Effects Of Free Ammonia On Phosphorus Removal Performance Of Polyphosphate Accumulating Organisms And Microbial Population Structure In EBPR System

In order to explore the influence of free ammonia（FA）on phosphorus removal performance and microbial population structure of polyphosphate accumulating organisms（PAOs）in the EBPR system.In this experiment,a Sequencing Batch Reactor（SBR）was used,with activated sludge rich in PAOs as the research object,and the influence of different FA concentrations on the performance of the EBPR system was investigated.The non-matrix inhibition kinetic model is used to perform nonlinear fitting to the experimental data.Based on16S rRNA high-throughput sequencing technology,in-depth exploration of the diversity of activated sludge microorganisms under different FA concentrations.Based on the results of batch experiments and microbial diversity analysis of different FA concentration inhibition,using correlation analysis and structural equation models,the correlation between FA,kinetics,and functional bacteria was established and statistically analyzed.It aims to provide theoretical support for optimizing biological phosphorus removal technology,so as to obtain an efficient and stable wastewater biological phosphorus removal method.Based on the test of the effect of different FA on the performance of the EBPR system,it was found that 18 various FA concentrations were obtained by controlling the NH₄⁺-N,p H value and temperature of the influent.With the increase of FA concentration gradient,the degradation rate of volatile fatty acids（VFA）decreased linearly,and at 50 mg FA·L^-1,it decreased to 6.12mg COD·g VSS^-1·h^-1.0.2 mg FA·L^-1 is the inflection point of phosphorus removal performance.When FA<0.2 mg FA·L^-1,FA promotes the activity of PAOs;when FA>0.2 mg FA·L^-1,FA has an effect on PAOs activity Inhibition.In the promotion process,FA can significantly promote anaerobic phosphorus release than aerobic phosphorus absorption.When the FA concentration is higher than 0.6 mg FA·L^-1,the inhibitory effect of FA on aerobic phosphorus absorption is stronger than that of anaerobic phosphorus release.Based on the non-matrix inhibition kinetic model,the experimental data is nonlinearly fitted.The results show that the Hellinga model can better describe the effects of FA on anaerobic phosphorus release(R²=0.90,F=882.9,P=2.0×10^-15)and aerobic phosphorus absorption(R²=0.96,F=998.2,P=7.8×10^-16),and the inhibition threshold of FA on the process of anaerobic phosphorus release is significantly higher than that of aerobic phosphorus absorption,and the inhibition constant（K_I）of anaerobic phosphorus release is 3.57～3.65 times higher than that of aerobic phosphorus absorption.Based on 16S rRNA high-throughput sequencing technology,we investigated 8 various FA concentrations(0 mg·L^-1,0.2 mg·L^-1,1.0 mg·L^-1,4.0 mg·L^-1,10 mg·L^-1,15 mg·L^-1,25mg·L^-1and 50 mg·L^-1)system community structure changes,further explore their differences.Sequencing results show that Good’s coverage index covers a range of 97.0%～98.6%,and the sequencing depth is sufficient to cover most microorganisms.Proteobacteria is the main dominant species at the phylum level.With the increase of FA concentration,its proportion in the sample rose from 31.68%to 63.29%,which has certain adaptability to the presence of FA in the system.Gammaproteobacteria gradually rises to 59.85%with the increase of FA concentration,and the relative abundance becomes stable at high FA concentration.According to the clustering results of species at the genus level,Ca.Accumulibacter and Ca.Competibacter are in a cluster,and the relative abundance difference of the flora is small,but the relative abundance change trend of Tetrasphaera is more in line with the influence of FA on the phosphorus removal performance.Based on the LEf Se analysis,statistically significant dominant species of samples with low,medium and high concentrations are Chloroflexi,Bacteroidetes,and Betaproteobacteriales.Based on the principle of statistical analysis,using the results of batch experiments inhibited by various FA concentrations and microbial diversity sequencing,the correlation analysis of FA concentration,kinetics and system community structure was carried out to explore the comparative rate of different FA concentrations and the relative abundance of functional bacteria degree of causality.The one-sample Kolmogorov-Sminov test based on the non-parametric test shows that the commensurate phosphorus rate(r _PO4,AN),the specific phosphorus uptake rate(r _PO4,Ox)and the progressive significance of Ca.Competibacter are greater than 0.05,in line with normal distribution,can be used for Pearson correlation analysis.The analysis results show that r _PO4,AN and r _PO4,Ox are highly positively correlated,and Ca.Competibacter is highly negatively correlated with r _PO4,AN,r _PO4,Ox,which is consistent with the fact that Ca.Competibacter has no phosphorus removal ability.Based on the Spearman correlation analysis,FA is highly negatively correlated with r _PO4,AN,r _PO4,Ox and Tetrasphaera,respectively,and positively correlated with Ca.Accumulibacter and Ca.Competibacter.Establish a structural equation model（SEM）for path analysis.Tetrasphaera has a strong positive drive for r _PO4,AN（path coeff.=0.17,P=0.11）and r _PO4,Ox（path coeff.=0.72,P<0.001）.The model explained that Tetrasphaera played a key role in the effect of different FA concentrations on the phosphorus removal performance of the EBPR system.