摘要

      细菌抗生素耐药基因（ARGs）是一种新兴的环境污染物，通过病原菌对人体健康构成极大威胁。高通量定量PCR（HT qPCR）和宏基因组技术是目前应用于水环境ARGs监测的两种常用手段。然而，目前对不同的ARG剖面分析方法认识不足，影响了对水环境中ARG的认识，极大地限制了这两种方法的进一步应用。为了填补这一知识空白，本研究同时采用这两种方法对淡水水库中的ARGs进行时空对比研究。我们发现亚基因组方法检测到更多的ARG亚型，比HT-qPCR检测到更多的杆菌肽抗性基因。总的来说，HT qPCR和宏基因组分析都揭示了ARGs与细菌类群之间相似的ARG动态模式和共生模式，以及ARGs与环境因子之间的关系。我们的结果表明，不同的ARG分析方法对理解细菌ARGs的影响可能很小或可以忽略不计。HT-qPCR方法具有省时、绝对定量、对生物信息学技能要求低的优点，但与宏基因组方法相比，存在PCR扩增和引物偏向性高、引物依赖性高、ARG亚型定量能力相对较低等缺点。我们认为HT-qPCR方法可用于常规的水环境监测，而宏基因组方法可用于综合调查，以获得更多的ARG亚型信息。我们的数据可以为选择合适的ARG剖面方法用于水环境中细菌ARGs的监测和风险评估提供有用的参考。

       Bacterial antibiotic resistance genes (ARGs), a kind of emerging environmental pollutants, greatly threat human health through pathogenic bacteria. High-throughput quantitative PCR (HT-qPCR) and metagenomic approaches are two popular tools applied in aquatic environmental ARGs monitoring. However, current poor knowledge of different ARG profiling approaches' impacts on the understanding of the ARGs in aquatic environments greatly limit the further field application of these two approaches. For filling such knowledge gap, this study simultaneously employed these two approaches to examine and compare the ARGs in a freshwater reservoir across space and time. We found metagenomic approach detected more ARG subtypes and much higher bacitracin resistance genes' abundances than HT-qPCR. In general, HT-qPCR and metagenomics analyses both revealed similar ARG dynamic patterns and co-occurrence patterns between ARGs and bacterial taxa as well as the relationships between ARGs and environmental factors. Our results indicated the impacts of different ARG profiling approaches on the understanding of bacterial ARGs might be minor or negligible. HT-qPCR approach has the superiorities of time-saving, absolute quantification, low requirement for bioinformatics skills but also has some drawbacks including higher PCR amplification & primer bias, higher primer dependency and relative lower ARG subtype quantification capability compared to metagenomic approach. We suggest HT-qPCR approach can be employed for routine aquatic environmental monitoring, and metagenomic approach could be applied in comprehensive surveys for getting more ARG subtype information. Our data can be a useful reference for choosing right ARG profiling approaches for bacterial ARGs monitoring and risk assessment in aquatic environments.

      https://www.sciencedirect.com/science/article/pii/S0048969719335107?via%3Dihub