Fig. 2: The linkage of the AS resistomes to microbiomes.

a Relationships detected by Procrustes analysis between the resistomes and bacterial community structure as measured by 16S genes extracted from the metagenomes. Metagenomic shotgun sequencing was performed for all activated sludge samples, and the 16S sequences were extracted and grouped at the genus level using Metaxa2. b Relationships detected by Procrustes analysis between the resistomes and bacterial community structure as measured by the 16S amplicon sequencing data. The dotted ends of lines represent the resistome position, while the undotted ends represent the bacteriome position. Vegan Procrustes test ‘protest’ with 999 permutations yielded a matrix-matrix correlation coefficient of 0.74 (protest, p = 0.001) for metagenome 16S-based bacterial community structure, and a matrix-matrix correlation coefficient of 0.70 (protest, p = 0.001) for 16S amplicon-based bacterial community structure. c The association between the ARG abundance (total ARG abundance and the top four major ARG groups) and the relative abundance of top 15 major bacterial phyla from 16S rRNA gene amplicon data (16S) or metagenomes (Shotgun). The circle-filled color corresponds to Spearman’s correlation coefficient. The asterisks ‘*’ denote significant correlations (two-sided p < 0.05 after adjustment for multiple testing). d The phylogenetic tree of metagenome-assembled genomes (MAGs) from global AS samples. The leaf colors indicate phylum groups. The bar heights outside the circle are proportional to the ARG count annotated in MAGs, and red bars represent the MAGs carrying multi-species mobile ARGs. Inner rings show the resistance gene abundances of the five major drug classes, with darker colors indicating higher abundances. e The mean count and relative abundances of ARGs encoding major resistance mechanisms or drug classes across phylogenetic groups. Error bars indicate standard deviations. Numbers on the top indicate the number of MAGs belonging to the phylogenetic groups. Source data are provided as a Source Data file.