Individual And Combined Effect Of Copper And Tetracycline On Soil Ammonia-oxidizing Process:A Mechanism Study

Antibiotics and heavy metals have been widely used as the additives in the livestock and poultry breeding industry,while which would be released to the animal wastes if their dosage exceeded the normal requirement.With the long-term addition of animal wastes as a kind of organic manure to soils,excessive accumulation of antibiotics and heavy metals has been commonly observed,likely imposing threat to the soil quality and the healthy of micro-ecological environments and human being.Due to the enormous application,copper and tetracycline were chosen as the representatives to assess their individual and combined effect on the ammonia-oxidizing process together with how the related parameters varied with the changes in this process.Meanwhile,efforts have also been made to screen the resistant bacteria for joint pollution of copper and tetracycline and reveal the underlying resistance mechanisms,with the aim to develop a new bio-remediation strategy for this emerging environment problem.The primary conclusions are as the following:(1)Two strains of resistant bacteria for joint pollution of copper and tetracycline were obtained,namely ZFS-1X and ZFS-2M,which were further identified as Acinetobacter johnsonii and Fusarium solani,respectively.Since they were proved to use tetracycline as the sole carbon source and poss tolerance to copper toxicity,we use them in the following remediation experiments after enrichment culture.(2)The distribution of copper species in soil is closely related to soil properties and its concentration.The content of oxidizable form was the highest in the soil treated with single and combined pollution,while the content of residual form was the highest in the soil treated with resistant bacteria.The reducible form was predominant in soil with low addition of copper,while the content of acid soluble form was highest in medium and highly copper contaminated soil.The percentage of acid-soluble copper increased with the increase in the addition of copper,and decreased with the prolongation of culture time.The activity of copper would be increased with the low concentration of tetracycline,and no observable effect on the content of acid-soluble copper was found in the presence of medium and high concentration of tetracycline and resistant bacteria.(3)The degradation of tetracycline in soil is related to soil properties,and exogenous bacteria will change the degradation process of tetracycline.The degradation of tetracycline in the single and combined treatment occurred mainly in the first 14 days.Copper ions had no impact on the degradation of tetracycline of low concentration,but affect degradation of medium and high concentration of tetracycline with inconsistent trend.The screened resistant bacteria can effectively promote the degradation of tetracycline and reduce the residual rate,with the order of efficiency decreasing as the following: Mixed bacterial solution >2M>1X.(4)The distribution of ammonia-oxidizing archaea(AOA)and ammonia-oxidizing bacteria(AOB)varied with the physical and chemical properties of soil,and pollution stress has inhibitory effect on ammonia-oxidizing microorganisms.The AOB is the dominate bacteria in the single and combined treatment,whereas the AOA is the dominant flora in soil treated with the exogenous bacteria.The abundance of AOA gene gradually increased and exceeded that of AOB during the process of cultivation in the single and compound treatment.This change could be observed after 56 d in the control experiments,and only 14 d and 28 d were needed in single and combined pollution treatment.The rate of AOB inhibited in corresponding pollution treatment was larger than AOA in the early and middle of the culture period,and was smaller than that of AOA in the later stage.In the exogenous bacteria treatment,the percentage of AOA gene abundance exhibited a downward trend in the process of culture,while it was always dominant.We find that AOB is more sensitive to pollutants and AOA is more tolerant,and they will complement with each other in driving ammonia oxidation.The driving effect of AOA is greater than that of AOB under environmental stress.(5)The net effect(inhibition or promotion)of single and combined pollution of copper and tetracycline on soil respiration,soil enzymes,ammonium nitrogen,nitrate nitrogen,AOA and AOB differs with their concentrations.The correlation among these indicators was different.Highly positive correlation between ammonium nitrogen and AOB were found,which can be used as the characteristic indicator of AOB.The exogenous bacteria would enhance the inhibition of the combined pollution on AOA,AOB and nitrification and promotion on ammonification and the activity of catalase.This effect was more evident in the mixed bacteria or 2M bacteria,while they had no significant effect on the activity of sucrase.The inhibitory effect of compound pollution on ammonia-oxidizing was intensified by exogenous bacteria in the early and middle stage of incubation,and the inhibitory effect of compound pollution on ammonia-oxidizing was alleviated through improving the degradation of tetracycline by exogenous bacteria in the later stage.