Optimal Start-up And Microbial Community Evolution Of Composite Flocs-biofilm Anammox System Treating Municipal Wastewater

The single-stage of partial nitrification-anammox（SPNA）is a novel energy-saving and efficient nitrogen removal process when compared with conventional nitrification-denitrification process.Integrated fixed-biofilm activated sludge（IFAS）reactor can achieve dominant growth of nitrifying bacteria and anaerobic ammonia oxidizing bacteria（AnAOB）in flocs and biofilm,respectively,and make it possible to independently control the sludge retention time（SRT）of flocs and biofilm,which is conducive to achieving stable mainstream SPNA process.However,the SPNA-IFAS process treating municipal wastewater still has several problems that need to be solved,such as the seeding partial nitrification sludge was not common,and it is difficult to maintain stable partial nitrification during long-term operation,which lead to the deterioration of SPNA-IFAS system and limit its application.To solve the above problems,the project explored the main factors on start-up of partial nitrification system by inoculating nitrification sludge.Based on that,the optimal start-up strategy of SPNA-IFAS system treating municipal wastewater was established by inoculating nitrification sludge and anammox biofilm.In addition,the evolution and mechanism of microbial community in flocs and biofilm were investigated.Finally,the influence mechanism of influent C/N ratios fluctuation on nitrogen removal performance and the interaction of microbial community were investigated in the SPNA-IFAS reactor.The main contents and conclusions were as follows:（1）Effects of specific biomass NH₄⁺-N load（BNL）and ammonia oxidizing bacteria（AOB）activity on start-up performance of partial nitrification system.The partial-nitrification reactor fed with synthetic municipal wastewater was started up by inoculating nitrification sludge under intermittent aeration condition.Firstly,the effect of BNL on the start-up of partial nitrification system was studied by regulating the NH₄⁺-N load.The results showed that the increase of BNL（from 15.0 to 67.8 mg/（L·VSS））was conducive to the accumulation of NO₂^--N（39.9%）in the reactor.When BNL decreased to 60.1 mg/（L·VSS）,the average NO₂^--N accumulation rate（NAR）decreased from 39.9%to 30.8%,indicating an unstable accumulation of NO₂^--N.In addition,when BNL increased,the reactor with high AOB activity（41.3 mg/（L·h））achieved the accumulation of NO₂^--N（30.3%）on day 14,while the reactor with low AOB activity（14.5 mg/（L·h））reached the NAR of 18.1%on day 41,indicating that high activity of AOB could accelerate the start-up of partial nitrification.Finally,the effect of BNL on the start-up of partial nitrification was investigated by regulating SRT.The results showed that the NAR was low（11.5%）under long SRT condition,then increased to 89.6%and remained stable for 90 days when the SRT was reduced to 30 d,indicating that the increase of BNL by reducing SRT was conducive to the realization of stable partial nitrification.（2）Rapid start-up of SPNA-IFAS system through inoculating nitrification sludge and anammox biofilm:Nitrogen removal performance and the evolution of microbial communityThe SPNA-IFAS system fed with synthetic municipal wastewater started up by inoculating nitrification sludge and anammox biofilm under intermittent aeration condition.Nitrogen removal performance and the evolution of microbial community with reduced SRT were investigated.The average effluent NO₃^--N concentration was 7.2 mg/L,with total nitrogen（TN）removal efficiency of 88.5%on day 2.The results showed that the SPNA-IFAS reactor started up rapidly by inoculating nitrification sludge and anammox biofilm.Both the abundance and maximum activity of nitrite oxidizing bacteria（NOB）in flocs decreased obviously during the start-up process.The abundance of anaerobic ammonium oxidizing bacteria（AnAOB）in flocs increased from 0.213%to 0.346%despite the sludge retention time（SRT）of flocs decreased to 60 days,the AnAOB in biofilm was 0.434%.That meant AnAOB gradually enriched in flocs and accounted for a fairly high proportion.The inhibition of NOB,partial denitrification and increased aerobic＿chemoheterotrophy function in flocs might be the main reasons for AnAOB enrichment.In addition,simultaneous fermentation,partial denitrification and anammox reaction might occur in the biofilm,which further improved long-term stability of the reactor.（3）Effects of influent C/N ratios fluctuation on nitrogen removal performance and the interaction of microbial community in SPNA-IFAS systemThe effects of influent C/N ratios fluctuation（C/N=3.43,2.58,1.71,2.86 and 2.14）on the nitrogen removal performance and microbial community interaction of the SPNA-IFAS system were explored.When C/N ratio increased from 1.71 to 2.86,the average TN removal efficiency gradually increased from 79.0%to 81.9%.When C/N ratio decreased from 2.86 to 2.14,the average TN removal efficiency gradually decreased to 78.4%.The results showed that the increase of C/N ratio was beneficial to improve the nitrogen removal performance.When C/N ratio increased,the abundances of AnAOB in flocs and biofilm increased from 2.1×10⁸and 9.2×10⁸to 4.6×10⁸and 1.5×10⁹copies/L,respectively,which was consistent with the variation of nitrogen removal performance.The mechanism of the enrichment of AnAOB was analyzed by combining the variations of microbial populations with sludge particle size distribution:1)When C/N ratio increased,the activity of AOB increased from 17.2 to 19.8 mg/（L·h）,while the activity of NOB decreased from 10.5 to 9.7 mg/（L·h）.The AOB/NOB ratio gradually increased from 1.7to 2.0,which was conducive to partial nitrification-anammox reaction and the growth and enrichment of AnAOB.2)Partial denitrification-anammox reaction might occur in the reactor.When C/N ratio increased,Thauera and Candidatus＿Competibacter,which were considered the key functional bacteria involved in the partial denitrification reaction, increased from 0.450%and 3.703%to 1.959%and 7.025%in the biofilm,respectively.In addition,the abundance of DNB_nitritegene gradually dropped from 2.3×10¹¹to 1.7×10¹¹copies/L in the biofilm,which was further conducive to the partial denitrification reaction and provided more NO₂^--N for AnAOB,promoting the growth and enrichment of AnAOB.3)The predicted sequences classified as aerobic＿chemoheterotrophy gradually increased from 10.034%to 12.434%in flocs when C/N ratio increased,which can provide anoxic environment for AnAOB and facilitate AnAOB enrichment.4)The proportion of large flocs gradually increased from 30.601%to 46.705%when C/N ratio increased,which further provided more micro-anoxic environment for AnAOB and was critical for the enrichment of slow-growing AnAOB.