Effects And Mechanism Of Livestock Manure Returning On Antibiotic Resistance Genes In Soil

Treatment of lagoonal fecal water and returning it to field after multi-stage oxidation pond processing are widely used in large scale farming,especially dairy cattle breeding.In the above process,antibiotic resistance gene（ARGs）is a new type of environmental hazard,which will be introduced into the soil environment with lagoonal manure and increase the risk of soil resistance.In this study,the effluent from a three-stage oxidation pond in a large-scale dairy farm was used as liquid fertilizer,and four application rates of 0,10.3,21.5 and 43.1 t/acre were set.Two maize planting cycles were continuously applied.High-throughput sequencing,fluorescence quantitative PCR and multivariate analysis were used to explore the effects of different application amounts of manure in oxidation pond on the occurrence characteristics of ARGs and MGEs in soil,the varieties of bacterial and fungal community composition and environmental characteristic,and the main influencing factors and driving mechanism of ARGs abundance change were analyzed.The study found that:（1）When the manure application rate was 43.1 t/acre,the total relative abundances of ARGs and MGEs were high,which were 0.310 and 0.874 copies/16 S r RNA copies,respectively.The total relative abundance of ARGs in the background soil was low（0.033 copies/16 S r RNA copies）.With the increase of manure application rate,the total relative abundance of ARGs in the treatment group increased significantly,and the total relative abundance of MGEs also showed an upward trend.Among the MGEs in each group,the relative abundance of int I2 was high,and that of Tn916/1545 was low.The total relative abundance of MGEs in each group was higher than that of ARGs.There were significant differences in the total relative abundance of ARGs（P < 0.05）and MGEs（P < 0.01）under different application rates.In each group,MGEs were significantly correlated with ARGs,soil physical and chemical factors and ARGs/MGEs.Int I1 and int I2 were significantly correlated with a variety of ARGs,which could lead to common horizontal transfer of different ARGs.Soil moisture content and total nitrogen were widely correlated with ARGs and MGEs,but p H was not correlated with ARGs and MGEs.（2）The effect of different application amount of dairy manure on microbial community structure was significant.Under the treatment of 21.5 t/acre,Chao1 and Shannon index of soil bacteria and fungi were higher,and species diversity was better.Actinobacteria,Proteobacteria and Chloroflexi were the main phyla in the bacterial community.When the application rate was 43.1 t/acre,Proteobacteria replaced Actinobacteria to become a relatively high phyla in the bacterial community.Ascomycota and Basidiomycota were the main fungi in the soil.The application of fecal water can promote the abundance of most microbial genera.（3）The host bacterial network analysis showed that the potential host species of ARGs and MGEs were different under different treatment conditions.In different treatments,ARGs and MGEs may be transferred in different host bacteria,and the abundance of potential host microorganisms can cause the corresponding ARGs/MGEs abundance change.In ARGs and MGEs,there were more types of potential host bacteria corresponding to erm B and erm F,which were 10 and 9,respectively.There were more types of potential host fungi corresponding to sul2,erm F,tet C,int I1 and int I2（10 in each）,and bacteria were more likely to become potential hosts than fungi.（4）Redundant analysis showed that soil available nitrogen and water content had high explanatory rates for the abundance change of potential host bacteria（interpretation rates were 10.7% and 10.0%,P < 0.01）.Variance decomposition analysis showed that the synergistic interaction of microbial community structure,MGEs and soil environment played a leading role in shaping the relative abundance of ARGs in soil,and the explanation rate was 46.4%.The explanation rate of microbial community in single factor was higher（20.9%）,followed by MGEs（16.0%）and soil physical and chemical indicators（8.9 %）.Soil physical and chemical indicators can indirectly affect the abundance of ARGs by changing the microbial community structure,and microbial communities can also use MGEs to indirectly affect ARGs.