Microbial Single-Cell Transcriptomics Links Gut Microbiota Functional States to Metabolic Remodeling in Mice

Overview

This repository contains the analysis code and data processing pipelines associated with the paper:
“Microbial Single-Cell Transcriptomics Links Gut Microbiota Functional States to Metabolic Remodeling in Mice”

Repository Structure

scripts/

Original scripts used in the study are organized within this directory.

Input data

Large input files required to run the analysis scripts are available on figshare: DOI: 10.6084/m9.figshare.32685366

Input data

Large input files required to run the analysis scripts are available on figshare:

DOI: 10.6084/m9.figshare.32685366

Please download and extract the corresponding Figure*_Input archives into each figure's Input/ directory before running the scripts.

Input data

Large input files required to run the analysis scripts are available on figshare:

DOI: 10.6084/m9.figshare.32685366

Please download and extract the corresponding Figure*_Input archives into each figure's Input/ directory before running the scripts.

1. Reference_genome_comparison_and_species_classification/

This directory contains all scripts used for benchmarking the three reference genome resources and performing species identification using a Kraken2-based classifier. The included scripts cover:

FASTQ preprocessing for taxonomic classification:
Read filtering and quality control, Barcode/UMI extraction
Species classification scripts:
Batch classification of FASTQ files with the previously described Kraken2-based workflow
Parsing, summarizing, and exporting genus-level and species-level reports
Reference genome comparison utilities:
Scripts to evaluate mapping / assignment rates across the three reference sets
Scripts to compare database content FastANI-based genome similarity comparison between reference catalogs

Each script contains detailed parameters, tool versions, and instructions for execution.

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2. smClassify_and_downstream_analysis/

This directory contains all downstream R scripts used to generate the figures in the manuscript. Each script corresponds directly to a specific figure or figure panel.

Included scripts:

Figure 2
smClassify pipeline scripts
Community composition summaries

Figure 3
Cell clustering and dimensionality reduction
Differential gene expression analysis
KEGG pathway enrichment
Functional-cluster distribution
Species proportion analysis
Subcluster characterization

Figure 4
PLS-DA analysis
Differential metabolite analysis
Network construction and visualization
KEGG pathway mapping
Metabolite boxplots and dotplots (amino acids, bile acids, LCFA, PUL-related metabolites)

Figure 5
Analysis of community-embedded functional states
Cross-species functional complementation within the gut microbiota

Figure 6
Subpopulation analysis of M. gordoncarteri
Disrupted nitrogen- and stress-response programs linked to host nitrogen imbalance