Evaluation of nitrogen removal processes and microbial communities in eight full-scale municipal wastewater treatment plants

VIEWS - 115 (Abstract) 46 (PDF) 5 (Supplementary Materials)
Jia Yan, Jie-Hui Xie, Si-Ji Wang, Hong-Guo Zhang, Jia-Peng Wu, Yi-Guo Hong


Wastewater treatment plants (WWTPs) face great challenges in developing countries, such as China, due to increasingly strict integrated wastewater discharge standard, especially for ammonium. Thus, it is very important to understand the limiting factor for ammonium removal in WWTPs. In this study, samples from 8 full-scale municipal WWTPs with different treatment processes in Pearl River Delta (PRD) area were evaluated. Bacterial and archaeal communities were investigated by high-throughput sequencing, activity of aerobic ammonium and nitrite oxidation, denitrification and anammox processes were evaluated. Nitrite, nitrate and TP concentration were strongly correlated with bacterial and archaeal composition in WWTPs based on canonical correspondence analysis (CCA). Aerobic ammonium oxidation activities were over 10 times lower than aerobic nitrite oxidation, denitrification and anammox activities in WWTPs samples, which indicated nitrogen removal activities in WWTPs were limited by aerobic ammonium oxidation, because of lack of nitrite/nitrate for denitrification and anammox. Moreover, a technology-dependent shaping of microbial diversity and nitrogen removal activity was observed. The highest bacterial diversity and nitrogen removal activities were achieved in sequencing batch reactor (SBR) and Modified Anaerobic-Anoxic-Oxic (MAAO) processes, which implied the advantage of these two treatment technologies in nitrogen removal. Therefore, this study suggested enhancing activity of aerobic ammonium oxidation might be a potential solution, for promoting ammonium removal and benefiting sustainable management of WWTPs in future.


microbial community, WWTPs, nitrogen removal, activated sludge, Pearl River Delta


American Public Health Association (APHA), American Water Works Association (AWWA) and Water Environment Federation (WEF), 2005, Standard Methods for the Examination of Water and Wastewater. Washington, D.C.:USAhttp://www.doi.org/10.3354/meps210223

Arrigo, K.R., 2005. Erratum: marine microorganisms and global nutrient cycles. Nature, 437, 349-355.http://www.doi.org/10.1016/s0025-326x(02)00474-5

Bai, Y., Sun, Q., Wen, D. and Tang, X., 2012. Abundance of ammonia-oxidizing bacteria and archaea in industrial and domestic wastewater treatment systems. FEMS Microbiology Ecology, 80, 323-330. http://www.doi.org/10.1007/s00267-010-9440-3

Brenner, A., 2010. Removal of nitrogen and phosphorus compounds in biological treatment of municipal wastewater in Israel. Israel Journal of Chemistry, 46, 45-51.http://www.doi.org/10.1155/2010/914159

Castellano-Hinojosa, A., Correa-Galeote, D., Carrillo, P., Bedmar, E.J. and Medina-Sánchez, J.M., 2017. Denitrification and biodiversity of denitrifiers in a high-mountain mediterranean lake. Frontiers in Microbiology, 8, 1911. http://www.doi.org/10.2307/1941340

Cébron, A. and Garnier, J., 2005. Nitrobacter, and Nitrospira, genera as representatives of nitrite-oxidizing bacteria: detection, quantification and growth along the lower seine river (France). Water Research, 39, 4979-4992. http://www.doi.org/10.1007/s00253-011-3408-y

Cheema, S., Zeyer, J. and Henneberger, R., 2015. Methanotrophic and methanogenic communities in Swiss alpine fens dominated by Carex rostrata and Eriophorum angustifolium. Applied and Environmental Microbiology, 81, 5832.http://www.doi.org/10.1371/journal.pone.0113603

Cloern, J.E., 2001. Our evolving conceptual model of the coastal eutrophication problem. Marine Ecology Progress Series, 210, 223-253.http://www.doi.org/10.1038/nature03911

Cortés-Lorenzo, C., Sipkema, D., Rodríguez-Díaz, M., Fuentes, S., Juárez-Jiménez, B., Rodelas, B., Smidt, H. and González-López, J., 2014. Microbial community dynamics in a submerged fixed bed bioreactor during biological treatment of saline urban wastewater. Ecological Engineering, 71, 126-132. http://www.doi.org/10.1073/pnas.1013488108

