Temporal characteristics of algae-denitrifying bacteria co-occurrence patterns and denitrifier assembly in epiphytic biofilms on submerged macrophytes in Caohai Lake,SW China

Denitrifying bacteria in epiphytic biofilms play a crucial role in nitrogen cycle in aquatic habitats.However,little is known about the connection between algae and denitrifying bacteria and their assembly processes in epiphytic biofilms.Epiphytic biofilms were collected from submerged macrophytes(Patamogeton lucens and Najas marina L.)in the Caohai Lake,Guizhou,SW China,from July to November 2020 to:(1)investigate the impact of abiotic and biotic variables on denitrifying bacterial communities;(2)investigate the temporal variation of the algae-denitrifying bacteria co-occurrence networks;and(3)determine the contribution of deterministic and stochastic processes to the formation of denitrifying bacterial communities.Abiotic and biotic factors influenced the variation in the denitrifying bacterial community,as shown in the Mantel test.The co-occurrence network analysis unveiled intricate interactions among algae to denitrifying bacteria.Denitrifying bacterial community co-occurrence network complexity(larger average degrees representing stronger network complexity)increased continuously from July to September and decreased in October before increasing in November.The co-occurrence network complexity of the algae and nirS-encoding denitrifying bacteria tended to increase from July to November.The co-occurrence network complexity of the algal and denitrifying bacterial communities was modified by ammonia nitrogen(NH_(4)^(+)-N)and total phosphorus(TP),pH,and water temperature(WT),according to the ordinary least-squares(OLS)model.The modified stochasticity ratio(MST)results reveal that deterministic selection dominated the assembly of denitrifying bacterial communities.The influence of environmental variables to denitrifying bacterial communities,as well as characteristics of algal-bacterial co-occurrence networks and the assembly process of denitrifying bacterial communities,were discovered in epiphytic biofilms in this study.The findings could aid in the appropriate understanding and use of epiphytic biofilms denitrification function,as well as the enhancement of water quality.

Effects of long-term elevated CO_2 on N_2-fixing,denitrifying and nitrifying enzyme activities in forest soils under Pinus sylvestriformis in Changbai Mountain

A study was conducted to determine the effects of elevated CO2 on soil N process at Changbai Mountain in Jilin Province, northeastern China （42°24＂N, 128°06＂E, and 738 m elevation）. A randomized complete block design of ambient and elevated CO2 was established in an open-top chamber facility in the spring of 1999. Changpai Scotch pine （Pinus sylvestris var. sylvestriformis seeds were sowed in May, 1999 and CO2 fumigation treatments began after seeds germination. In each year, the exposure started at the end of April and stopped at the end of October. Soil samples were collected in June and August 2006 and in June 2007, and soil nitrifying, denitrifying and N2-fixing enzyme activities were measured. Results show that soil nitrifying enzyme activities （NEA） in the 5-10 cm soil layer were significantly increased at elevated CO2 by 30.3% in June 2006, by 30.9% in August 2006 and by 11.3% in June 2007. Soil denitrifying enzyme activities （DEA） were significantly decreased by elevated CO2 treatment in June 2006 （P 〈 0.012） and August 2006 （P 〈 0.005） samplings in our study; no significant difference was detected in June 2007, and no significant changes in N2-fixing enzyme activity were found. This study suggests that elevated CO2 can alter soil nitrifying enzyme and denitrifying enzyme activities.

Isolation of Denitrifying Bacteria in Sea Sediment and Simulated Experiment of Removing Nitrate-N from Seawater

The objective of this research was to isolate denitrifying bacteria from sea sediment and simulate the removal efficiency of nitrate-N by denitrifying bacteria from seawater. The result showed that the isolated denitrifying bacteria could effectively remove nitrate-N from seawater. About 90 % of nitrate-N was removed by denitrifying bacteria from seawater within a week in the simulated experiment I （the initial concentration of nitrate-N was 100 mg/L）. The removal efficiency of nitrate-N reached about 70 % within one day in the simulated experiment Ⅱ （initial concentration of nitrate-N was 1 mg/L）. The final removal efficiency was about 98 % and 85 % in the simulated experiments Ⅰ and Ⅱ, respectively. It was found that there was positive correlation between the concentration of nitrate-N and the number of denitrifying bacteria in seawater. Lots of denitrifying bacteria would disappear and the seawater would become transparent once the process of bioremediation was completed.

