Distribution Of Antibiotic Resistant Bacteria And Horizontal Transfer Characteristics Of Antibiotic Resistance Genes In Urban Landscape Waters

Urban landscape water bodies are closely related to residents,they are vulnerable to be contaminated and became a medium for the spread of antibiotic resistant bacteria,then pose a huge threat to human health.Although it has been recognized that the environment plays a key role in the development of bacterial resistance,the research on antibiotic resistant bacteria in the environment is still insufficient,especially the characteristics of antibiotic resistant bacteria in urban water environment in close contact with human beings,are urgent to be clarified.In this study,the main landscape water bodies in Xi’an were selected,the resistance of the total heterotrophic flora to different types of antibiotics and the population composition of the main resistant bacteria were investigated.The horizontal transfer characteristics of plasmid-mediated resistance genes and the effect of UV disinfection on it were studied in depth to reveal the distribution of antibiotic resistant bacteria and the horizontal transfer of major antibiotic resistant genes in urban landscape waters,and to provide a scientific basis for the propagation control of drug-resistant bacteria in urban landscape waters.In this study,six landscape water bodies（XQ,FQ,HC,TH,SY and LH）with different supply sources in Xi’an were selected.Water samples were collected monthly from April to December 2018.Resistant plate culture was used to investigate the content of total heterotrophic flora and its resistance rate to sulfamethoxazole（SMZ）,tetracycline（TET）,ciprofloxacin（CIP）,ampicillin（AMP）and cefotaxime（CTX）.The diversity of CTX resistant bacteria was studied by high-throughput sequencing.A strain of CTX resistant strain L4 isolated from LH was used as a donor strain,and Escherichia coli NK5449 was used as a recipient strain to study the change of the conjugative gene transfer frequency after different doses of ultraviolet radiation.The related antibiotic resistance genes and type I integron integrase genes on donor bacteria and conjugon were determined by PCR to confirm the horizontal transfer of antibiotic resistance genes.The results showed that the total heterotrophic flora content of the six landscape water bodies was 4.50×10³².29×10⁶ CFU/100mL;the resistance rates to SMZ,AMP and CTX were 4.99%²2.12%,5.05%¹1.37%and 4.85%¹7.64%,respectively,the resistance rate to TET and CIP were only 0.24%⁰.59%and 0.46%⁰.90%,respectively.Therefore,resistance to SMZ,AMP and CTX were the main Types of antibiotic resistance of landscape water bacteria in Xi’an.It was found that there was no significant difference in the content of five resistant bacteria in SY and LH,HC and TH（p>0.05）.The average content of CTX resistant bacteria in XQ using surface water hydration was 1.39×10⁴ CFU/100mL,while the average content of CTX resistant bacteria in FQ Lake using reclaimed water was only 3.47×10³ CFU/100mL,there is a significant difference between the two waters（p<0.05）.Based on the OTUs annotation results of high-throughput sequencing,the CTX resistant strains of different water bodies belong to 6 phylums,12 classes,21 families,and 21 genuses.Except for the CTX resistant bacteria in the HC,mainly the Bacteroides,the CTX resistant bacteria in the other five landscape waters are dominated by the Proteobacteria and the thick-walled bacteria.The CTX resistant bacteria in the HC are mainly Bacteroides,and the CTX resistant bacteria in the other five landscape waters are mainly Proteobacteria and Thick-walled bacteria.The types and abundances of CTX resistant bacteria in different water bodies are very different at the genus level.Compared with water bodies that use reclaimed water to replenish water,there are more CTX resistant bacteria species at the genus level in landscape waters that use surface water to replenish water.When the UV irradiation dose is 0,20,40 and 80 mJ/cm²,the conjugative gene transfer frequency of donor bacteria L4 and NK5449 are 3.63×10^-3,2.08×10^-4,1.17×10^-55 and 5.8×10^-6,respectively,indicating that UV disinfection can control the transfer of antibiotic resistance genes.After PCR detection,the plasmids of donor bacteria and conjugon were found to contain TEM,CTX-M and intI1 genes,indicating that the cefotaxime resistance of the donor bacteria was transferred into the recipient bacteria by conjugate transfer of plasmid.And classⅠintegrons may play an important role in this process.