Enrichment of ANAMMOX bacteria from conventional activated sludge entrapped in poly(vinyl alcohol)/sodium alginate gel Hyokwan Bae a,⇑ , Minkyu Choi a , Changsoo Lee b , Yun-Chul Chung a , Young Je Yoo c,⇑ , Seockheon Lee a a Center for Water Resource Cycle Research, Korea Institute of Science and Technology, 39-1 Hawolgok-Dong, Seongbuk-Gu, Seoul 136-791, Republic of Korea b School of Urban and Environmental Engineering, Ulsan National Institute of Science and Technology (UNIST), Ulsan 689-798, Republic of Korea c Interdisciplinary Program of Bioengineering, Seoul National University, San 56-1, Shilim-Dong, Kwanak-Gu, Seoul 151-742, Republic of Korea highlights  Conventional activated sludge was utilized for ANAMMOX enrichment.  PVA/alginate beads efficiently retained biomass in an enrichment reactor.  Exponential growth rate was enhanced with a higher nitrogen loading rate.  The bead size adversely affected the lag period with limited mass transport.  The Candidatus ‘Brocadia sinica’-related species was identified. article info Article history: Received 14 January 2015 Received in revised form 18 June 2015 Accepted 25 June 2015 Available online 3 July 2015 Keywords: Anaerobic ammonium oxidation Poly(vinyl alcohol)/sodium alginate bead Enrichment Conventional activated sludge Start-up abstract The inoculation of conventional activated sludge is a prospective option against transferring anaerobic ammonium oxidation (ANAMMOX) biomass from a currently existing ANAMMOX process in places where large amounts of ANAMMOX biomass are not available. In this study, poly(vinyl alcohol)/sodium alginate gel beads were utilized as an immobilization system to entrap activated sludge. Despite the longer start-up periods than those in previous examples using the ANAMMOX inoculum, the acclimation of ANAMMOX bacteria in the gel beads was successful. The maximum nitrogen removal rate of 1.12 kg N/m 3 -day was established at a nitrogen loading rate of 1.26 ± 0.04 kg N/m 3 -day with a total nitro- gen removal efficiency of 88.9%. The exponential growth rate in the initial phase was enhanced by a higher nitrogen loading rate. The area-to-volume ratio of the gel beads was the significant control factor for the lag period, showing an inverse relationship due to the internal mass transport limitation. Based on the 16S rRNA gene, a sharp increase in the amount of ANAMMOX bacteria was identified along with an increase in the nitrogen removal rate. The responsible ANAMMOX bacteria were the species related to Candidatus ‘Brocadia sinica’. Ó 2015 Elsevier B.V. All rights reserved. 1. Introduction The combination of partial nitritation (PN) and anaerobic ammonium oxidation (ANAMMOX) represents a new cost-effective nitrogen removal process which provides a short-cut pathway for nitrogen removal (Eqs. (1) and (2)). ANAMMOX bacteria have a unique metabolic pathway through which nitrogen is removed by oxidizing NH 4 + with NO 2  as an elec- tron acceptor. Through this reaction, N 2 (g) and a small amount of NO 3  are produced under autotrophic and anaerobic conditions (Eq. (2)). Compared to conventional nitrification and denitrification processes, the PN-ANAMMOX process theoretically reduces the costs of aeration and an addition of organic carbon to 42% and http://dx.doi.org/10.1016/j.cej.2015.06.111 1385-8947/Ó 2015 Elsevier B.V. All rights reserved. Abbreviations: CAS, wastewater treatment system using conventional activated sludge; PAB, pre-cultured ANAMMOX biomass; PN, partial nitritation, oxidizing reaction of a half of NH 4 + to NO 2  ; ANAMMOX, anaerobic ammonium oxidation; AOB, ammonia-oxidizing bacteria; AOA, ammonia-oxidizing archaea; NOB, nitrite-oxidizing bacteria; A2O, anaerobic-anoxic-oxic; SBR, sequencing batch reactor; MBR, membrane bioreactor; CSTR, continuously stirred tank reactor; PVA, poly(vinyl alcohol); PVA/SA, mixture of poly(vinyl alcohol) and sodium alginate; PEG, poly(ethylene glycol); 16S rRNA, small subunit ribosomal RNA; FISH, fluorescence in situ hybridization; HRT, hydraulic retention time; SRT, solid retention time; NLR, nitrogen loading rate; NRR, nitrogen removal rate; TN, total nitrogen; TSS, total suspended solid; VSS, volatile suspended solid; FNA, free nitric acid; A b /V b , the area-to-volume ratio of the PVA/SA gel beads; l, specific growth rate; l max , maximum specific growth rate; Ks NH4 , half-saturation constant for NH 4 + ; Ks NO2 , half-saturation constant for NO 2  ; EPS, extracellular polymeric substances. ⇑ Corresponding authors. Tel.: +82 2 958 6796; fax: +82 2 958 5839 (H. Bae). E-mail addresses: lotus@kist.re.kr (H. Bae), yjyoo@snu.ac.kr (Y.J. Yoo). Chemical Engineering Journal 281 (2015) 531–540 Contents lists available at ScienceDirect Chemical Engineering Journal journal homepage: www.elsevier.com/locate/cej