Determination Of The Fate Of Antibiotic Resistance Genes And The Response Mechanism Of Plants During Enhanced Antibiotic Degradation In A Bioelectrochemical-constructed Wetland System

The extensive use of antibiotics and their continuous release to the environment have promoted the proliferation of resistant bacteria(antibiotic resistant bacteria,ARB)and resistance genes(antibiotic resistance genes,ARGs),posing a potential threat to human health.Chloramphenicol(CAP)is a broad-spectrum antibiotic that has been widely used.Due to the biological toxicity of nitro and chlorine substituents,CAP is difficult to be degraded by typical biological wastewater treatment processes.Bioelectrochemical(BES)systems can decrease the biotoxicity of CAP by using biocathode to promote the reduction of chlorinated substituents and the conversion of nitro groups to ammonia substituents,which will increase its biodegradability.Simultaneously,the constructed wetland(CW)system together with the substrate,microorganism,and plant can enhance antibiotics removal.The coupling of bioelectrochemical systems and constructed wetland(BES-CW)can improve the removal efficiency of refractory organic pollutants and broadened the removal route of antibiotics.However,in the process of antibiotic removal by BES-CW,bacteria will develop drug resistance under the selection pressure of antibiotics.Therefore,it is of significance to investigate the fate of ARB and ARGs in the BES-CW.In this study,a combined BES-CW system was developed for CAP removal.The CAP removal capability and the abundance of ARGs in BES-CW were examined under different operating conditions.With the help of high-throughput sequencing results,the correlation analysis between the abundance of horizontal transfer marker genes(type Ⅰ integron,intI1)and the abundance of ARGs clarified the contribution of horizontal transfer and host bacteria to the changes in ARGs,and revealed the ARGs in BES-CW.At the same time,antibiotics will be absorbed by plants and accumulated in plant tissues.Therefore,this study finally took the model plant Arabidopsis thaliana as the research object to investigate the toxicity mechanism of CAP to Arabidopsis.The main research results are summarized as follows:(1)In this study,we investigated the effects of voltage and initial concentration of CAP on system performance.With the increase of voltage,the removal of CAP,NO3--N and TN in wastewater was enhanced.At a high initial concentration of CAP,the removal efficiency of CAP was improved,while the removal efficiency of nutrients was inhibited.In addition,CAP can accumulate in the plant,and the underground part is more than the overground part.(2)The distribution and transfer mechanism of ARGs were determined based on the correlation analysis between microbial community and ARGs.ARGs are mainly distributed at the cathode in the BES-CW system.Along the direction of influent flow,the abundance and expression of ARGs decreased.The mechanism of voltage and CAP concentration on ARGs abundance is different,which is shown as follows:The shift of ARGs potential hosts,rather than horizontal gene transfer,was the major factor for altering ARGs under different voltages.While different CAP concentrations,the horizontal transfer and the shift of ARGs potential hosts are the main reason for the change of ARGs.(3)In this study,the main root elongation of Arabidopsis thaliana was significantly inhibited after exposure to different concentrations of CAP(100 nM,300 nM,500 nM,1000 nM,2000 nM and 4000 nM).The detection of reactive oxygen species(ROS)levels in roots by staining showed that with the increase of CAP concentration,ROS in plants significantly increased.Through root morphological analysis of root tissue,it was found that CAP can shorten the cell length of meristematic and mature regions,which is consistent with the accumulation of ROS in the root.