Details all analysis related to the BREATHE Sputum bacteriome analysis
This github repository houses R notebooks of codes used in the manuscript Abotsi RE et al. "Sputum bacterial load and bacterial composition correlate with lung function and are altered by long term azithromycin treatment in children with HIV-associated chronic lung disease" published in Microbiome 2023. https://doi....
The following README gives an overview of the overall structure of the repository, and important notes on how to run the scripts.
BREATHE_Sputum_bacteriome_published_2023
1.Microbiome_published_material -- contains manuscript (pdf) and supplementary material (pdf, xlsx) published in BMC Microbiome
2.Preprocessing_QC_generating_phyloseq -- contains scripts used for sequence processing to generating first phyloseq object using TADA - Targeted Amplicon Diversity Analysis using DADA2, implemented in Nextflow found here https://github.com/h3abionet/TADA. The output from the pipeline is also included.
3.Decontamination_steps -- houses R scripts used to check extraction efficiency,reproducibility between and within run, in silico decontamination etc
4.Generating_final_phyloseq_object-- houses R scripts used to generate final phyloseq object made up of the final ASVs, samples and metadata retained after decontamination step
5.16S_copies_Alpha_diversity-- houses R scripts used to determine the 16S copies and alpha diversity results and figures
6.Within-patient_change_in_beta_diversity-- houses R scripts used to generate the within-patient change in beta diversity results and figures
7.Between_group_beta_diversity-- houses R scripts used to generate the between-group beta diversity results and figures
8.PCoA-- houses R scripts used to generate the PCoA plots to visualise beta diversity
9.PERMANOVA-- houses R scripts used to conduct PERMANOVA tests
10.Rel.abun.plots.microeco-- houses R scripts used to generate the barplots of relative abundance and group mean barplots
11a.DA_analysis_AZM_only_Baseline_Week48 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the AZM arm only from baseline and 48 week visit using 10 methods
11b.DA_analysis_AZM_only_Baseline_Week72 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the AZM arm only from baseline and 72 week visit using 10 methods
11c.DA_analysis_AZM_only_Week48_Week72 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the AZM arm only from 48 and 72 week visit using 10 methods
11d.DA_Analysis_AZM_Placebo_Baseline -- houses R scripts used to conduct ddifferentially abundant taxa analysis on samples from the AZM and Placebo arm at baseline using 10 methods
11e.DA_Analysis_AZM_Placebo_Week48 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the AZM and Placebo arm at week 48 using 10 methods
11f.DA_Analysis_AZM_Placebo_Week72 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the AZM and Placebo arm at week 72 using 10 methods
11g.DA_analysis_Placebo_only_Baseline_Week48 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the Placebo arm only from baseline and 48 week visit using 10 methods
11h.DA_analysis_Placebo_only_Baseline_Week72 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the Placebo arm only from baseline and 72 week visit using 10 methods
11i.DA_analysis_Placebo_only_Week48_Week72 -- houses R scripts used to conduct differentially abundant taxa analysis on samples from the Placebo arm only from 48 and 72 week visit using 10 methods
11j. DA_Analysis_Supplementary_Tables -- houses supplementary tables from the differentially analysis test on samples from the AZM arm only (Baseline and 48 weeks, and 48 and 72 weeks) and from AZM and Placebo arms at 48 weeks using 10 methods
12.ANCOM2_Analysis_AZM_only_allvisits-- houses R scripts used to determine the differentially abundant taxa in AZM arm at all visits using Ancom2
13.Simper_analysis-- houses R scripts used to conduct SIMPER analysis
14.Associations-- houses R scripts used to determine associations between clinical and socio-demographic factors and alpha diversity
|-- 16S_copies -- contains scripts used to run linear mixed effect models on 16S rRNA copies and clinical and socio-demographic factors
|-- Change_Aitchison_distance_lung_function -- contains scripts used to run linear regression on within-participant change in beta diversity (Aitchison's distance) and within-participant change in lung function metrics- FEV1z and FVCZ
|-- Change_genus_abun_Lung_function -- contains scripts used to run linear regression on within-participant change in the relative abundances of selected genera and within-participant change in lung function metrics- FEV1z and FVCZ
|-- Genus_association_analysis -- contains scripts used to run linear mixed effect models on the relative abundances of selected genera and clinical and socio-demographic factors using MaAsLin2
|-- Shannon_diversity_lme -- contains scripts used to run linear mixed effect models on Shannon diversity indices and clinical and socio-demographic factors
|-- stata_analysis_16S_paper -- contains scripts used to manually generate trial arm and visit interaction terms for used in the linear mixed effect models
15.Final_DA_heatmaps-- houses R scripts used to determine the differentially abundant taxa using the other 9 methods and generating heatmap from the results
Data dictionary -- houses excel files of variables and their interpretation
Figures_made_inppt-- houses differentially abundant taxa analysis results summary in figures on a Microsoft Powerpoint file. Also contains figure on number of samples collected and analysed