Meta-transcriptomic analysis of host-microbiota interactions in a longitudinal study in CF patients

患有囊性纤维化（CF）的患者会出现下呼吸道的多微生物感染以及慢性炎症宿主反应，导致肺功能的稳步下降。下一代测序技术已经破译了呼吸道微生物群的组成。然而，许多研究只在横断面设计中完成，但需要纵向研究来了解改变的微生物组在疾病进化中的作用。在德国肺研究中心（DZL）资助的一个项目框架下，我们目前正在通过16S rRNA测序分析一大批囊性纤维化患者的微生物群进化。自2013年以来，我们在海德堡转化肺研究中心（TLRC）的一项纵向研究中纳入了318名患者，同时收集了6500多份样本。在这项研究中，现在已经证明微生物群的进化和宿主的免疫反应是密切相关的。对宿主和微生物群的功能相互关系进行公正的研究需要一种元转录组学方法，而这不在当前研究的范围内。我们进行了一项双RNAseq的试点研究，并证明了用痰样本进行meta转录组分析以研究宿主和微生物群的功能输入的可行性。我们现在想把重点放在一个定义明确的患者亚群上，深入分析微生物群进化和宿主反应之间的相互作用。我们的队列中的患者每3-4个月定期采样一次，以及当他们出现恶化时。在每次访问中，将采集痰样本进行双RNAseq元转录组分析。基于先前的分析，我们可以定义临床疾病发展不同的亚组，从而允许信息丰富的研究设计。一个特别的焦点将是共生细菌的功能，这实际上是有争议的讨论。因此，这个附加项目提案的目标是在我们已经建立的CF队列中使用元转录组学来分析宿主和微生物群的功能相互关系。以下问题将被探讨：微生物转录组和宿主转录组在一年内有多稳定？微生物转录组如何与宿主转录组相关？微生物优势对活性微生物群稳定性的影响是什么？它是否会改变人类转录组？急性加重对微生物组和宿主的短期和长期功能变化的影响是什么？微生物和宿主转录组如何与临床结果相关？

Patients suffering from cystic fibrosis (CF) develop polymicrobial infections of the lower airways as well as a chronic inflammatory host response resulting in a steady decline in lung function. Next generation sequencing has deciphered the composition of the airways’ microbiota. However, many studies have only been done in cross-sectional designs, yet longitudinal studies are needed to understand the role of the altered microbiome in the evolution of the disease. We are currently analyzing the microbiome evolution via 16S rRNA sequencing in a large cohort of cystic fibrosis patients in the framework of a project funded by the German Center for Lung Reasearch (DZL). Since 2013, we have included 318 patients in a longitudinal study with meanwhile over 6,500 samples at the translational lung research center (TLRC) in Heidelberg. During the study it has now turned out that microbiome evolution and host immune responses are closely linked. An unbiased study of the functional interrelation of host and microbiota requires a meta-transcriptomic approach which is not in the scope of the running study. A pilot dual RNAseq study was performed by us and shows the feasibility of meta-transcriptomic analysis with sputum sample to study the functional input from both, host and microbiota. We would now like to focus on a well-defined subset of patients to analyze in depth the interplay of microbiome evolution and host response over time. Patients within our cohort are sampled regularly every 3-4 months as well as when they suffer from exacerbation. On each of those visits, sputum samples will be taken for dual RNAseq meta-transcriptomic analysis. Based on the previous analysis we can define subgroups that differ in clinical disease development therefore allowing an informative study design. A special focus will be the function of commensal bacteria which is actually controversially discussed.The objective of this add-on project proposal therefore is to use meta-transcriptomics in our already established CF cohort to analyze the functional interrelation of host and microbiota. The following questions will be approached: How stable are the microbial transcriptome and the host transcriptome over a year? How does the microbial transcriptome correlate with the host transcriptome? What is the impact of microbial dominance on the stability of the active microbiota and does it modify the human transcriptome? What is the impact of exacerbation on short term and long term functional changes of the microbiome and of the host? How do microbial and host transcriptome correlate to the clinical outcome?