Denitrifying phosphorus removal in a step-feed CAST with alternating anoxic-oxic operational strategy

A bench-scale cyclic activated sludge technology （CAST） was operated to study the biological phosphorus removal performance and a series of batch tests was carried out to demonstrate the accumulation of denitrifying polyphosphate-accumulating organisms （DNPAOs） in CAST system. Under all operating conditions, step-feed CAST with enough carbon sources in influent had the highest nitrogen and phosphorus removal efficiency as well as good sludge settling performance. The average removal rate of COD, NH4^＋-N, PO4^3--P and total nitrogen （TN） was 88.2%, 98.7%, 97.5% and 92.1%, respectively. The average sludge volume index （SVI） was 133 mL/g. The optimum anaerobic/aerobic/anoxic （AOA） conditions for the cultivation of DNPAOs could be achieved by alternating anoxic/oxic operational strategy, thus a significant denitrifying phosphorus removal occurred in step-feed CAST. The denitrification of NO^x--N completed quickly due to step-feed operation and enough carbon sources, which could enhance phosphorus release and further phosphorus uptake capability of the system. Batch tests also proved that polyphosphate-accumulating organisms （PAOs） in the step-feed process had strong denitrifying phosphorus removal capacity. Both nitrate and nitrite could be used as electron acceptors in denitrifying phosphorus removal. Low COD supply with step-feed operation strategy would favor DNPAOs accumulation.

Influence of pH on short-cut denitrifying phosphorus removal

Through a series of experiments using denitrifying phosphorus-accumulating sludge in sequencing batch reactors （SBRs）, the variations of the intracellular polymers during the anaerobic phosphorus release process at different pH values were compared, the probable reasons for different performances of phosphorus removal were examined, and system operations in a typical cycle were investigated. The results show that the phosphorus removal rate was positively correlated with pH values in a range of 6.5-8.5. When the pH value was 8.0, the anaerobic phosphorus release rate and anoxic phosphorus uptake rate of the activated sludge were 20.95 mg/（g, h） and 23.29 mg/（g, h）, respectively; the mass fraction of poly-13-hydroxybutyrate （PHB） increased to 62.87 mg/g under anaerobic conditions; the mass fraction of polyphosphate was 92.67 mg/g under anoxic conditions; and the effluent concentration of total phosphorus （TP） was 1.47 mg/L. With the increase of pH, the mass fraction of acetic acid and PHB also increased, and the absorption rate of acetic acid was equal to the disintegration rate of polyphosphate. When the pH value was above 8.0, biological phosphorus removal was achieved by chemical phosphorus precipitation, and the phosphorus removal rate decreased.

Identification and Metabolic Mechanism of Non-fermentative Short-cut Denitrifying Phosphorus-removing Bacteria

To investigate the characteristics and metabolic mechanism of short-cut denitrifying phospho- rus-removing bacteria （SDPB） that are capable of enhanced biological phosphorus removal （EBPR） using nitrite as an electron acceptor, an aerobic/anoxic sequencing batch reactor was operated under three phases. An SDPB-strain YC was screened after the sludge enrichment and was identified by morphological, physiological, biochemical properties and 16S rDNA gene sequence analysis. Denitrifying phosphorus-removing experiments were conducted to study anaerobic and anoxic metabolic mechanisms by analyzing the changes of chemical oxygen demand （COD）, phosphate, nitrite, poly-fl-hydroxybutyrate （PHB）, and glycogen. The results show that strain YC is a non-fermentative SDPB similar to Paracoccus denitrificans. As a kind of non-fermentative bacteria, the energy of strain YC was mainly generated from phosphorus release （96.2%） under anaerobic conditions with 0.32 mg P per mg synthesized PHB. Under anoxic conditions, strain YC accumulated 0.45 mg P per mg degraded PHB, which produced most of energy for phosphate accumulation （91.3%） and a little for glycogen synthesis （8.7%）. This metabolic mechanism of strain YC is different from that of traditional phosphorus-accumulating organisms （PAOs）. It is also found that PHB, a kind of intracellular polymer, plays a very important role in denitrifying and accumulating phosphorus by supplying sufficient energy for phosphorous accumulation and carbon sources for denitrification. Therefore, monitoring AP/APHB and ANO2 -N/APHB is more necessary than monitoring AP/ACOD, ANO2 -N/ACOD, or AP / ANO2 -N.

