| China has a vast area of the sea and wide coastal beach,and is the world's largest mariculture country.In recent years,with the expansion of the scale of aquaculture in China,aquaculture has caused tremendous pressure on the environment and produced a series of environmental problems.,Antibiotics are the main drug to prevent disease in aquaculture,however long term repeated use of an antibiotic will not only reduce the efficacy of antibiotics and it is possible to induce antibiotic resistance genes(ARGs)in animals,which it is not effective to treat diseases caused by pathogenic bacteria.ARGs can migrate and transform in soil,water,groundwater and other environmental media,eventually passed into animal and the human body through the food chain.Once the human pathogens have acquired such ARGs,it may be a threat to human health.The pollution of antibiotics and ARGs has become a global environmental problem,and the potential ecological risks caused by it have been paid more and more attention by governments and researchers.In our country,because the use of antibiotics has not been good supervision,the antibiotic concentration is often too high during the use of antibiotics to resulting high residual concenteation in the detection of sediments along the coast of China,which leads to the emergence,spread and aggregation of antibiotic resistance gene(ARGs).Because the existence of antibiotic resistance is very serious in China's marine aquaculture environment,it is necessary to carry out the detection of antibiotic resistance in intensive aquaculture areas along the coast of china.In this experiment,sediment samples were collected from 11 typical water areas along the coast of China,sampling locations including Dalian,Tangshan,Penglai,Lianyungang,Qidong,Xiangshan,Ningde,Zhanjiang,Lianshui,Dongshan,Meiji reef.First of all,it selected two sulfonamides(sulfadiazine and sulfamethoxazole),two tetracyclines(tetracycline and oxytetracycline)and two quinolones(ciprofloxacin and enrofloxacin)as the object of study after consulting the data,it was carried out which is quantitative analysis of antibiotics in sediment samples at each sampling site by solid phase extraction(SPE)and HPLC-MS/MS(High-performance liquid chromatography–tandem mass spectrometry).Then,the samples were sequenced by Illumina MiSeq,the experimental data were processed by using Qiime software and mothur software and the bacterial community in the samples was analyzed by abundance analysis,Alpha analysis and community structure analysis.Next,a single colony was isolated and selected,and part of the mixture was sequenced by Illumina MiSeq,antibiotic resistance bacteria at each sampling point was analyzed by abundance analysis,Alpha analysis and community structure analysis.Another part needed to be sequenced by 16 S rDNA,the samples were compared by extraction of total DNA and by fluorescence quantitative PCR and Illumina HiSeq2000 sequencing,through the mothur software and R software,the antibiotic resistance genes in each sampling point were quantitatively analyzed,and the detection rate and abundance of ARGs species were analyzed.Finally,by comparing the situation of antibiotic contamination in the sediments of different cultivation areas,diversity of bacterial community in sediment and antibiotics and comparing the diversity of antibiotic resistant bacteria and their resistance genes under different antibiotics in different cultivation areas to master the pollution status of antibiotics and resistance genes in the intensive aquaculture areas along the eastern coast of China and to provide a reference for the follow-up study of coastal aquaculture in eastern China.The results of this study were as follows:(1)Antibiotic contamination was present at each experimental site,the contamination of tetracyclines and sulfonamides was more serious than that of quinolones.The data shows the highest concentration of tetracycline hydrochloride in PL(Penglai)area was 1230.49 ?g/kg,the concentration of sulfadiazine in PL(Penglai)area was 714.56 ?g/kg and the concentration of sulfamethoxazole in TS(Tangshan)area was 395.25 ?g/kg;(2)There are some differences in the richness of bacterial communities in different areas,however Firmicutes and Proteobacteria are the main bacteria in the sediment of the typical coastal waters of Eastern china,bacillus has obvious advantages in all bacterial groups;(3)In most of the breeding waters,the resistance to tetracycline and sulfonamide bacteria in the community,Firmicutes occupy an absolute advantage,in the resistant bacteria community,Proteobacteria has the absolute advantage.In addition,the diversity of antibiotic resistant bacteria in the Meiji reef is the lowest in all test points;(4)Antibiotic resistance genes(ARGs)are ubiquitous in the coastal waters of China,the results of sul1 and sul2 in PL(the result of sul1/16 S rDNA is 6.87×10-2,the result of sul2/16 S rDNA is 8.99×10-2)are the highest.tet B gene abundance was highest(1.57×10-2)in TS area.In addition,the proportion of baca are large in all ARGs,DL is the highest(87.77%),QD is the lowest(30.81%).Through the above experimental results,it hopes that it can provide a reference basis for environmental protection and sustainable development of coastal aquaculture area in China's eastern in the future.Under the premise of ensuring the production of aquaculture,but also pay attention to environmental protection,so as to obtain a win-win situation. |
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