DZIF Microbiome sequencing facility
Within the German Center of Infection Research (DZIF), we have established a core NGS sequencing facility giving support to different research projects aiming at analyzing microbiota changes and its relation to different pathological manifestations. In addition, we are conducting different clinical studies where the analysis of intestinal and gastric microbiota is a major focus to understand disease outcome. Two of the most relevant projects currently ongoing include the assessment of microbiota changes upon H. pylori infection in vivo and understanding the role of the microbiome during organ transplantation. The bioinformatics analysis is completed broadly following the Mothur MISEQ SOP (Schloss et al. 2013 AEM) for OTUs and DADA-2 pipeline in R (Callahan et al. 2016 Nat. Methods) for ASVs generation, respectively and EzBioCloud 16S bacterial database for alignment and taxonomy purposes (Yoon et al. 2017 IJSEM). Other platforms like the Trimmomatic (trimming PE reads generated by Illumina MiSEQ; Bolger et al. 2014 Bioinformatics), CutAdapt (removal of adapter sequences, primers, poly-A tails and other unwanted sequences; Marcel M. 2011 EMBnet J.), Pandaseq (Assembling PE reads generated by Illumina MiSEQ; Masella et al. 2012 BMC bioinform.), FastTree (making phylogenetic trees which could also be then utilized for calculating phylogenetic indices like weighted and unweighted UniFrac; Price et al. 2010 PLoS One), Lefse Galaxy (discovery of significant microbial signatures between different study groups; Segata et al. 2011, Genome Biol.) were utilized as per the need arrived. R packages, Primer-6 and Graphpad Prism 8 were utilized for the statistical analyses. The generated data is stored in a pseudonymized form in a publicly available database like NCBI SRA or ENA and will be available for further research.