Impact of Long-Term Fertilization on Community Structure of Ammonia Oxidizing and Denitrifying Bacteria Based on amoA and nirK Genes in a Rice Paddy from Tai Lake Region,China

Ammonia oxidizing （AOB） and denitrifying bacteria （DNB） play an important role in soil nitrogen transformation in natural and agricultural ecosystems. Effects of long-term fertilization on abundance and community composition of AOB and DNB were studied with targeting ammonia monooxygenase （amoA） and nitrite reductase （nirK） genes using polymerase chain reaction- denaturing gradient gel electrophoresis （PCR-DGGE） and real-time PCR, respectively. A field trial with different fertilization treatments in a rice paddy from Tai Lake region, centre East China was used in this study, including no fertilizer application （NF）, balanced chemical fertilizers （CF）, combined organic/inorganic fertilizer of balanced chemical fertilizers plus pig manure （CFM）, and plus rice straw return （CFS）. The abundances and riehnesses of amoA and nirK were increased in CF, CFM and CFS compared to NF. Principle component analysis of DGGE profiles showed significant difference in nirK and amoA genes composition between organic amended （CFS and CFM） and the non-organic amended （CF and NF） plots. Number of amoA copies was significantly positively correlated with normalized soil nutrient richness （NSNR） of soil organic carbon （SOC） and total nitrogen （T-N）, and that of nirK copies was with NSNR of SOC, T-N plus total phosphorus. Moreover, nitrification potential showed a positive correlation with SOC content, while a significantly lower denitrification potential was found under CFM compared to under CFS. Therefore, SOC accumulation accompanied with soil nutrient richness under long-term balanced and organic/inorganic combined fertilization promoted abundance and diversity of AOB and DNB in the rice paddy.

Effect of carbon source and nitrate concentration on denitrifying phosphorus removal by DPB sludge

Effect of added carbon source and nitrate concentration on the denitrifying phosphorus removal by DPB sludge was systematically studied using batch experiments, at the same time the variation of ORP was investigated. Results showed that the denitrifying and phosphorus uptake rate in anoxic phase increased with the high initial anaerobic carbon source addition. However once the initial COD concentration reached a certain level, which was in excess to the PHB saturation of poly-P bacteria, residual COD carried over to anoxic phase inhibited the subsequent denitrifying phosphorus uptake. Simultaneously, phosphate uptake continued until all nitrate was removed, following a slow endogenous release of phosphate. High nitrate concentration in anoxic phase increased the initial denitrifying phosphorus rate. Once the nitrate was exhausted, phosphate uptake changed to release. Moreover, the time of this turning point occurred later with the higher nitrate addition. On the other hand, through on-line monitoring the variation of the ORP with different initial COD concentration, it was found ORP could be used as a control parameter for phosphorus release, but it is impossible to utilize ORP for controlling the denitrificaion and anoxic phosphorus uptake operations.

Operation of three parallel AN/AO processes to enrich denitrifying phosphorus removing bacteria for low strength wastewater treatment

Three parallel anaerobic-anoxic/anaerobic-aerobic （AN/AO） processes were developed to enrich denitrifying phosphorus removal bacteria （DPB） for low strength wastewater treatment. The main body of the parallel AN/AO process consists of an AN （anaerobic-anoxic） process and an AO （anaerobic-aerobic） process. In the AO process, the common phosphorus accumulating organisms （PAOs） was dominate, while in the AN process, DPB was dominate, The volume of anaerobic zone（Vana）：anoxie zone（Vano） ： aerobic zone （Vaer） for the parallel AN/AO process is 1：1：1 in contrast with a Vana：Vaer and Vano：Vaer of 1：2 and 1：4 for a traditional biological nutrient removal process （BNR）. Process 3 excels in the 3 processes on the basis of COD, TN and TP removal. For 4 month operation, the effluent COD concentration of process 3 did not exceed 60 mg/L; the effluent TN concentration of process 3 was lower than 15 mg/L; and the effluent TP concentration of process 3 was lower than 1 mg/L.

