16S-FASAS: an integrated pipeline for full-length 16S rRNA gene sequencing data analysis

16S rRNA full-length amplicon sequencing based on next generation sequencing platform (16S-FAS-NGS) is an emerging cost-effective technology with higher accuracy and resolution in species identification. We developed 16S full-length amplicon sequencing analysis software (16S-FASAS), a new flexible tool for 16S full-length amplicon sequencing data analysis. 16S-FASAS consists of data quality control, de novo assembly, annotation and visualization modules. We verified the performance of 16S-FASAS on both mock and real samples. In mock communities, we proved that taxonomy assignment by MegaBLAST with 16S-FASAS was superior to operational taxonomic unit (OTU) clustering with Quantitative Insights Into Microbial Ecology 2 (QIIME2), resulting in correct species-level classification of 85.7% (6/7) of taxa and the expected community structure. When further applied to fecal samples, we found that the full-length 16S rRNA gene datasets generated contigs grouped into 79 and 85 genera, while only 22% (17/79) and 19% (16/85) of which were detected by the V4V5 datasets with Illumina platforms. Moreover, full-length 16S rRNA gene sequencing technology showed better resolutions than V4V5 in analyses of relative abundance frequency. 16S-FASAS is a valuable tool that helps researchers process and interpret the results of full-length 16S rRNA gene sequencing. Depending on the full-length amplicon sequencing technology, we can make a more accurate report on bacterial complexity of microbiome samples by the 16S-FASAS pipline. 16S-FASAS is freely available for use at https://github.com/capitalbio-bioinfo/FASAS.