Computational protocol: Microbiome Dynamics of a Polychlorobiphenyl (PCB) Historically Contaminated Marine Sediment under Conditions Promoting Reductive Dechlorination

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Protocol publication

[…] Forward and reverse reads were trimmed for quality using Trimmomatic v. 0.32 () with the settings SLIDINGWINDOW:5:3 and MINLEN:275. The trimmed forward and reverse reads were merged using FLASH v. 1.2.7 () with the settings -m 25 -M 200. The merged reads were dereplicated and formatted for use in the UPARSE workflow (). The dereplicated reads were clustered, using the usearch v. 7.0.1090 -cluster_otus command with default settings. OTU abundances were estimated using the usearch v. 7.0.1090 -usearch_global command with -id 0.97. Taxonomy was assigned using the RDP classifier () as implemented in the parallel_assign_taxonomy_rdp.py script in QIIME (), using the MiDAS database v.1.20 (). The results were analyzed in R () through the Rstudio IDE using the ampvis package v.1.9.1 ().Evenness (H) and the taxonomic distinctness (TD) indices were used to describe the biodiversity in the original marine sediment and during the microcosm study and were calculated using Past version 3.10. [...] The 16S rRNA gene sequences were analyzed with the ARB software () using the SILVA 16S rRNA SSU Reference database release 102 (). Sequences were analyzed for chimera formation using the chimera-checking tool QIIME Software. The phylogenetic tree was constructed using the maximum likelihood method RAxML (). For the construction of the phylogenetic tree with sequences from the original marine sediment a total of 18 16S rRNA gene sequences were used and Chlamidyae was chosen as outgroup (Supplementary Table ).For the construction of the phylogenetic tree with sequences from marine sediment after lactate addition a total of 20 rRNA gene sequences were used and Acidobacteria was chosen as outgroup (Supplementary Table ). […]

Pipeline specifications

Software tools Trimmomatic, UPARSE, USEARCH, RDP Classifier, QIIME, ampvis, ARB, RAxML
Applications Phylogenetics, 16S rRNA-seq analysis
Organisms Bacteria, Dehalococcoides mccartyi
Chemicals Polychlorinated Biphenyls, Sulfur, Lactic Acid