Fig. 1

Metapangenomic workflow. a Pangenome construction. (1) All putative protein-coding gene sequences (colored block arrows) are extracted from each bacterial genome (colored bacilli above genes) to be included in the pangenome and (2) clustered into homologous gene clusters via blastp results grouped by the Markov Clustering Algorithm (sequence variants cartoonized as shades of the same color). (3) These gene clusters become the central dendrogram of the pangenome. Note that the gene clusters are organized by occurrence in genomes, not based on the order found in a particular genome. The detection of each gene cluster in a genome is visualized by filling in to indicate the presence or absence of each gene cluster across the genomes. The genomes are ordered by a dendrogram (top right) based on each genome’s gene cluster content. b Metagenomic mapping. (1) The exact genomes as above in a are used as a reference onto which reads from metagenomes are mapped. Gene-level coverage for each gene is then calculated. (2) These coverages are plotted for all genes from a given genome to show that genome’s gene-level representation in those samples. (3) Environmental representation is evaluated for each gene to decide whether that gene is environmentally core (gene’s median coverage > 0.25 of the genome’s median coverage across metagenomes from that environment) or environmentally accessory (gene’s median coverage < 0.25 of the genome’s median coverage). c Metapangenome construction. The environmental representation from b is then summarized for all genes and then overlaid onto the pangenome created in a—the inner layers show the genomic representation of the pangenome, and the outer layer shows the environmental representation of the pangenome. This outer layer summarizes the fraction of genes in each gene cluster that were environmentally core or accessory in those metagenomes (callout)