Nitrate input enriched the antibiotic resistance genes in lake sediments by shaping co-host community and promoting horizontal gene transfer

The impact of various non-antibiotic factors on antibiotic resistance has garnered widespread attention. However, there has been little investigation into whether the coexistence of nutrients with antibiotic resistance genes (ARGs) in aquatic ecosystems contributes to the increasing abundance of ARGs. We employed a microcosm experiment and metagenomic analysis to investigate the impact of nitrate on ARG profiles in lake sediments. Our results revealed that increased nitrate input correspondingly elevated the abundance of sediment ARGs, virulence factor genes (VFGs), mobile genetic elements (MGEs), and nitrate reduction genes (NRGs). Among the metagenome-assembled genomes (MAGs) harboring ARGs found by binning analysis, nitrate inputs increased the abundance of 78.4 % ARG-carried MAGs, especially in genera Nitrosomonas and Sulfuriomonas. All MAGs carrying ARGs simultaneously encoded NRGs, suggesting that ARG-NRG co-hosts are important factors for ARG proliferation. Co-localization and Pearson's correlation analyses suggested that nitrate input most likely accelerated the acquisition of ARGs by particular bacterial taxa via horizontal gene transfer (HGT). Genes involved in HGT, including those related to reactive oxygen species production, membrane permeability, ATP synthesis, and pili synthesis, were also upregulated by nitrate input, thus potentially enhancing ARG transfer. Based on the partial least squares path modeling analysis, abundances of genes involved in HGT (r = 0.43) and ARG-NRG co-hosts (r = 0.41) had the highest direct positive impact on the ARG abundance. Our study suggests the increased nitrate levels may drive the dissemination of antibiotic resistance, consequently affecting human health.