Supplementary Dataset and Workflows for 'Soil origin affects gut microbiota and immune response in a wild rodent'

Dataset and Workflows for "Soil origin affects gut microbiota and immune response in a wild rodent" Description of the data and file structure This dataset includes code to preprocess short read data deposited in ENA SRA (PRJEB87226) using QIIME2 and subsequent data analysis using R with related packages, processed 16S and ITS2 datasets required for the R pipeline, and reproducible nf-core workflow reports for RNA sequencing and differential expression analysis. Please see project GitHub repo for updates on scripts. Files and variables File: readme.md Description: This file. File: FECES_16S_QIIME2.sh Description: Bank vole faecal 16S sequence data pre-processing pipeline using QIIME2. File: ITS2_QIIME2_vsearch.sh Description: Bank vole faecal ITS2 sequence data pre-processing pipeline using QIIME2. File: dna-sequences_16S.fasta Description: Denoised and decontaminated 16S V3-V4 DNA sequences produced by QIIME2 pipeline. File: dna-sequences_ITS2_c97.fasta Description: Denoised and decontaminated 16S V3-V4 DNA sequences clustered at 97% sequence identity produced by QIIME2 pipeline. File: feature-table_ITS2_c97.tsv Description: Decontaminated fungal ITS2 OTU abundance table produced by QIIME2 pipeline. File: feature-table_16S.tsv Description: Decontaminated bacterial 16S ASV abundance table produced by QIIME2 pipeline. File: rooted-tree_16S.nwk Description: Rooted phylogenetic tree for bacterial 16S data. File: metadata_feces-soil_16S.txt Description: Metadata file for 16S samples to be loaded in R. Variables SampleID: Sample ID Sample_ID: Sample alias Pup_ID: Subject ID of the experimental animal Sample_type: Sample type. Sampling_timepoint: Sampling time pre- (T1) or post (T2) treatment. is.neg: Identifier for negative control used in DECONTAM in QIIME2 pipeline. pup_birth_date: Birth date of subject. cage: Cage number of subject. sex: Sex of subject. Soil_location: City or national park from which experimental soil was collected. Soil_type: Main experimental factor - Control, Park or Urban. Pup_head: Subject head width at birth Pup_wt: Subject body weight at birth (g) End_head: Subject head width at end of experiment (mm) End_wt: Subject body weight at end of experiment (g) Testis_wt: Left testis weight (g) (males only) Sampling_date: Date of endpoint sampling Mother_head: Head width of the subject's mother at subject's weaning (mm). Mother_wt: Body weight of the subject's mother at subject's weaning (g). Mother_ID: ID of subject's mother. Used as random intercept in linear mixed models to control for genetic component. Mother_type: Information on subject's mother. FEAST: Subsetting factor for sourcetracking using FEAST. Env: Sample name in format required for FEAST source tracking. SourceSink: Source or Sink population identifier for FEAST. id: Sample identifier in format required for FEAST. sample_accession: ENA sample accession ID File: metadata_ITS2_c97.tsv Description:Metadata file for ITS2 samples to be loaded in R. Variables As above. File: taxonomy_ITS2_c97.guilds.txt Description: Taxonomic classifications for ITS2 data, including FUNGuild annotations. File: taxonomy_16S.tsv Description: Taxonomic classifications for 16S data. File: firstdistances_bacteria_bray.tsv Description: Table of Bray-Curtis first distances of faecal bacteria pre- and post-treatment (=distance of the same subject between two time points). Data produced by QIIME2 pipeline, to be plotted in R. Variables SampleID: Sample ID SubjectID: Sample alias Distance: Distance Group: Experimental factor File: firstdistances_bacteria_jaccard.tsv Description: Table of Jaccard first distances of faecal bacteria pre- and post-treatment (=distance of the same subject between two time points). Data produced by QIIME2 pipeline, to be plotted in R. Variables As above File: firstdistances_fungi_bray.tsv Description: Table of Bray-Curtis first distances of faecal fungi pre- and post-treatment (=distance of the same subject between two time points). Data produced by QIIME2 pipeline, to be plotted in R. Variables As above File: firstdistances_fungi_jaccard.tsv Description: Jaccard first distances of faecal fungi pre- and post-treatment (=distance of the same subject between two time points). Data produced by QIIME2 pipeline, to be plotted in R. Variables As above File: FEAST_F_contributions.txt Description: Combined fungal source contributions from soil and food to faecal communities. Data produced in "faecal_metataxonomics_markdown.html" pipeline and externally edited in excel. Used as input for FEAST figure. Data also available in excel file Supplementary Data SI8. Variables Sample: Sample ID Soil: Source contribution of soil mixture to which subject was exposed. Food: Source contribution of food pellets. Unknown: Unknown source component. Group: Experimental factor Location: Origin of soil mixture File: faecal_metataxonomics_corestats.html Description: Microbial diversity analyses in R markdown. All required input files are published along this pipeline. File: faecal_metataxonomics_FEAST.html Description: Microbial source tracking analysis in R markdown. All required input files are published along this pipeline. File: faecal_metataxonomics_diffabund.html Description: Microbial differential abundance analysis in R markdown. All required input files are published along this pipeline. File: nfcore_RNAseq.html Description: Nextflow nf-core/RNAseq workflow report and settings for preprocessing of bank vole colonic mRNA reads. Includes extensive quality control and run settings. File: nfcore_differentialabundance.html Description: Nextflow nf-core/differentialabundance workflow report for exploratory data analysis and differential expression analysis. File: medoid_subsampling.R Description: R script for unsupervised subsampling for RNA extraction based on *a priori* knowledge of amplicon sequencing data. File: soil2gut.network.R Description: Script for microbial co-occurrence network analysis. Code/software Amplicon sequencing data preprocessing: QIIME2 distribution qiime2-amplicon-2024.10. Preprocessing pipeline and settings are described in Materials & Methods and files "FECES_16S_QIIME2.bash" and "ITS2_QIIME2_vsearch.bash". RNAseq data preprocessing and differential expression analysis: Data was processed using nf-core/rnaseq v3.18.0 (doi: [10.5281/zenodo.1400710](https://doi.org/10.5281/zenodo.1400710)) of the nf-core collection of workflows ([Ewels *et al.*, 2020](https://doi.org/10.1038/s41587-020-0439-x)), utilising reproducible software environments from the Bioconda ([Grüning *et al.*, 2018](https://doi.org/10.1038/s41592-018-0046-7)) and Biocontainers ([da Veiga Leprevost *et al.*, 2017](https://doi.org/10.1093/bioinformatics/btx192)) projects. The pipeline was executed with Nextflow v24.10.3 ([Di Tommaso *et al.*, 2017](https://doi.org/10.1038/nbt.3820)). Full details available in "nfcore_RNAseq.html" and "nfcore_differentialabundance.html". Data analysis: R 4.3.1. Analysis pipeline is available in R markdown file "faecal_metataxonomics_markdown.html". All required input files are included in this release. Required packages are displayed in the markdown file.