Dosta, J., Galí, A., Benabdallah, E.T., Macé, S. and Mata-Alvarez, J., 2007. Operation and model description of a sequencing batch reactor treating reject water for biological nitrogen removal via nitrite. Bioresource Technology, 98, 2065-2075.http://www.doi.org/10.1016/S0958-1669(00)00211-1

Emamjomeh, M.M., Tahergorabi, M., Farzadkia, M. and Bazrafshan, E., 2017. A review of the use of earthworms and aquatic worms for reducing sludge produced: an innovative ecotechnology. Waste and Biomass Valorization, 10, 1-15. http://www.doi.org/10.1038/nature16459

Ettwig, K.F., Shima, S., van, de, Pas-Schoonen, K.T., Kahnt, J., Medema, M.H., Op, den, Camp, H.J.M., Jetten, M.S.M. and Strous, M., 2008. Denitrifying bacteria anaerobically oxidize methane in the absence of archaea. Environmental Microbiology, 10, 3164-3173. http://www.doi.org/10.1128/aem.68.3.1312-1318.2002

Fan, X.Y., Gao, J.F., Pan, K.L., Li, D.C. and Dai, H.H., 2017. Temporal dynamics of bacterial communities and predicted nitrogen metabolism genes in a full-scale wastewater treatment plant. RSC Advances, 7, 56317-56327. http://www.doi.org/10.1099/00207713-47-1-150

Figuerola, E.L. and Erijman, L., 2010. Diversity of nitrifying bacteria in a full-scale petroleum refinery wastewater treatment plant experiencing unstable nitrification. Journal of Hazardous Material, 181, 281-288.http://www.doi.org/10.1038/NGEO1683

Fisher, T.R., 1985. Review: the marine nitrogen cycle. Ecology, 1, 315-316.http://www.doi.org/10.1128/aem.00402-06

Fisk, L.M., Maccarone, L.D., Barton, L. and Murphy, D.V., 2015. Nitrapyrin decreased nitrification of nitrogen released from soil organic matter but not amoA gene abundance at high soil temperature. Soil Biology and Biochemistry, 88, 214-223. http://www.doi.org/10.1111/j.1574-6941.2012.01296.x

Gantner, S., Andersson, A.F., Alonso-Sáez, L. and Bertilsson, S., 2011. Novel primers for 16s rRNA-based archaeal community analyses in environmental samples. Journal of Microbiological Methods , 84, 12-18. http://www.doi.org/10.1016/j.watres.2015.07.005

Gonzalez-Martinez, A., Rodriguez-Sanchez, A., van, Loosdrecht, M.C.M., Gonzalez-Lopez, J. and Vahala, R., 2016. Detection of comammox bacteria in full-scale wastewater treatment bioreactors using tag -454-pyrosequencing. Environmental Science and Pollution Research, 23, 25501-25511. http://www.doi.org/10.1007/s11356-016-7914-4

Guo, J., Ling, N., Chen, H., Zhu, C., Kong, Y.L., Wang, M., Shen, Q.R. and Guo, S.W., 2017. Distinct drivers of activity, abundance, Diversity and composition of ammonia-oxidizers: evidence from a long-term field experiment. Soil Biology and Biochemistry, 115, 403-414.

Hai, R., Wang, Y., Wang, X., Li, Y. and Du, Z.Z., 2014. Bacterial community dynamics and taxa-time relationships within two activated sludge bioreactors. PLOS One, 9, e90175.http://www.doi.org/10.1016/j.biortech.2018.02.012

Harter, J., Weigold, P., El-Hadidi, M., Huson, D.H., Kappler, A. and Behrens, S., 2016. Soil biochar amendment shapes the composition of n 2 o-reducing microbial communities. Science of The Total Environment, 562, 379-390.

