Assembly and analytical validation of a metagenomic reference catalog of human gut microbiota based on co-barcoding sequencing

Abstract

Human gut microbiota is associated with human health and disease, and is known to have the second-largest genome in the human body. The microbiota genome is important for their functions and metabolites; however, accurate genomic access to the microbiota of the human gut is hindered due to the difficulty of cultivating and the shortcomings of sequencing technology. Therefore, we applied the stLFR library construction method to assemble the microbiota genomes and demonstrated that assembly property outperformed standard metagenome sequencing. Using the assembled genomes as references, SNP, INDEL, and HGT gene analyses were performed. The results demonstrated significant differences in the number of SNPs and INDELs among different individuals. The individual displayed a unique species variation spectrum, and the similarity of strains within individuals decreased over time. In addition, the coverage depth analysis of the stLFR method shows that a sequencing depth of 60X is sufficient for SNP calling. HGT analysis revealed that the genes involved in replication, recombination and repair, mobilome prophages, and transposons were the most transferred genes among different bacterial species in individuals. A preliminary framework for human gut microbiome studies was established using the stLFR library construction method.

Keywords: horizontal gene transfer (HGT); insertions/deletions (INDELs); metagenomic; single nucleotide polymorphism (SNPs); stLFR.