Anammox Enrichment Research Articles

Enrichment of anaerobic ammonium oxidation (anammox) bacteria for short start-up of the anammox process: a review

The application of the anaerobic ammonium oxidation (anammox) reaction in a biological nitrogen removal system to treat wastewater has become of great interest since its discovery. The anammox reaction is performed by anammox bacteria that belong to the Planctomycete phylum. The reaction occurs in the presence of ammonium using nitrite as the substrate under anaerobic conditions. However, the bacteria have an extremely slow growth rate and stringent metabolic conditions that cause difficulty in culturing and applying the system for wastewater treatment. Anammox enrichment has a long start-up period for the anammox process that hinders researchers using laboratory and full-scale systems for the first time. Many attempts have been made to culture anammox to establish a successful anammox culture with a shorter start-up period for the anammox reaction and high nitrogen removal activity. This paper reviews previous studies on anammox enrichment with emphasis on (i) inoculum selection, (ii) enrichment techniques and (iii) factors influencing anammox enrichment. This review will assist researchers in planning and designing an appropriate anammox enrichment. The findings should widen the application of anammox in biological nitrogen removal systems for nitrogenous wastewater.

Development of the combined nitritation–anammox process in an upflow anaerobic sludge blanket (UASB) reactor with anammox granules

The combined nitritation–anaerobic ammonium oxidation (anammox) process represents a promising innovative technology for biological nitrogen removal, especially for treating wastewater with low chemical oxygen demand (COD) to ammonium ratios. In the present study, a laboratory-scale upflow anaerobic sludge blanket (UASB) reactor, which has been operated under low oxygen levels (⩽0.5mgL−1) for over 280days, was shown to remove about 88% of the supplied total nitrogen loads without nitrite/nitrate accumulation. During the start-up period, the reactor was initially inoculated with enriched anammox granules and fed with synthetic wastewater, where the ammonium concentration was fixed but nitrite concentration was gradually decreased to zero. As the influent NO2−-N/NH4+-N ratio decreased, O2 supply was increased gradually to facilitate the conversion of ammonium to nitrite by ammonium oxidation bacteria (AOB). Our results showed that the nitritation–anammox granules with porous structures were enriched and featured good settling ability. Microbial community composition was investigated by fluorescence in situ hybridization (FISH), which demonstrated the coexistence of AOB and anammox bacteria in the granules. The two groups of bacteria exhibited an overlapping growth style, which can improve ammonium availability to anammox bacteria and facilitate the immediate consumption of nitrite (produced by AOB) by anammox bacteria.

Appropriate Fe (II) addition significantly enhances anaerobic ammonium oxidation (Anammox) activity through improving the bacterial growth rate.

The application of anaerobic ammonium oxidation (Anammox) process is often limited by the slow growth rate of Anammox bacteria. As the essential substrate element that required for culturing Anammox sludge, Fe (II) is expected to affect Anammox bacterial growth. This work systematically studied the effects of Fe (II) addition on Anammox activity based on the kinetic analysis of specific growth rate using data from batch tests with an enriched Anammox sludge at different dosing levels. Results clearly demonstrated that appropriate Fe (II) dosing (i.e., 0.09 mM) significantly enhanced the specific Anammox growth rate up to 0.172 d−1 compared to 0.118 d−1 at regular Fe (II) level (0.03 mM). The relationship between Fe (II) concentration and specific Anammox growth rate was found to be well described by typical substrate inhibition kinetics, which was integrated into currently well-established Anammox model to describe the enhanced Anammox growth with Fe (II) addition. The validity of the integrated Anammox model was verified using long-term experimental data from three independent Anammox reactors with different Fe (II) dosing levels. This Fe (II)-based approach could be potentially implemented to enhance the process rate for possible mainstream application of Anammox technology, in order for an energy autarchic wastewater treatment.

Impact of seeding on the start-up of one-stage deammonification MBBRs

Treating nitrogen-rich reject water from anaerobically digested sludge with deammonification has become a very beneficial side stream process. One common technique is the one-stage moving bed bioreactors (MBBRs), which in comparison with the other deammonification techniques can be started up without seeding anammox bacteria. This study investigated the impact of biofilm seeding on the start-up of one-stage deammonification MBBRs. Two lab-scale reactors were run in parallel with partial nitritation for 56 days until 11% of the carrier area in one reactor was replaced with fully developed deammonification biofilm to work as the seeding material. The seeded reactor started nitrogen reduction immediately up to a plateau of 1.3 g N m−2 d−1; after another 54 days on day 110, the reduction significantly increased. At the same time, the non-seeded reactor also started to reduce nitrogen due to deammonification. The development was followed with both nitrogen analyses and fluorescence in situ hybridization analyses. On day 134, the biofilm in both reactors contained>90% anammox bacteria and reached maximum nitrogen removal rates of 7.5 and 5.6 g N m−2 d−1 in the seeded and non-seeded reactor, respectively. Over 80% of the inorganic nitrogen was reduced. In conclusion, the seeding did not contribute to a shorter start-up time or the achieved anammox enrichment, although it did contribute to a partial, immediate nitrogen reduction. The boundary conditions are the most important factors for a successful start-up in a deammonification MBBR system.

