Refining microbiome diversity analysis by concatenating and integrating dual 16S rRNA amplicon reads.

Understanding the role of human gut microbiota in health and disease requires insights into its taxonomic composition and functional capabilities. This study evaluates whether concatenating paired-end reads enhances data output for gut microbiome analysis compared to the merging approach across various regions of the 16S rRNA gene. We assessed this approach in both mock communities and Korean cohorts with or without ulcerative colitis. Our results indicate that using the direct joining method for the V1-V3 or V6-V8 regions improves taxonomic resolution compared to merging paired-end reads (ME) in post-sequencing data. While predicting microbial function based on 16S rRNA sequencing has inherent limitations, integrating sequencing reads from both the V1-V3 and V6-V8 regions enhanced functional predictions. This was confirmed by whole metagenome sequencing (WMS) of Korean cohorts, where our approach improved taxa detection that was lost using the ME method. Thus, we propose that the integrated dual 16S rRNA sequencing technique serves as a valuable tool for microbiome research by bridging the gap between amplicon sequencing and WMS.