Denitrifying Bacteria in Paddy Soils of the Taihu Lake Basin,China

This study attempted to determine the characteristics of the communities, the ecological factors, and the denitrifying enzyme activity for denitrifying bacteria found in the paddy soils of the Taihu Lake Basin, China. Samples of the six main soil types of the basin were taken from paddy fields with different fertilities. The total numbers of bacteria and denitrifying bacteria in the high fertility soils were much more than those in low fertility soils, and the number of denitrifying bacteria accounted for 49% to 80% of the total number of bacteria. The O2 content was an important ecological factor that affected denitrification. Of test the strains isolated from the paddy soils in the Taihu Lake Basin, some (e.g., Pseudomonas spp.) grew well under low oxygen partial pressure, while others (e.g., Bacillus spp.) had no strict predilection with O2 content. Another critical ecological factor was the nitrogen concentration. Three selected denitrifying bacteria grew better in aculture medium with 135 instead of 276mg L^-1 nitrogen. At the same time 67% of the test strains were able to reduce NO3^- to NO2^- and 56% had N2O reductase.

Denitrifying phosphorous removal in anaerobic/anoxic SBR system with different startup operation mode

To achieve stable and efficient nitrogen and phosphorus removal and to investigate the characteristics of the A/A SBR enriched with denitrifying phosphorus removal bacteria(DPB),the whole course of startup was studied with two reactors operated in different mode.The reactor I was operated under anaerobic/settling/anoxic/settling mode,and the reactor II was operated under anaerobic/anoxic/settling mode.Differences between the two reactors in removal efficiency of COD,nitrogen and phosphorus were examined.The results indicated that efficient performance could be achieved in both reactors with different startup operation mode,while the phosphorus removal efficiency was improved sooner in reactor I than in reactor II,which suggested that reactor I would supply a more favorable condition for DPB proliferation.Meanwhile,it was observed that the amount of organic substrates consumption had a linear correlation to that of phosphorus release in anaerobic phase when DPB was accumulated in the A/A SBR denitrifying phosphorus removal system.

Community structure and assembly of denitrifying bacteria in epiphytic biofilms in a freshwater lake ecosystem

Denitrifying bacteria are a crucial component of aquatic ecosystem in nitrogen cycle.However,the denitrifying bacterial community dynamics and structure in epiphytic biofilms remain unexplored.The abundance of denitrification gene(nir)and structure of nirS-denitrifying bacterial community in the epiphytic biofilms collected in July and November of 2018 from a typical plateau lake(Caohai Wetland,Guizhou,China)were studied by Real-time Quantitative Polymerase Chain Reaction(qPCR)and highthroughput sequencing.Results show that the gene abundance of nirK was higher than that of nirS(P<0.05),and it was significantly different during the growth period(July)than the decline period(November).The denitrifying bacterial species was similar in the two months and shared 76.18%of OTUs.Proteobacteria(56.55%±22.15%)was the dominant phylum in all the samples.Epiphytic biofilms between growth period and decline period displayed significantly different microbial community structures due to differences in species abundance.Water temperature was the crucial factor that affected the denitrifying microbial community structure in our study.Environmental factors explain only partially the dynamic characteristics of denitrifying microbial communities,implying that the stochastic processes affected the construction of denitrifying microbial communities.As the null model analysis results show,dispersal limitation(stochastic)and undominated processes significantly influenced the assembly of denitrifying microbial communities.This study broadened our understanding of the denitrifying bacterial community structure and its function on epiphytic biofilms in freshwater ecosystems with new information provided.

