Phylosymbiosis with limited functional divergence in bee gut microbiota

# Repository overview This repository contains the scripts, workflows, and associated data resources used for the study **“Phylosymbiosis with limited functional divergence in bee gut microbiota”**. The study investigates the evolutionary and ecological processes shaping gut microbiome variation across bees (Anthophila), testing whether phylosymbiosis, the tendency for closely related hosts to harbour more similar microbial communities, can emerge without strict microbial inheritance. Using gut bacterial community data from 1,989 samples representing 60 bee genera worldwide and spanning all major bee families, we combined comparative phylogenetic approaches, host trait analyses, microbial phylogenies, and predicted functional profiling to disentangle the contributions of host evolutionary history and environmental context in structuring bee gut microbiomes. The repository includes the computational workflows, statistical analyses, and data processing steps required to reproduce the main findings of the study. All scripts are organized to facilitate transparency, reproducibility, and further investigation of host–microbe evolutionary dynamics in bees. --- # Repository contents ## 1. Data.zip This folder contains the raw and processed datasets required to perform the analyses, including: * **ASV abundance table:** `asv_table.merge.txt`* **Predicted KEGG functional profiles of bacterial communities:** `kegg_abundance.rds`* **Bacterial phylogenetic tree:** `seq2.afa.treefile`* **Bacterial taxonomy assignment:** `taxonomy.vsearch`* **Sample names and identifiers:** `samples.csv`* **Bee phylogenetic tree:** `BEE_mat6b_genera_p8pmAa.treefile`* **Standardized bee taxonomy:** `bee_taxonomy_paths.csv`* **Sample metadata from the different projects included in this study:** `Bee_Metadata.zip`* **Bee trait database extracted from the literature:** `Traits.zip`* **Raw ASV sequences:** `asv_raw.zip` --- ## 2. Scripts.zip This folder contains the R and shell scripts used for data processing, quality control, merging, phylogenetic reconstruction, functional prediction, and statistical analyses. * `0_1_Bee taxonomy.R` and `0_2_Bee_Trait_Database.R` are used to harmonize bee taxonomy across projects and assign host traits at the subgenus level, respectively.* Bacterial 16S rRNA sequences were aligned using `muscle5.sh`, and bacterial phylogenetic trees were reconstructed using `iqtree.sh`. Bacterial functional profiles were predicted using `picrust.sh`.* Scripts `1_XXX.R` to `11_XXX.R` are numbered according to the workflow described in the manuscript. They progress from raw data processing to statistical modelling, visualization, and generation of the main results.* `metabarcoding_tools_0-1a.R` contains custom functions used throughout the analyses.* `Summary_Table_Metadata.R` generates Supplementary Table 1. --- ## 3. Outputs This folder contains intermediate and final outputs generated by the scripts in `Scripts.zip`. The main outputs correspond to comparative phylogenetic models assessing: * **Taxonomic diversity:** H* **Phylogenetic diversity:** PD* **Functional diversity:** FD Models based on beta diversity analyses are indicated with the suffix `_comp`. Results generated at taxonomic levels other than genus are provided in: `Bac_Taxo_levels.zip` --- ## 4. Plots This folder contains the figures generated for the study, including the main and supplementary plots. The original bee images used for Figure 5 are provided separately in: `Bee_Pictures.zip`