Study On The Composition,abundance,elimination Of Bacterial Communities And Antibiotic Resistance Genes From The Aquaculture Sediments

In recent years,with the increasing demand for aquatic products,the aquaculture industry has ushered a period of rapid development in China.The pond aquaculture has gradually become an important part of freshwater aquaculture industry,which provides a large number of aquatic products for China every year.With the rapid development of freshwater aquaculture,the environmental problems caused by aquaculture activity have also attract people’s extensive attention,especially the emergence of antibiotic resistant bacteria(ARB)and resistance genes(ARGs)has been detected in the aquaculture sediments.The enrichment and propagation of ARB and ARGs were accelerated by the pollutants and antibiotics in aquaculture ponds,which also enter the external environment with aquaculture wastewater and further cause some harm to the ecosystem.Therefore,sediments of differnte aquaculture ponds were collected to investigated the structure of bacterial community,the distribution of ARB and the c haracteristics of ARGs.these results could provide evidence for the health of aquatic products and the protection of the ecological environment.The sediment samples were collected from the aquaculture farms in Anhui,Fujian,Guangxi,Hainan and Shaanxi provinces.Subsequently,the gene cloning,real-time quantitative PCR(qPCR),16 S rRNA high-throughput sequencing,drug medium screening,metagenomics analysis and simulation methods were used to study the structure of bacterial community,the abundance and diversity of ARB and ARGs,the antibiotics resistance phenotypes,genotypes of Escherichia coli(E.coli)and the efficiency of NaClO remove the culturable bacteria and ARGs in the aquaculture environment.We also used network analysis and structural equation modeling(SEM)to analyze the co-occurrence patterns of bacterial communities(phylum and genus)and ARGs.The research results are as follows:(1)The Proteobacteria has been found the highest abundance at the aquaculture ponds from 5 provinces in China,followed by Firmicutes,Bacteroidetes,Chloroflexia and Desulfobacterota.These bacterial phyla accounted for more than 60% of the total bacterial community.The higher abundance of Campilobacterota was detected in Fujian aquaculture ponds.Similarly,a higher abundance of Halobacterota was detected in Hainan aquaculture ponds.At the genus level,Anaeromyxobacter and Geothermobacter were the dominant genera in Anhui ponds;Pseudomonas,Sulfurimonas,and Sulfurovum were the dominant genera in Fujian ponds;Robiginitalea and Candidatus were the dominant genera in Guangxi ponds;Methanobacteria,Methanoregula and Leptolinea were the dominant genera in Hainan ponds;Thiobacillus,Sphingomonas and Erysipelothrix are the dominant genera in Shaanxi ponds.(2)We found 27 potential pathogenic bacteria with a higher abundance in the sediments from 4 aquaculture farms of Shaanxi Province.Among them,Salmonella,Larenella,Enterococcus,Klebsiella and Aeromonas have the highest relative abundance in Shangluo farms;Mycobacterium,Acinetobacter,Toxobacter,Pseudomonas and Haemophilus have the highest relative abundance in Ankang farms;Bdellovibrio,Campylobacter,Listeria,Eubacter and Staphylococcus have the highest relative abundance in Heyang farms;the relative abundance of all kinds of pathogenic bacteria in Xian farms was lower than other farms.(3)A total of 422 ARGs were obtained from sediment of 4 farms ponds in Shaanxi Province by the method of metagenomics analysis,which includes aminoglycosides,betalactams,macrolides,multidrug,sulfonamides,fluoroquinilone,rifamycin,tetracyclines,MLS and polypeptide gene,most of those belong to the efferent pump mechanism and antibiotic inactivation mechanism.Among them,β-lactams resistance genes had the highest relative abundance,which included 120 ARGs.Multidrug resistance genes and Aminoglycoside resistance genes had the higher relative abundance,which has 59 and 55 ARGs,respectively.The number of Rifamycin resistance genes were the least,which only have 7 ARGs.At these ARGs,TEM-116 has the highest abundance,which was concentrated in the Shangluo farms.Following by ade F,chr B and Tri A are concentrated in Heyang farms,and the Tri A is a disinfectant resistant gene.SEM analysis indicated that the richness and diversity of these ARGs affected the contents of TN and TP and the diversity of bacterial community in the environment.Meanwhile,Salmonella,Acinetobacter,Aeromonas and Bdelvibrio have positively correlated with the antibiotic resistance genes of β-lactams,MLS,quinolones and sulfonamides.Enterococcus and Streptococcus have positively correlated with the antibiotic resistance genes of rifamycin.(4)63 E.coli whose isolated from these sediments has been shown to resist multiple antibiotic.Among them,23 strains showed resistance to DOX + CIP + AMP;19 strains showed resistance to OXY + DOX + AMP;17 strains showed resistance to OXY + DOX + CIP;4 strains showed resistance to OXY + DOX + EFT.At the 63 strains,89% of the strains carried 8 or more ARGs,and 36 strains carried the Intl1 genes,9 strains carried the Intl2 genes and 21 strains carried the ISCR1.(5)The NaClO whose containing 3mg/L effective chlorine could effectively remove the culturable bacteria in aquaculture water;the 9 mg/L effective chlorine could remove part of plasmid ARGs in aquaculture water.In conclusion,aquaculture sediments not only become a reservoir for ARGs,but also become a reservoir for some potential pathogens.The pollution status of ARGs in aquaculture farms was caused by long-term effects.Therefore,we suggest that reasonable use of NaClO could effectively reduce the risk of pathogen and ARGs transmission to the environment.