Microbial Characterization of Denitrifying Sulfide Removal Sludge Using High-Throughput Amplicon Sequencing Method

The denitrifying sulfide removal(DSR) process has recently been studied extensively from an engineering perspective. However, the importance of microbial communities of this process was generally underestimated. In this study, the microbial community structure of a lab-scale DSR reactor was characterized in order to provide a comprehensive insight into the key microbial groups in DSR system. Results from high-throughput sequencing analysis revealed that the fraction of autotrophic denitrifiers increased from 2.34 % to 10.93% and 44.51% in the DSR system when the influent Na Cl increased from 0 g/L, to 4 g/L and 30 g/L, respectively. On the contrary, the fraction of heterotrophic denitrifiers decreased from 61.74% to 39.57%, and 24.12%, respectively. Azoarcus and Thiobacillus were the main autotrophic denitrifiers, and Thauera was the main hetetrophic denitrifier during the whole process. This study could be useful for better understanding the interaction between autotrophs and heterotrophs in DSR system.

Simultaneous Nitrogen and Phosphorus Removal by Denitrifying Dephosphatation in a (AO)_2 Sequencing Batch Reactor

A 24 L working volume reactor was used for the research on simultaneous phosphorus (P) and nitrogen (N) removal by denitrifying dephosphatation in an anaerobic-oxid-anoxic-oxid sequencing batch reactor ((AO)_2SBR) system. The durations of each phase are: anaerobic 1.5 h, aerobic 2.5 h, anoxic 1.5 h, post-aerobic 0.5 h, settling 1.0 h, fill 0.5 h. The successful removal of nitrogen and phosphorus is achieved in a stable (AO)_2SBR. The effluent P concentrations is below 1 mg/L, and the COD,TN and P average removal efficiency is 88.9%, 77.5% and 88.7%, respectively. The batch experiment results show that the durations of aerobic and anoxic phase influence the P removal efficiency. Some feature points are found on the DO, ORP and pH curves to demonstrate the complete of phosphate release and phosphate uptake. These feature points can be used for the control of (AO)_2 SBR.

Enhanced Performance of Denitrifying Sulfide Removal Process by 1,2-Naphthoquinone-4-Sulphonate

The denitrifying sulfide removal(DSR) process with bio-granules comprising both heterotrophic and autotrophic denitrifiers can simultaneously convert nitrate, sulfide and acetate species into di-nitrogen gas, elemental sulfur and carbon dioxide, respectively, at high loading rates. This study has determined that the reaction rate of sulfide oxidized into sulfur could be enhanced in the presence of 1,2-naphthoquinone-4-sulphonate(NQS). The presence of NQS mitigated the inhibition effects of sulfide species on denitrification. Furthermore, the reaction rates of nitrate and acetate to nitrogen gas and CO_2, respectively, were also promoted in the presence of NQS, thereby enhancing the performance of DSR granules. The advantages and disadvantages of applying the NQS-DSR process are discussed.

Continuous treatment of azo acid dyes by photo-dependent denitrifying sludge

Simultaneous removals of dye and nitrate by photo dependent denitrifying sludge(PDDS) have been demonstrated in a continuous flow bench scale reactor. The best C/N for the degradation of azo dyes by PDDS was 1.5. The specific removal rate of azo dye AB92 decreased with a decrease in hydraulic retention time and increased with a decrease in solids retention time. The degradation rate of TOC decreased with a decrease in hydraulic retention time. AB92, which has nitro and hydroxyl substitutions in non para positions, was uniquely degraded. During continuous flow treatment experiments using PDDS, complete degradation of azo dyes AB92 and AO20 at influent concentrations of 40 mg/L and 30 mg/L, respectively, was achieved with an HRT of 16.

Effect of Zinc Oxide Nanoparticles on Denitrification and Denitrifying Bacteria Communities in Typical Estuarine Sediments

For revealing the effects of increasing of zinc oxide nanoparticles(ZnO NPs)on denitrification and denitrifying bacteria communities in estuarine sediments,the surface sediments of two typical estuaries(the Yangtze River Estuary and the Yellow River Estuary)were added with medium concentration(170mgL−1)and high concentration(1700mgL−1)of ZnO NPs for anaerobic cul-ture in laboratory.The concentration of NO_(3)^(−)and NO_(2)^(−),the reductase activity and denitrification rate were measured by physico-chemical analysis,nirS gene abundance and denitrifying bacteria communities by molecular biological methods.The results showed that ZnO NPs inhibited NO_(3)^(−), NO_(2)^(−)reduction process and NO_(3)^(−), NO_(2)^(−)reductase activity,and a stronger inhibition effect resulting from the higher ZnO NPs concentration.ZnO NPs decreased nirS gene abundance and community diversity of denitrifying bacteria.In addition,the inhibition degree of ZnO NPs on the denitrification process of sediments in different estuaries was different.These results were of great significance for evaluating the potential ecological toxicity and risks of nanomaterials in estuaries.

