LotuS2: An Ultrafast and highly accurate tool for amplicon sequencing analysis . LotuS2

Background: Amplicon sequencing is an established and efficient method for profiling microbiomes. However, many available tools for analysis of amplicon data require both bioinformatics skills and high computational power to process big datasets and only some of them allow for long read amplicon data analysis. To bridge this gap, we developed the LotuS2 (Less OTU Scripts 2) pipeline, which enables the uncomplicated analysis of raw amplicons sequencing data, extremely fast and with high precision. Results: LotuS2 is available from GitHub, conda or via a Galaxy web interface. Six different sequence clustering algorithms and extensive pre- and postprocessing options are available in LotuS2, allowing for flexible data processing by both expert and naïve users. We benchmarked the QIIME2, DADA2, mothur and LotuS2 on four independent datasets. LotuS2 was on average 13 times faster than other pipelines and technical replicates of gut or soil microbiomes were consistently more reproducible when analyzed with LotuS2. LotuS2 also reconstructed the mock community most precisely, recovering the highest fraction of true positive genera and species enabled through the LotuS2 `seed extension` algorithm, 82% of reconstructed 16S rRNA genes, were 100% identical to the known references, the highest fraction achieved by any pipeline on the Mock community. Conclusion: LotuS2 is a slim and easy-to-use pipeline that is fast, precise and streamlined. High data usage rates and reliability enable cost-efficient microbiome analysis that can be executed on laptop computers and is freely available at http://lotus2.earlham.ac.uk/.