Microbiome Analysis

Microbiome research has witnessed significant growth within the field of biomedical research, becoming one of the most rapidly advancing areas. Coined by Joshua Lederberg, the term “microbiome” was proposed “to signify the ecological community of commensal, symbiotic, and pathogenic microorganisms that share our body space and have been all but ignored as determinants of health and disease”. The microbiome encompasses all microorganisms along with their genomic data, structural elements, metabolites, and environmental conditions within a specific environment. Advancements in genomic technologies, including next-generation sequencing (NGS) have enabled in-depth exploration of microbial communities, yielding numerous insights into the intricate relationship between the human microbiome, health, and disease. Particularly, it could be shown that the complex ecological communities of trillions of microorganisms associated to humans are not only crucially involved in digestion and fermentation processes, but also exhibit protective and structural functions within the human host. Changes in community composition (i.e. dysbiosis) have been linked to various systemic diseases in humans, including inflammatory bowel disease (IBD), allergies, diabetes, skin conditions and neurological disorders. Given these links between microbiome and (chronic) diseases, and the potential for targeted interventions, current research focus on understanding how signals from human-associated microorganisms influence the human immune responses and disease aetiology and how this could be harnessed for precision medicine efforts in the coming years.

Since its establishment in 2014, the Microbiome Platform at the Institute of Clinical Molecular Biology (IKMB) has been at the forefront of microbiome research, applying standardized high-throughput methods to make significant contributions to the research agenda of Kiel University. While collaborating primarily with scientists/physicians of the University Medical Center Schleswig Holstein (UKSH) and the University of Kiel (CAU), the Microbiome Platform is also listed within the Research Infrastructure portal of the DFG (https://risources.dfg.de/detail/RI_00517_en.html) and collaborates as well with external institutes in Germany and abroad. Over the past 8 years, 200 national as well as international collaborative projects, processing a remarkable number of over 100,000 samples.

To ensure high quality of sample preparation with respect to extraction and 16S rDNA sequencing, the Microbiome Platform also participates in the INSTAND EQA scheme “Bacterial Genome Detection – Molecular Genetic Analysis of the Human Microbiome (580)” yearly. The actual certification applies to both regions used (V1V2 and V3V4) and was awarded in July 2023.

Fig. 1 The Microbiome Platform has profound knowledge and protocols for efficient DNA extraction methods, encompassing human, animal, and plant samples.

Sample Collection and Processing
Based on the profound knowledge within the microbiome research field, we are advising and assisting with writing applications and study planning to ensure that applications meet the requirements for downstream analysis and research purposes. In addition, we are providing Standard Operating Procedures (SOPs) for the optimal collection and storage of (human) fecal or other relevant samples for the planned study. The extraction methods for various sample types (stool, swabs, soil) originated from various hosts and environments and analysis tools are scalable, partly automated and used in experiments from just a few observation points to larger cohorts of thousands of patients.

Sequencing Strategies
The molecular techniques used for the description of microbiota are either based on marker genes, which allow taxonomic investigation on the species level (Who is there? Amplicon high-throughput sequencing) or use broader metagenomic methods to analyse the metabolic repertoire of respective biological states, e.g. metagenomic (Who is there and what could be their function?) and metatranscriptomic sequencing (Who is there and what cellular processes are they performing?).

Amplicon sequencing strategies established and offered:

• 16S rDNA V1V2 for detection of Bacteria
• 16S rDNA V3V4 for detection of Bacteria
• 16S rDNA nested-approach for detection of Archaea
• ITS2 rDNA for detection of Fungi

Metagenome and metatranscriptome sequencing

Data Analysis
The Microbiome Platform employs the DADA2 workflow for quality control and processing amplicon raw sequencing data of large datasets (https://benjjneb.github.io/dada2/bigdata.html) resulting in abundance tables of amplicon sequence variants (ASVs) tailored the sequenced regions in our lab, which can be found here. If required, further taxonomic annotation using Bayesian classifier and databases like SILVA is performed and provided to collaborators. The Platform also offers two up-to-date pipelines for the processing of metagenomic raw sequencing data, available for both internal and external users (https://github.com/ikmb/MAGScoT and https://github.com/ikmb/TOFU-MAaPO).

Team and Contact 
The conducted work in the laboratory is expertly managed and conducted by highly trained technicians, and the data is meticulously organized in an internal database. For registering a new project, please access this database or contact the platform leader.

The regular platform meeting is open for collaborators who want to discuss or prioritize their ongoing projects at the IKMB and takes place bi-weekly Wednesdays at 08:25 AM (odd weeks only).