Research On The Prevalence And Distribution Characteristics Of Antibiotic Resistance Genes In Pig Farms And Their Surroundings

The pollution of antibiotic resistance genes(ARGs)has been identified as a hot issue of global concern.Acquiring ARGs from Antibiotic Resistance Bacteria(ARB)will spread with the host bacteria in the environment.Despite intensive animal husbandry is believed to a major contributor to the increased environmental burden of ARGs,little information is available regarding the corresponding ARGs associated with animal farms,which has attracted attention.Therefore,high-throughput real-time PCR was used to investigate the prevalence and distribution of ARGs in some pig farms and their surrounding environments in many provinces in China.Metagenome approach to investigate the effects of anaerobic fermentation on ARGs and microbial communities in farm wastewater.This study provides basic data for mastering the prevalence and distribution of ARGs in pig farms in China.In this study,a total of 189 samples of pig manure,soil and sewage from pig farms in five provinces of Fujian,Hainan,Jiangsu,Jiangxi and Zhejiang were collected.The high-throughput real-time PCR technique was used to compare the relative abundance and diversity detection of 285 ARGs in 16 categories.The results showed that a variety of ARGs were detected,and the profiles of ARGs were similar,among them the relative abundance of aminoglycoside ARGs,macrolide-lincomycin-streptozotocin B(MLSB)ARGs and tetracycline ARGs were detected at a higher rate,about 59.26%.The relative abundance of ARGs in pig manure was the highest,followed by sewage and finally soil.In this study,samples of Fujian pig farms were further analyzed by metagenomic high throughput sequencing.14 drug resistance genes were detected,among which tetracycline resistance genes,macrolide resistance genes and aminoglycoside resistance genes were higher detected in all samples,which consistents with the datas detected by High-throughput real-time PCR technique.Sulfonamide resistance genes,phenylpropanol resistance genes,trimethoprim resistance genes and?-lactam resistance genes are only high in soil and sewage.In addition,after the anaerobic treatment of the three pig farms sewage,the relative abundance of the total resistance genes was greatly reduced.The relative abundance of each type of drug resistance gene in the effluent was lower than that of the influent.It can be infered that sewage anaerobic fermentation treatment has a certain removal effect on drug resistance genes.Howere,the relative abundance of drug resistance genes varies from farm to farm.After oxygen fermentation erm(42),tet(M),tet(O),cfx A,bla _VEB,aph(2')-Ic,tet(Y)not only not be removed,but increased.The bacterial community structure of the influent before anaerobic fermentation treatment and effluent after anaerobic fermentation treatment were slightly different,and Helicobacteraceae,Saccharospirillaceae and Veillonellales increased,which infered that some bacteria added after anaerobic fermentation may be potential hosts for increased abundance of drug resistance genes after anaerobic fermentation.Analyze the intrinsic relationship between bacterial community and drug resistance genes,and select 7 dominant bacteria groups,of which Escherichia coli is the main dominant flora.E.coli has the best correlation with colistin resistance gene,fosfomycin resistance gene,nitroimidazole resistance gene,quinolone resistance gene and rifampic resistance gene,so E.coli may be the host bacteria of these classes of antibiotic resistance genes.It was also that found Eubacterium limosum may be the host of the multidrug resistance gene cfr(C),which needs to be verified by subsequent experiments.In this study,high-throughput real-time PCR and metagenomic sequencing techniques were used to find that various drug-resistance genes were contaminated in the farms and surrounding environment,especially the relative abundance of aminoglycosides,tetracyclines and MLSB resistance genes were the highest.After further analysis of the structural diversity and abundance of drug-resistant genes and bacteria in sewage samples before and after anaerobic treatment by used metagenomic sequencing technology,it was found that the relative abundance of total drug-resistant genes decreased after anaerobic fermentation treatment,but some drug-resistant genes showed an increasing trend.Further investigation into the internal relationship between the composition and structure of bacteria and ARGs revealed that this may be caused by the increase of bacterial abundance caused by the structural changes of bacteria after anaerobic fermentation.This study provides basic data for ecological risk assessment of drug resistance in livestock and poultry breeding environment,and provides methodological reference for epidemic distribution monitoring of drug resistance genes in breeding environment and tracing of important drug resistance genes.