Effects Of Iron And Copper Ions On The Heterotrophic Nitrification And Aerobic Denitrification Of Arthrobacter Nicotianae D51

With the increasing emissions of nitrogen contaminants,biological denitrification has been widely concerned by researchers due to its high efficiency,environmentally friendly and low cost.Currently,many heterotrophic nitrification-aerobic denitrification bacteria were investigated for their ability to convert nitrogen-containing compounds into gaseous products by nitrification and denitrification under aerobic conditions.Although most heterotrophic nitrifying bacteria have the ability of aerobic denitrification,they involve different genera and have different characteristics in heterotrophic denitrification performance and nitrogen removal pathways.Therefore,it is necessary to isolate new heterotrophic denitrifying bacteria from different environment and further study their nitrogen removal characteristics to enrich the species of denitrifying microorganism and the theory of biological denitrification.In this study,a new hypothermic biological denitrification bacterium was obtained by using the methods of enrichment,isolation and purification.It was identified based on morphology and molecular biology analyses.We have investigated the removal effects of this bacterium on different nitrogen sources and the production of denitrification product N₂O.On this basis,the removal effects of this bacterium on the mixed nitrogen source were studied in order to indicating the simultaneous nitrification and denitrification of this bacterium.In addition,the denitrification characteristics of this bacterium in the presence of Cu and Fe ions were also investigated.The main findings as follows:?1?Strain D51,was isolated from long-term flooded paddy fields in GuiZhou Province,China.It was identified as Arthrobacter nicotianae,gram positive bacteria,based on morphological,phospholipid fatty acids?PLFAs?,and 16S rRNA gene sequence analyses.This is the first study to show that Arthrobacter nicotianae can perform heterotrophic nitrification and aerobic denitrification?HNAD?in wastewater,which expand the species of denitrifying microorganism.?2?Further tests demonstrated that strain D51 had the capability to use nitrite,nitrate and ammonium as the sole nitrogen source in 15?,respectively.The ammonia and total nitrogen?TN?removal rate were 95.21%and 86.48%when using ammonia nitrogen?51.59mg/L?as the sole nitrogen source,respectively.The biomass nitrogen?Bio-N?accumulation was 25.21mg/L?about 48.87%of ammonia nitrogen?,and the nitrification products?nitrate and/or nitrite?were not detected.When using nitrate?53.78mg/L?and nitrite?58.39mg/L?nitrogen as the sole nitrogen source,nitrate and nitrite nitrogen removal rate were 74.04%and 70.11%,total nitrogen removal rate were72.92%and 56.37%,biomass nitrogen accumulation were 16.52mg/L?about 30.72%of nitrate nitrogen?and 18.45 mg/L?about 31.60%of nitrite nitrogen?,respectively.?3?We have detected the net production of N₂O were 0.0271,0.0144,and0.2163mg/L when the ammonia,nitrate and nitrite nitrogen as the sole nitrogen source under the condition that strain D51 cultured at 15?while shaking at 150 rpm for 48 h,respectively.These results further confirmed that strain D51 could conduct heterotrophic nitrification-aerobic denitrification under aerobic conditions.N₂O is a greenhouse gas,but it produces little in this process and does little harm to the environment.?4?The ammonia,nitrate and total nitrogen removal rates were 98.62%,58.53%and 68.60%when the ammonia?28.21mg/L?and nitrate?29.30mg/L?nitrogen as the mixed nitrogen sources,the nitrite nitrogen was detected in this process,respectively.When ammonia?29.40mg/L?and nitrite nitrogen?28.46mg/L?were used as mixed nitrogen sources,the removal rates of ammonia,nitrite and total nitrogen were 95.88%,65.95%and 73.87%,respectively.Therefore,strain D51 can utilize ammonium and nitrate or nitrite nitrogen simultaneously,and has the function of simultaneous nitrification and denitrification.The addition of nitrate or nitrite nitrogen would not affect the removal rate of ammonia nitrogen,but the addition of ammonia nitrogen would inhibit the removal rate of nitrate and nitrite nitrogen.?5?The effects of Fe³⁺on heterotrophic nitrification was lower than that on aerobic denitrification.The removal rate of nitrate nitrogen?52.42mg/L?reach up to 100%,and the average removal rate was 2.18mg/L/h in the presence of 15mg/L Fe³⁺.When added10mg/L Fe³⁺into medium,the removal rate of nitrite nitrogen?61.17mg/L?reached to97.45%,and the average removal rate was 1.24mg/L/h.The nitrate and nitrite nitrogen removal rate increased by nearly 30%and 36%compared with the medium without Fe³⁺,respectively.?6?The cell growth and nitrification of strain D51 would be inhibited when adding Cu²⁺into medium.However,the cell growth and aerobic denitrification of strain D51were promoted when Cu²⁺?0.05mg/L?was added to the denitrification medium.The average nitrate and nitrite nitrogen removal rate were 0.96mg/L/h and 0.80mg/L/h in the presence of Cu²⁺?0.05mg/L?when nitrate and nitrite nitrogen as the sole nitrogen source,respectively.Compared with control treatment,the nitrate and nitrite nitrogen removal rate increased by about 8%and 14%,respectively.However,the denitrification of strain D51 would be inhibited when the concentrations of Cu²⁺>0.1mg/L.The above results have demonstrated that Arthrobacter nicotianae D51 had the capability to conduct heterotrophic nitrification–aerobic denitrification and simultaneous nitrification-denitrification under aerobic conditions.This bacterium could still remove ammonium nitrogen in wastewater rapidly and efficiently though ammonium and nitrate or nitrite nitrogen were used as mixed nitrogen sources.Moreover,strain D51 had the capability to use ammonium,nitrate and nitrite nitrogen in the presence of Cu²⁺or Fe³⁺.Therefore,this bacterium can be used as a candidate strain for denitrification of nitrogen pollution wastewater containing Cu²⁺and Fe³⁺in autumn and winter,which is of great significance for the protection of environmental water bodies and the regeneration and utilization of water resources in our country.