Factors affecting biological denitrifying dephosphatation in anaerobic/anoxic/aerobic sequencing batch reactor

This study was conducted to verify and discuss the denitrifying dephosphatation under different levels of nitrate concentration and retention time of anoxic/aerobic process in a Sequencing Batch Reactor (SBR). The results of tests demonstrated that there were two kinds of phosphorus-accumulating organisms (PAOs) in the biological excess phosphorus removal (BEPR) system. One was non-DNPAOs that could only use oxygen as terminal electron acceptors, the other was denitrifying PAOs (DNPAOs) that could use both nitrate and oxygen as terminal electron acceptors. Phosphorus uptake efficiency could be attained under anoxic period ranging from 28.7%-96.7% in an anaerobic/anoxic/aerobic system. Experimental results showed that nitrate concentration and retention time of anoxic/aerobic process were the key factors affecting the course of denitrifying dephosphatation.

Nitrogen Removal Performance of Denitrifying Ammonium Oxidation System in Treating Sulfamethoxazole-laden Secondary Wastewater Effluent

In this study,nitrogen removal performance of the denitrifying ammonium oxidation(DAO)process was investigated when treating sulfamethoxazole(SMX)-laden secondary wastewater effluent.The influent SMX concentration showed negligible effect on efficiencies for removal of nitrate and COD.However,the ammonium ions removal rate was moderately reduced,when the influent SMX concentration in wastewater reached 6 mg/L.Total nitrogen removal efficiency remained as high as 76.77%towards the day 158 at the end of experiment.Candidatus_Brocadia and Candidatus_Kuenenia were the functional anammox strains.The unclassified_f__Rhodobacteraceae sp.was predominant heterotrophic denitrifying strain in the studied reactor.The concentrations of soluble extracellular polymeric substances in sludge obviously increased from 16.76 mg/g VSS to 32.31 mg/g VSS,which might protect the nitrogen removal strains from high-concentration SMX.This result provides a theoretical and technical foundation for the application of denitrifying ammonium oxidation process in treating sulfamethoxazole-laden secondary wastewater effluent.

NirS-type denitrifying bacteria in aerobic water layers of two drinking water reservoirs: Insights into the abundance, community diversity and co-existence model

The nirS-type denitrifying bacterial community is the main drivers of the nitrogen loss process in drinking water reservoir ecosystems.The temporal patterns in nirS gene abundance and nirS-type denitrifying bacterial community harbored in aerobic water layers of drinking water reservoirs have not been studied well.In this study,quantitative polymerase chain reaction(qPCR)and Illumina Miseq sequencing were employed to explore the nirS gene abundance and denitrifying bacterial community structure in two drinking water reservoirs.The overall results showed that the water quality parameters in two reservoirs had obvious differences.The qPCR results suggested that nirS gene abundance ranged from(2.61±0.12)×10^(5) to(3.68±0.16)×10^(5) copies/mL and(3.01±0.12)×10^(5) to(5.36±0.31)×10^(5) copies/mL in Jinpen and Lijiahe reservoirs,respectively.The sequencing results revealed that Paracoccus sp.,Azoarcus sp.,Dechloromonas sp.and Thauera sp.were the dominant genera observed.At species level,Cupriavidus necator,Dechloromonas sp.R-28400,Paracoccus denitrificans and Pseudomonas stutzeri accounted for more proportions in two reservoirs.More importantly,the co-occurrence network analysis demonstrated that Paracoccus sp.R-24615 and Staphylococcus sp.N23 were the keystone species observed in Jinpen and Lijiahe reservoirs,respectively.Redundancy analysis indicated that water quality(particularly turbidity,water temperature,pH and Chlorophyll a)and sampling time had significant influence on the nirS-type denitrifying bacterial community in both reservoirs.These results will shed new lights on exploring the dynamics of nirS-type denitrifying bacteria in aerobic water layers of drinking water reservoirs.