Pollution Characteristics Of Antibiotic Resistance Genes In Livestock Waste And Farmland Soil Of Manure Application In Small And Medium-sized Livestock Farms

There are a large number of small and medium-sized livestock farms in our country,which are mainly distributed in rural areas.Livestock waste is mostly applied to farmland soil after simple composting treatment,and the breeding model also belong to extensive farming,which brings great pressure to the rural ecological environment and severely aggravated environmental pollution.Especially,the pollution of antibiotic resistance genes（ARGs）has attracted wide attention in livestock waste and farmland soil environment.Aiming at the unclear pollution problem of ARGs in small and medium-sized livestock breeding environment,and the unknown impact of livestock-derived ARGs on the farmland environment,a total of 48 typical small and medium-sized livestock farms were selected in Yuqiao reservoir in Jizhou area and Erhai Lake basin of Dali in this study,including 16 pig farms,16 cow farms and 16 chicken farms.All of 189livestock waste and soil samples were collected from these livestock farms.Meanwhile,the detection and analysis of 23 ARGs in 8 classes were carried out by combining the methods of qualitative PCR and real-time fluorescence q PCR.The purpose of this study was to find out the distribution characteristics,pollution level and influence of ARGs on the surrounding farmland environment in livestock waste of small and medium-sized livestock farms in surrounding areas of drinking water sources in sensitive basins.The main research results of this study are as follows:（1）ARGs pollution was prevalent in livestock waste in small and medium-sized livestock farms in Erhai Lake basin,Dali.The sulfonamide sul1,sul2,tetracycline tetO,tet Q,tet W,tet L,tet X,macrolides erm B,erm C,quinolone qnr S,qnr B,oqx B,streptomycin aad A,str A,str B andβ-lactam resistance genes bla_{amp C},bla _OXA-1 and bla _TEM-1 had high detection rate.Among them,the pollution level of common genes tetO,tet Q,tet W and high-risk genes bla _OXA-1,bla_{amp C} and bla _TEM-1 were relatively high,with the relative abundance ranges of 10^-3～10⁰ and 10^-4～10^-1copies/16S copies,respectively.Additional,the pollution of ARGs was different in different livestock waste.Among different livestock waste,the pollution levels of ARGs in chicken farms was the highest,with an abundance of 10^-4 to 10⁰ copies/16S copies.Meantime,the ARGs abundance in sow and layer waste were higher than that in piglets/fattening pig and broiler waste,respectively.The application of livestock manure had also a certain effect on soil ARGs.The ARGs abundance in the soil increased by 36 times,30 times,and 11 times after the application of pig,chicken,and cattle manure,respectively.Therefore,application of livestock waste could significantly increase ARGs abundance in soil environment.（2）The common sulfonamide sul1,sul2,tetracycline tetO,tet Q,tet W,tet L,tet X,macrolides erm B,erm C,streptomycin aad A,str A,str B were prevalent in livestock waste of small and medium-sized livestock farms in Yuqiao reservoir in Jizhou area,with the detection rate of 100%.Among them,the pollution level of tetracycline resistance genes tetO,tet Q,tet W and macrolide resistance gene erm B were most severe.Meantime,β-lactam resistance genes bla_{amp C},bla _OXA-1 and bla _TEM-1 and multiyear resistance genes mcr-1,which were closely related to human health,were also widely distributed in livestock waste in small and medium-sized livestock farms.The pollution levels of bla _OXA-1、bla_TEM-1and bla_{amp C} were higher,with the relative abundance of10^-5～10^-2 copies/16S copies.Additional,compared with pig waste and cattle waste,the pollution level of ARGs was more serious in chicken waste.And the relative abundance of most ARGs were significantly higher in sow waste than that waste of piglet and fattening pig in different growth stages.The ARGs abundance in the soil increased by 18 times,14 times,and 8 times after the application of chicken,pig,and cattle manure,respectively.Therefore,there was a great risk of ARGs transmission in manure returned to the field.（3）Through a comparative analysis,it was found that the pollution level of ARGs in livestock waste of small and medium-sized livestock farms in Erhai Lake basin was higher 1～1.5 orders of magnitude than that in livestock manure in livestock farms surrounding Yuqiao reservoir.Meantime,the effect of ARGs on soil of animal manure application in Erhai Lake basin was greater than that in soil of animal manure application in Yuqiao reservoir.