Herlemann, D.P., Labrenz, M., Jürgens, K., Bertilsson, S., Waniek, J.J. and Andersson, A.F., 2011. Transitions in bacterial communities along the 2000 km salinity gradient of the baltic sea. The ISME Journal, 5, 1571-1579. http://www.doi.org/10.1016/j.biortech.2012.04.061

Herrmann, M., Opitz, S., Harzer, R., Totsche, K.U. and Küsel, K., 2017. Attached and suspended denitrifier communities in pristine limestone aquifers harbor high fractions of potential autotrophs oxidizing reduced iron and sulfur compounds. Microbial Ecology, 74, 1-14. http://www.doi.org/10.1371/journal.pone.0113603

Hsu, S.C., Lai, Y.C., Hsieh, P.H., Cheng, P.J., Wong, S.S. and Hung, C.H., 2014. Successful enrichment of rarely found Candidatus anammoxoglobus propionicus from leachate sludge. Journal of Microbiology and Biotechnology, 24(7), 879-887. http://www.doi.org/10.1016/j.jhazmat.2010.05.009

Hu, B.L., He, Z.F., Sha, G., Cai, C., Lou, L.P., Zheng, P. and Xu, X.H., 2014. Cultivation of nitrite-dependent anaerobic methane-oxidizing bacteria: impact of reactor configuration. Applied Microbiology and Biotechnology , 98, 7983-7991. http://www.doi.org/10.1371/journal.pone.0090175

Hu, B.L., Zheng, P., Tang, C.J., Chen, J.W., van, der, Biezen, E., Zhang, L., Ni, B.J., Jetten, M.S.M., Yan, J., Yu, H.Q. and Kartal, B., 2010. Identification and quantification of anammox bacteria in eight nitrogen removal reactors. Water Research, 44, 5014-5020.http://www.doi.org/10.1016/j.ibiod.2017.01.022

Hu, M., Wang, X., Wen, X. and Xia, Y., 2012. Microbial community structures in different wastewater treatment plants as revealed by 454-pyrosequencing analysis. Bioresource Technology, 117, 72-79. http://www.doi.org/10.1016/j.ecoleng.2014.07.025

Huang, X.P., Huang, L.M. and Yue, W.Z., 2003. The characteristics of nutrients and eutrophication in the pearl river estuary, South China. Marine Pollution Bulletin, 47, 30-36.http://www.doi.org/10.1016/j.micres.2014.12.013

Jetten, M.S.M., Wagner, M., Fuerst, J., van, Loosdrecht, M., Kuenen, G. and Strous, M., 2001. Microbiology and application of the anaerobic ammonium oxidation (‘anammox’) process. Current Opinion in Biotechnology, 12, 283-288.http://www.doi.org/10.1016/j.scitotenv.2016.03.220

Kartal, B., Rattray, J., van, Niftrik, L.A., van, de, Vossenberg, J., Schmid, M.C., Webb, R.I., Schouten, S., Fuerst, J.A., Damste, J.S., Jetten, M.S. and Strous, M., 2007. Candidatus “anammoxoglobus propionicus” a new propionate oxidizing species of anaerobic ammonium oxidizing bacteria. Systematic and Applied Microbiology, 30(1), 39-49. http://www.doi.org/10.1016/j.scitotenv.2017.08.275

Könneke, M., Bernhard, A.E., de, la, Torre, J.R., Walker, C.B., Waterbury, J.B. and Stahl, D.A., 2005. Isolation of an autotrophic ammonia-oxidizing marine archaeon. Nature, 437, 543-546.http://www.doi.org/10.1038/ismej.2011.41

Langone, M., Yan, J., Haaijer, S.C.M., Op, den, Camp, H.J., Jetten, M.S. and Andreottola, G., 2014. Coexistence of nitrifying, anammox and denitrifying bacteria in a sequencing batch reactor. Frontiers in Microbiology, 5, 28.http://www.doi.org/10.1016/j.mimet.2010.10.001

Le, C., Zha, Y., Li, Y., Sun, D., Lu, H. and Yin, B., 2010. Eutrophication of lake waters in china: cost, causes, and control. Environmental Management, 45, 662-668.http://www.doi.org/10.1128/AEM.66.11.5066-5072.2000

Li, L., Qian, G.S., Ye, L.L., Hu, X.M., Yu, X. and Lyu, W.J., 2018. Research on the enhancement of biological nitrogen removal at low temperatures from ammonium-rich wastewater by the bio-electrocoagulation technology in lab-scale systems, pilot-scale systems and a full-scale industrial wastewater treatment plant. Water Research, 140, 77. http://www.doi.org/10.1128/aem.00062-07

