating that this is a Candida specific effect. Importantly, we validated that the 3 pathogens formed robust biofilms in vivo (an anaerobic environment) using a dextran sodium sulfate (DSS) model of murine ulcerative colitis. Metabolomic analyses of supernatants from TSBs identified 11 significantly increased metabolites compared to SSBs or DSBs. Of these, 5-oxoproline (5-OP), was the most elevated metabolite (131-fold higher in TSBs compared to SSBs or DSBs). Also, Indole-3-acetic acid (IAA), formed as a result of Lactobacilli metabolizing tryptophan, which can act as an endogenous ligand for aryl hydrocarbon receptors (AhR), mediating innate lymphoid cell (ILC) production of IL-22, that in-turn triggers the inhibition of CA colonization, was also significantly increased. These 2 metabolites influence candidal virulence factors (filamentation, adhesion and auto-aggregation). Although these data are significant and indicate a strong polymicrobial interaction in CD, they do not establish the underlying mechanisms of these microbial interactions. This proposal focuses on determining the mechanism/s underlying the interaction of mixed species microbes using complementary in vitro and in vivo approaches. Our hypothesis is that SM, EC and Candida (CT and CA), in the setting of biofilms, interact cooperatively to exacerbate intestinal inflammation and exacerbate symptoms To address the mechanism(s) we will: (1) Determine the mechanism(s) metabolites formed by CT, EC, and SM in TSBs alter intestinal outcomes; (2) Identify gene changes underlying Candida modifications that increase pathogenicity using microbial mutant strains and RNAseq approaches; and (3) Determine whether exposure of TSBs to antibacterial, antifungal or metabolite inhibitors modulates the severity of intestinal inflammation in vivo. via multispecies interactions.

肠道真菌群落(“MYC”)的作用及细菌群(BM)与真菌的相互作用 克罗恩病(CD)的社区相对被忽视。最近，我们比较了Gut BM和MYC CD患者的健康亲属，并显示热带假丝酵母菌(CT)的丰度 与粘质沙雷氏菌(SM)和大肠埃希菌(EC)呈正相关。 我们分析了 CD患者具有全部3种细菌的数量；CD患者表现出所有3种细菌的数量为30%， 相比之下，健康对照组的这一比例为9%。关键的是，在CD患者中，这三种微生物的丰度都高出6.2倍 与控制的对比。因此，这种微生物组合在CD患者的胃肠道中的流行率是显著的。 我们进一步表明，在体外，这些多微生物三重物种相互合作，形成了强大的生物膜(TSB)， 与单物种(SSB)或双物种(CT+EC或 CT+SM、DSB)。支持数据表明，CT、SM和EC之间的相互作用是特定的； 毛孢霉(Trichosporon spp.)(TC)或啤酒酵母(SF)对生物被膜的形成无促进作用。然而，初步的 使用白色念珠菌(CA)作为对照对照的数据，据其他人报道，CD患者的水平上升，确实如此 引起TSB生物被膜形成增多，表明这是念珠菌特有的作用。重要的是，我们验证了 这3种病原体在使用葡聚糖钠的体内(厌氧环境)形成了强大的生物膜 硫酸盐(DSS)建立小鼠溃疡性结肠炎模型。11株TSB上清液的代谢组学分析 与SSB或DSB相比，代谢物显著增加。其中，5-羟基脯氨酸(5-OP)含量最高 代谢物升高(TSB比SSB或DSB高131倍)。此外，吲哚-3-乙酸(IAA)，形成 由于乳杆菌代谢色氨酸，色氨酸可以作为芳香烃的内源配体 受体(AhR)，介导先天淋巴样细胞(ILC)产生IL-22，进而触发CA的抑制 移民率也显著增加。这两种代谢物影响念珠菌毒力因子 (丝化、粘合和自动聚集)。尽管这些数据意义重大，并表明 CD中的多个微生物相互作用，它们没有建立这些微生物相互作用的潜在机制。 这项建议的重点是确定混合物种微生物相互作用的潜在机制/S 体外和体内互补的方法。 我们的假设是SM、EC和假丝酵母菌(CT和CA)，在 设置生物膜，协同作用加剧肠道炎症和加重症状 解决机制(S)我们将：(1)确定机制(S) CT、EC和SM在TSB中形成的代谢物改变肠道结局；(2)识别基因变化 利用微生物突变株和RNAseq对提高致病性的念珠菌进行潜在的修饰 方法；以及(3)确定TSB是否暴露于抗菌、抗真菌或代谢物 抑制物在体内调节肠道炎症的严重程度。 通过多物种的相互作用。