Anthropogenic impacts on antibiotic resistance genes and their hosts from pristine to urban river using metagenomic and binning approaches

Highlights

• Anthropogenic pollution aided the emergence and spread of certain ARGs.

• Anthropogenic pollution increased the co-occurrence of multiple ARGs.

• Methanothrix soehngenii affiliated to archaea carrying ARGs and ICEs was retrieved.

• MGEs were the most predominant factor in shaping ARG profiles.

• Antibiotic resistance could be enriched through co-transfer with MRGs.

Abstract

Driven by anthropogenic pressure, Antibiotic resistance genes (ARGs) could transfer from the environmental resistome into human commensals or even pathogens. The transport of ARGs through aquatic ecosystems is crucial and has attracted attention. Here, we employed metagenomic and binning to compare ARGs profiles, their co-occurrence with metal resistance genes (MRGs) and mobile genetic elements (MGEs), and their hosts between pristine and anthropogenic influenced rivers and explore the ecological mechanisms underlying the dissemination of ARGs induced by anthropogenic activities. The significantly increased relative abundance of macrolide-lincosamide-streptogramins, vancomycin, β-lactam and sulfonamide resistance genes along the environmental gradient from pristine to polluted sediments implied that anthropogenic impact aided the emergence and dissemination of certain ARGs. At the lower reach of the Ba River, the higher ratios for contigs carrying more than one ARG suggested that anthropogenic pollution favored the co-occurrence of multiple ARGs. Anthropogenic pressures also increased the relative abundance of advantaged hosts, including Chloroflexi, Firmicutes and Euryarchaeota. At the lower reach of Ba River, Romboutsia timonensis carrying multiple ARGs and ICEs were successfully recovered, posing a serious threat to human health by affecting the metabolism of gut microbiomes. And Methanothrix soehngenii affiliated to archaea carrying multiple ARGs, MRGs and ICEs were also recovered from the lower Ba River. The partial least squares path modeling revealed that MGEs were the most predominant factors inducing the ARG profiles, and the antibiotic resistance could be enriched by co-transfer with MRGs. Furthermore, environmental factors could impact the ARG profiles indirectly by first influencing the ARGs’ hosts.

Introduction

The overuse of antibiotics in both clinical and non-clinical settings has increased selective pressure and led to increasing the emergence, spread and accumulation of antibiotic resistance genes (ARGs) and antibiotic resistance bacteria (ARB) in various environmental media, including wastewater treatment plants (WWTPs), livestock farms, aquaculture and receiving aquatic environments, which pose a potential threat to public and ecological health (Shi et al., 2021; Yang et al., 2021; Ye et al., 2021; Yu et al., 2020). Antibiotic resistance is further exacerbated through horizontal gene transfer (HGT) mediated by mobile genetic elements (MGEs) and vertical gene transfer by microbial proliferation (Li et al., 2019). Moreover, the co-selection of ARGs with metal resistance genes (MRGs) could contribute to the maintenance and proliferation of ARGs under anthropogenic levels of metal pollution in the environment (Seiler and Berendonk, 2012). In particular, the transportation of ARGs via aquatic environments is significant and has received great attention, among which sediments are an important reservoir of some ARGs (Qiao et al., 2018). Urban rivers, as the major water source and environmental carrier, are suffering from serious pollution from anthropogenic sources, including sewage from WWTPs and residents (Laffite et al., 2020; Schmeller et al., 2018). More importantly, these ARGs and ARB in the river system could return to aquatic animals and even humans via the food chain or recreation, which enhances the likelihood of global dissemination of ARGs.

ARGs are inherent characteristics of microorganisms in various natural environments where there is no apparent human activity impact, such as deep permafrost, deep ocean sediments and the terrestrial deep subsurface (Brown and Balkwill, 2009; Chen et al., 2013; Zhang et al., 2018). Whereas antibiotic resistance has been detected at high levels in human-impacted environments as a result of increasing selection pressure from contaminants such as antibiotics, heavy metals and other environmental factors (Chen et al., 2019). Recent studies have looked into the occurrences and distributions of ARGs in river environments, but the role of human activities in the prevalence and spread of ARGs, as well as the ecological mechanisms underlying them, has not been thoroughly investigated (Laffite et al., 2020; Na et al., 2021). Although some studies have used metagenomic or genomic to investigate the co-occurrence between ARGs and MGEs, MRGs or microbial communities, comprehensive information on the genetic linkage between ARGs and MRGs and their co-existence with MGEs and the accurate host bacteria in the river environment is limited (Hu et al., 2020; Li et al., 2017; Yang et al., 2019).

The development of metagenomic and binning technologies overcame the drawbacks of traditionally culture-dependent and amplification-based approaches, which provided more comprehensive insights into the profiles of ARGs in complex microbial communities and reconstructed genomes of ARG hosts in the environment (Liu et al., 2019; Van Goethem et al., 2018). In this study, we chose both the primitive and urban section of Ba Rivers, which showed an environmental gradient due to different degrees of anthropogenic contamination. And environmental factors including antibiotics, heavy metals, and nutrients were considered in sediments. The metagenomic assembly and binning approaches were adopted to decipher the profiles of ARGs, MGEs and MRGs, their genetic location on the plasmid, as well as the hosts of ARGs in sediments along the environmental gradient. The objectives were to (1) reveal the effects of anthropogenic inputs on the characteristics of ARGs, MRGs and MGEs; (2) explore the molecular mechanisms involved in the mobility and dissemination of ARGs by identifying their genetic location patterns; and (3) identify the influential factors on ARG profiles of sediments and the key driver.