Study On Denitrification Process And Microbial Community Metabolism Mechanism Of MFC-granular Sludge Coupling System

Anaerobic biological treatment technology has been widely used in the treatment of all kinds of wastewater,and can produce energy-methane.However,after anaerobic treatment,the effluent contains dissolved methane,which is an important greenhouse gas.Therefore,it has become a hot issue to actively explore new methods of methane resource utilization in wastewater treatment.In addition,the excessive discharge of nitrogen and its treatment failed to meet the standards have caused many disturbing and ecological environmental problems.The process of denitrifying anaerobic methane oxidation（DAMO）with nitrite/nitrate as electron acceptors by oxidizing methane pathway,which is used for denitrification and utilization of dissolved methane.Hence,it can provide a new method for the treatment of nitrogen wastewater.Therefore,by constructing a microbial fuel cell（MFC）-granular sludge coupling system,the denitrification process,utilization of dissolved methane,electrical performance,extracellular polymer substance（EPS）and its microbial community in the treatment of nitrogen-containing wastewater by using sodium acetate and methane as electron donors and carbon sources respectively,were investigated in this study.Meanwhile,the effect of growth factors addition on the denitrification process and microbial metabolic mechanism of the system were analyzed.The results were as follows:（1）At different nitrogen concentrations of the influent,the chemical oxygen demand（COD）removal efficiency of the system was the highest at theⅣphase（NO₃^--N and NO₂^--N concentrations of influent were 180 mg/L and 80 mg/L,respectively）,and the removal efficiency was reached 96.0%.The effluent concentration of NO₃^--N was more stable at theⅡphase（NO₃^--N and NO₂^--N concentrations of influent were 100 mg/L and 40 mg/L,respectively）,and the removal efficiency of NO₃^--N reached 99.0%.The electricity generation performance was better in theⅢphase（NO₃^--N and NO₂^--N concentrations were 140 mg/L and 60 mg/L,respectively）,and the maximum current density was 1859.1 m A/m³.The highest value of polysaccharide and protein content of loosely bound（LB）-EPS was 13.7 mg/g and 14.7 mg/g in the 5^thchamber of theⅡphase,respectively.On the whole,the content of polysaccharide and protein was higher at theⅣphase.From theⅠto theⅣphases of the system,the relative abundance of Proteobacteria decreased,but Chloroflexi,Firmicutes and Planctomycetes all increased.The relative abundance of Methanothrix increased to 12.3%in the 4^thchamber,indicating that methanogenic bacteria could co-exist with other bacteria in the system.（2）The COD removal efficiency of this system can be maintained at a high level when the hydraulic retention time（HRT）was 24 h,16 h and 12 h,respectively,and the highest average removal efficiency was 98.4±0.8%,while it was slightly decreased at 8 h,and the average removal efficiency was 96.7±2.2%.The removal efficiency of NO₃^--N was the highest at HRT of12 h and 8 h,and could be maintained above 99.5%.The effluent concentration of NO₂^--N was lower than 0.01 mg/L in each phase of HRT.The system had the best electricity generation performance at HRT of 24 h,and the maximum power density and output voltage of the 4^thwere471.2m W/m³and 608.1m V,respectively.When HRT was reduced to 8 h,the abundance of Proteobacteria,Thauera and Asoarcus with denitrification and nitrogen fixation were increased,especially in the 1^stand 2^ndchambers,which increased by 34.5%,16.5%,and 5.9%,and 17.7%,13.9%and 8.4%,respectively.In the Kyoto Encyclopedia of Genes and Genomes（KEGG）metabolism pathway,Metabolism at the Pathway Level 1 accounted for the highest proportion（50.0%）in each chamber,the proportion of Membrane Transport at the Pathway Level 3 was the highest（13.7%）,and Methane metabolism（2.0%）and Nitrogen metabolism（0.9%）played an important role,which provided a guarantee for the stable and efficient removal of organic matter and nitrogen in this system.（3）When betaine was selected as a growth factor,at the betaine addition of 1.0 g/L,the NO₃^--N and NO₂^--N removal was increased to 210 mg/L and 150 mg/L,respectively,and the concentration of dissolved methane was up to 17.8 mg/L.Electrochemical impedance spectroscopy（EIS）curve at 1.0 g/L of betaine addition indicated the minimum transfer resistance and the highest electron transport capacity in the 1^stto 5^thchambers,which was consistent with the results of electron transport system（ETS）activity,indicating that the increase of betaine promoted electron transport.Moreover,the increase of fluorescent substance in EPS showed that the structure and flocculation of granular sludge was more stable.As for the microbial community,when betaine increased from 0 g/L to 1.0 g/L,the dominant phylum changed from Euryarchaeota（42.0%-54.1%）to Proteobacteria（20.8%-50.7%）in the 1^stto 4^thchambers.The relative abundance of Methanothrix was significantly decreased（by 17.9%-37.4%）.Nitrospirae（15.4%）,Paracoccus（17.0%）and Nitrospira（15.4%）were found in the 5^thchamber,and the abundance of DAMO microorganism showed a slight decreasing trend,which may be due to the increase of methyl donors overmuch.When the addition of betaine was 0.5 g/L,the proportion of various metabolic pathways was relatively high in the 1^stchamber.In general,when betaine was added as a growth factor,the microbial community structure showed a higher growth metabolic activity.（4）When arginine was selected as a growth factor,the addition of arginine had little effect on the improvement of nitrogen removal,and the low dissolved methane limited the use of arginine as an electron donor for NO₃^-and NO₂^-reduction by DAMO microorganism.When the addition of arginine was 0.2 m M/L,the EIS radius was the smallest and the electrochemical activity was better in the 1^stto 5^thchambers.When the addition of arginine was 1.0 m M/L,the activities of 2-p-nitrophenyl tetrazolium chloride（INT）-ETS and 2,3,5-triphenyl tetrazolium chloride（TTC）-ETS in sludge were higher,reaching 17.02 mg INTF/（g TSS·h）and 1.99 mg TF/（g TSS·h）,and the fluorescence peak intensity of coenzyme F₄₂₀was increased.The influence of arginine on the microbial community of MFC-granular sludge coupling system was analyzed,indicating that Syntrophobacter(4.2%for the 1^stchamber,10.6%for the 2^ndchamber,15.9%for the 3^rdchamber,15.7%for the 4^thchamber)and Thermononas(7.3%for the 1^stchamber,7.0%for the 2^ndchamber,3.3%for the 3^rdchamber,3.4%for the 4^thchamber)were the dominant genus in the 1^stto 4^thchambers.In the Pathway Level 1 of KEGG metabolic pathway analyzed by metagenomic data in the 1^stchamber,metabolic clusters of Metabolism,Environmental Information Process,Genetic Information Processing and Cellular Processes accounted for21.9%,7.2%,5.4%and 4.6%,respectively.Meanwhile,amino acid metabolism and protein generation and transport played an important role in the system of arginine addition.