Computational protocol: Local domestication of lactic acid bacteria via cassava beer fermentation

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Protocol publication

[…] Sequence processing was conducted in QIIME () using MacQiime (version 1.6.0, http://www.wernerlab.org/software/macqiime; QIIME, RRID:OMICS_01521). Quality filtered forward reads (Phred score > 20; 250 bp) were binned with barcodes corresponding to the respective sample IDs. Operational taxonomic units (OTUs) were assigned at 99% genetic similarity. Representative OTU sequences were aligned to the Greengenes database (October 2012 version) and assigned taxonomic nomenclature with an RDP classifier. We rarified all samples to 19,000 sequences for even sampling depth; two samples significantly below that threshold were omitted from further analysis.We also conducted a manual BLAST search against the NCBI 16S isolate database for the top 10 OTUs for exploratory purposes (NCBI BLAST, RRID:nlx_84530). We included these species-level NCBI database matches for visualization and reporting. Results from OTU clustering matched to the Greengenes database using an RDP classifier within the QIIME pipeline were used for all analysis purposes.Community similarity was calculated in two different ways: with the phylogeny-based UniFrac metric (unweighted), and by calculating the number of OTUs shared between samples. Unweighted UniFrac uses phylogeny-based branch lengths generated from our rarified dataset, comparing their fractions between samples to quantify dissimilarity between communities without regard for species abundance. We determined if differences were significant using PERMANOVA (Adonis method) in QIIME. Using the UniFrac matrix, we generated a PCoA plot in QIIME and included it as . We then used the QIIME generated OTU table to conduct a one-way ANOVA in SPSS (SPSS, RRID:rid_000042, version 20.0.0) to investigate differences in the number of shared OTUs across households and across villages (). […]

Pipeline specifications

Software tools QIIME, RDP Classifier, BLASTN
Databases Greengenes
Applications Phylogenetics, 16S rRNA-seq analysis
Organisms Manihot esculenta, Homo sapiens
Chemicals Lactic Acid