Liu, F., Zhang, S.N., Wang, Y., Li, Y., Xiao, R.L., Li, H.F., He, Y., Zhang, M.M., Wang, D., Li, X. and Wu, J.S., 2016. Nitrogen removal and mass balance in newly-formed myriophyllum aquaticum mesocosm during a single 28-day incubation with swine wastewater treatment. Journal of Environmental Management, 166, 596-604. http://www.doi.org/10.1128/AEM.01541-09

Liu, H.Q., Lam, J.C., Li, W.W., Yu, H.Q. and Lam, P.K.S., 2017. Spatial distribution and removal performance of pharmaceuticals in municipal wastewater treatment plants in china. Science of The Total Environment, 586, 1162-1169.http://www.doi.org/10.1128/AEM.01519-15

Liu, X., Meng, L., Castelle, C.J., Probst, A.J., Zhou, Z.C., Pan, J., Liu, Y., Banfield, J.F. and Gu, J.D., 2018. Insights into the ecology, evolution, and metabolism of the widespread Woesearchaeotal lineages. Microbiome, 6, 102. http://www.doi.org/10.1016/j.syapm.2010.08.003

Ma, Q., Qu, Y,Y,, Zhang, X.W., Shen, W.L., Liu, Z.Y., Wang, J.W., Zhang, Z.J. and Zhou, J.T., 2015. Identification of the microbial community composition and structure of coal-mine wastewater treatment plants. Microbiological Research, 175, 1-5. http://www.doi.org/10.3389/fmicb.2014.00028

Martens-Habbena, W., Berube, P.M., Urakawa, H., de, la, Torre, J.R. and Stahl, D.A., 2009. Ammonia oxidation kinetics determine niche separation of nitrifying archaea and bacteria. Nature, 461, 976-979.https://www.doi.org/10.1016/j.ibiod.2018.05.010

Meng, H., Yang, Y.C., Lin, J.G., Denecke, M. and Gu, J.D., 2017. Occurrence of anammox bacteria in a traditional full-scale wastewater treatment plant and successful inoculation for new establishment. International Biodeterioration and Biodegradation, 120, 224-231.http://www.doi.org/10.1111/j.1462-2920.2012.02894.x

Nejidat, A., Khaimov, A., Ronen, Z. and Brenner, A., 2006. Enhanced nitrification in industrial wastewater after augmentation by soil nitrifying enrichments. International Journal of Environmental Technology and Management, 6, 489-496.http://www.doi.org/10.1007/BF02996984

Ortiz-Alvarez, R. and Casamayor, E.O., 2016. High occurrence of Pacearchaeota and Woesearchaeota (archaea superphylum DPANN) in the surface waters of oligotrophic high‐altitude lakes. Environmental Microbiology Reports, 8(2), 210-217.http://www.doi.org/10.1111/1758-2229.12370

Pan, K.L., Gao, J.F., Li, H.Y., Fan X.Y., Li, D.C. and Jiang, H., 2018. Ammonia-oxidizing bacteria dominate ammonia oxidation in a full-scale wastewater treatment plant revealed by DNA-based stable isotope probing. Bioresource Technology, 256, 152-159. http://www.doi.org/10.1111/1462-2920.13754

Park, H.D., Wells, G.F., Bae, H. and Francis, C.A., 2006. Occurrence of ammonia-oxidizing archaea in wastewater treatment plant bioreactors. Applied and Environmental Microbiology, 72, 5643-5647. http://www.doi.org/10.1186/s40168-018-0488-2

Posmanik, R., Gross, A. and Nejidat, A., 2014. Effect of high ammonia loads emitted from poultry-manure digestion on nitrification activity and nitrifier-community structure in a compost biofilter. Ecological Engineering, 140-147. http://www.doi.org/10.1007/978-3-642-30141-4

Qiu, Y., Shi, H. and He, M., 2010. Nitrogen and phosphorous removal in municipal wastewater treatment plants in china: a review. Int. J. Chem. Engineer, 2010, 10.http://www.doi.org/10.3389/fmicb.2017.01911

Rosenberg, E., Delong, E.F., Thompson, F. and Lory, S., 2013. The prokaryotes: prokaryotic physiology and biochemistry.http://www.doi.org/10.1007/s00248-017-0950-x

Saghaï, A., Gutiérrezpreciado, A., Deschamps, P., Moreira, D., Bertolino, P., Ragon, M. and López-García P., 2017. Unveiling microbial interactions in stratified mat communities from a warm saline shallow pond. Environmental Microbiology, 19(6), 2405.http://www.doi.org/10.1016/j.watres.2010.07.021.

