Resistome profiles of the Han River based on metagenome sequencing and high-throughput qPCR (HT-qPCR) analyses. a Sampling sites surveyed in this study shown on a map of the Han River (blue) along with human population density. b ARG density estimated from shotgun metagenomes and HT-qPCR. For metagenome reads, reads per kilobase of sequence per million mapped reads (RPKM) values of ARGs were normalized by the average RPKM of 40 single-copy genes (SCGs). For HT-qPCR data, copy numbers of targeted ARGs were normalized by those of bacterial 16S rRNA genes. c ARG diversity estimated by Chao richness index. The Chao index of each sample corresponds to the estimated number of ARGs present in the sample. d Heatmaps of ARG density distribution throughout the sampling sites according to antibiotic class. e Dendrogram of various sample parameters based on Pearson’s correlation distance matrix. The correlation with ARG density is shown to the right of each variable. *P < 0.05, **P < 0.01. Sample names corresponding to the “downstream” regions are highlighted with a red background in (b–d).