Schloss, P.D., Westcott, S.L., Ryabin, T., Hall, J.R., Hartmann, M., Hollister, E.B., Lesniewski, R.A., Oakley, B.B., Parks, D.H., Robinson, C.J., Sahl, J.W., Stres, B., Thallinger, G.G., Van, Horn, D.J. and Weber, C.F., 2009. Introducing mothur: open-source, platform-independent, community-supported software for describing and comparing microbial communities. Applied and Environmental Microbiology, 75, 7537-7541.http://www.doi.org/10.1016/j.watres.2005.10.006

Strous, M., Fuerst, J.A., Kramer, E.H.M., Logemann, S., Muyzer, G., van, de, Pas-Schoonen, K.T., Webb, R., Kuenen, J.G. and Jetten, M.S., 1999. Missing lithotroph identified as Planctomycete. Nature, 400, 446-449.http://www.doi.org/10.1128/AEM.68.8.4153-4157.2002

Sun, S.P., I, Nàcher, C.P., Merkey, B., Zhou, Q., Xia, S.Q., Yang, D.H., Sun, J.H. and Semts, B.F., 2010. Effective biological nitrogen removal treatment processes for domestic wastewaters with low C/N ratios: a review. Environmental Engineering Science, 27, 111-126. http://www.doi.org/10.1016/j.syapm.2006.03.004

Sun, W., Xiao, E.Z., Pu, Z.L., Krumins, V., Dong, Y.R., Li, B.Q. and Hu, M., 2018. Paddy soil microbial communities driven by environment- and microbe-microbe interactions: a case study of elevation-resolved microbial communities in a rice terrace. Science of The Total Environment , 612, 884-893.http://www.doi.org/10.4014/jmb.1401.01016

Takai, K. and Horikoshi, K., 2000. Rapid detection and quantification of members of the archaeal community by quantitative PCR using fluorogenic probes. Applied and Environmental Microbiology, 66, 5066-5072. http://www.doi.org/10.1007/s00253-012-4036-x

Tang, H.L. and Chen, H., 2015. Nitrification at full-scale municipal wastewater treatment plants: evaluation of inhibition and bioaugmentation of nitrifiers. Bioresource Technology, 190, 76-81. http://www.doi.org/10.1038/22749

Thamdrup, B. and Dalsgaard, T., 2002. Production of N2 through anaerobic ammonium oxidation coupled to nitrate reduction in marine sediments. Applied and Environmental Microbiology, 68, 1312-1318. http://www.doi.org/10.1111/j.1462-2920.2008.01724.x

Tourna, M., Stieglmeier, M., Spang, A., Könneke, M., Schintlemeister, A., Urich, T., Engel, M., Schloter, M., Wagner, M., Richter, A. and Schleper, C., 2011. Nitrososphaera viennensis, an ammonia oxidizing archaeon from soil. Proceedings of the National Academy of Sciences of the United States of America, 108, 8420-8425.http://www.doi.org/10.1007/s00253-014-5835-z

Van, Kessel, M.A.H.J., Speth, D.R., Albertsen, M., Nielsen, P.H., Op, den, Campet, H.J.M., Kartal, B., Jetten, M.S.M. and Lücker, S., 2015. Complete nitrification by a single microorganism. Nature, 528, 555-559.http://www.doi.org/10.1016/j.ecoleng.2013.10.033

Wang, J. and Gu, J., 2013. Dominance of Candidatus Scalindua species in anammox community revealed in soils with different duration of rice paddy cultivation in northeast china. Applied Microbiology and Biotechnology, 97(4), 1785-1798. http://www.doi.org/10.1016/j.jenvman.2015.11.020

Wang, Q., Garrity, G.M., Tiedje, J.M. and Cole, J.R., 2007. Naive bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy. Applied and Environmental Microbiology, 73, 5261-5267. https://www.doi.org/10.1016/j.soilbio.2015.05.029

Wang, S., Peng, Y., Ma, B., Wang, S.Y. and Zhu, G.B., 2015. Anaerobic ammonium oxidation in traditional municipal wastewater treatment plants with low-strength ammonium loading: widespread but overlooked. Water Research, 8, 66-75.https://www.doi.org/10.1016/j.soilbio.2017.09.007

Wang, Z., Zhang, X.X., Lu, X., Liu, B., Li, Y., Long, C. and Li, A., 2014. Abundance and diversity of bacterial nitrifiers and denitrifiers and their functional genes in tannery wastewater treatment plants revealed by high-throughput sequencing. PLOS One, 9, e113603.http://www.doi.org/10.1089/ees.2009.0100

Wang, Z., Zhang, X.X., Lu, X., Liu, B., Li, Y., Long, C. and Li, A., 2014. Abundance and diversity of bacterial nitrifiers and denitrifiers and their functional genes in tannery wastewater treatment plants revealed by high-throughput sequencing. PLOS One, 9, e113603.http://www.doi.org/10.1016/j.biortech.2006.04.033

Ward, B.B. and O'Mullan, G.D., 2002. Worldwide distribution of nitrosococcus oceani, a marine ammonia-oxidizing γ-proteobacterium, detected by PCR and sequencing of 16s rRNA and amoA genes. Applied and Environmental Microbiology, 68, 4153-7. http://www.doi.org/10.1016/j.scitotenv.2017.02.107

Xu, D., Gu, J., Li, Y., Zhang, Y., Howard, A., Guan, Y.D., Li, J.H. and Xu, H., 2016. Purifying capability, enzyme activity, and nitrification potentials in December in integrated vertical flow constructed wetland with earthworms and different substrates. Environmental Science and Pollution Research, 23, 273-281. http://www.doi.org/10.1038/nature08465.

Yan, J., Haaijer, S.C.M., Op, den, Camp, H.J.M., van, Niftrik, L., Stahl, D.A., Könneke, M., Rush, D., Sinninghe, Damsté, J.S., Hu, Y.Y. and Jetten, M.S.M., 2012. Mimicking the oxygen minimum zones: stimulating interaction of aerobic archaeal and anaerobic bacterial ammonia oxidizers in a laboratory-scale model system. Environmental Microbiology, 14, 3146-3158. http://www.doi.org/10.1504/IJETM.2006.010480

Yan, J., Op, den, Camp, H.J.M., Jetten, M.S.M., Hu, Y.Y. and Haaijer, S.C.M., 2010. Induced cooperation between marine nitrifiers and anaerobic ammonium-oxidizing bacteria by incremental exposure to oxygen. Systematic and Applied Microbiology, 33, 407-415. http://www.doi.org/10.1016/j.biortech.2015.04.063

Yan, J., Zhong, K.Q., Wang, S.J., Chen, Z.X., Hu, H.H., Jian, Z.Y., Wen, H.J. and Zhang, H.G., 2018. Carbon metabolism and sulfate respiration by a non-conventional Citrobacter freundii strain SR10 with potential application in removal of metals and metalloids. International Biodeterioration and Biodegradation. http://www.doi.org/10.1016/j.watres.2018.04.036

Zhang, T., Ye, L., Tong, A.H.Y., Shao, M.F. and Lok, S., 2011. Ammonia-oxidizing archaea and ammonia-oxidizing bacteria in six full-scale wastewater treatment bioreactors. Applied Microbiology and Biotechnology , 91, 1215-1225.http://www.doi.org/10.1007/s12649-017-9907-z

Zhu, G., Wang, S., Wang, W., Wnag Y., Zhou, L.L., Jiang, B., Op, den, Camp, H.J.M., Risgaard-Petersen, N., Schwark, L., Peng, Y.Z., Hefting, M.M., Jetten, M.S.M. and Yin, C.Q., 2013. Hotspots of anaerobic ammonium oxidation at land-freshwater interfaces. Nature Geoscience, 6, 103-107. http://www.doi.org/10.1007/s11356-015-5734-6

DOI: http://dx.doi.org/10.26789/AEB.2019.01.007


  • There are currently no refbacks.

Copyright (c) 2019 Jia Yan

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.