Anaerobic ammonium-oxidising bacteria: A biological source of the bacteriohopanetetrol stereoisomer in marine sediments

Bacterially-derived bacteriohopanepolyols (BHPs) are abundant, well preserved lipids in modern and paleo-environments. Bacteriohopanetetrol (BHT) is a ubiquitously produced BHP while its less common stereoisomer (BHT isomer) has previously been associated with anoxic environments; however, its biological source remained unknown. We investigated the occurrence of BHPs in Golfo Dulce, an anoxic marine fjord-like enclosure located in Costa Rica. The distribution of BHT isomer in four sediment cores and a surface sediment transect closely followed the distribution of ladderane fatty acids, unique biomarkers for bacteria performing anaerobic ammonium oxidation (anammox). This suggests that BHT isomer and ladderane lipids likely shared the same biological source in Golfo Dulce. This was supported by examining the BHP lipid compositions of two enrichment cultures of a marine anammox species (‘Candidatus Scalindua profunda’), which were found to contain both BHT and BHT isomer. Remarkably, the BHT isomer was present in higher relative abundance than BHT. However, a non-marine anammox enrichment contained only BHT, which explains the infrequence of BHT isomer observations in terrestrial settings, and indicates that marine anammox bacteria are likely responsible for at least part of the environmentally-observed marine BHT isomer occurrences. Given the substantially greater residence time of BHPs in sediments, compared to ladderanes, BHT isomer is a potential biomarker for past anammox activity.

C2/C3 fatty acid stress on anammox consortia dominated by Candidatus Jettenia asiatica

Anaerobic ammonium oxidation (anammox) is a promising technology for autotrophic removal of ammonium and nitrite. In order to expand its industrial application niche, effects of organic matter need more specific investigation. In this study, the effects of C2/C3 fatty acids stress on anaerobic ammonium oxidation (anammox) process were evaluated in batch experiments. BLAST search analysis of 16S rRNA sequences showed that the homology of the highly enriched anammox consortia used here and Candidatus Jettenia asiatica (J. asiatica) reached 99%. Results showed that low acetate (⩽30 mg/L) and propionate (⩽50 mg/L) concentration does not significantly influence ammonium oxidation. Higher acetate/propionate concentrations caused decrease of ammonium removal. A level of acetate no more than 240 mg/L caused the decrease of ammonium consumption rate by 33%, and 29% for propionate with (<400 mg/L). J. asiatica showed higher adaptability to propionate stress than acetate. Real-time quantitative PCR (qPCR) results reveal that anammox bacteria J. asiatica are capable of growing at the present of low-concentration acetate (⩽120 mg/L)/propionate (⩽200 mg/L). The anammox hzo gene concentrations reached to round 0.5 × 108–1.0 × 108 copies/ml after 36 days cultivation with C2/C3 fatty acid. However, there is no superiority for J. asiatica consortia to grow under organotrophic conditions compared to autotrophic ones.

Physiological and kinetic characterization of a suspended cell anammox culture

Anammox related technologies are currently widely applied for nitrogen removal from sewage sludge digester rejection water. Nevertheless, many aspects of the anammox process like the kinetic characteristics and the reaction stoichiometry are still subject of debate. Parameter values reported in literature are often hampered by mass transfer limitation or by the presence of a significant side population. In this study a membrane bioreactor (MBR) based method for growing a highly enriched anammox microbial community is described. The almost pure free-cells suspension of highly active anammox bacteria was used for detailed kinetic and stoichiometric analysis of the anammox process. The anammox culture enriched during this study had a biomass specific maximum growth rate of 0.21 d−1 which is higher than ever reported before in literature. Using an experimental methodology based on imposing dynamic process conditions combined with process modeling and parameter estimation, the intrinsic nitrite half saturation constant was identified to be as low as 35 μg-N L−1. This was confirmed to be an accurate estimation in the pH range of 6.8–7.5.

Comparison of nitrogen removal rates and nitrous oxide production from enriched anaerobic ammonium oxidizing bacteria in suspended and attached growth reactors

Attached growth-systems for the anaerobic ammonium oxidation (anammox) process have been postulated for implementation in the field. However, information about the anammox process in attached growth-systems is limited. This study compared nitrogen removal rates and nitrous oxide (N2O) production of enriched anammox cultures in both suspended and attached growth sequencing batch reactors (SBRs). Suspended growth reactors (SBR-S) and attached growth reactors using polystyrene sponge as a medium (SBR-A) were used in these experiments. After inoculation with an enriched anammox culture, significant nitrogen removals of ammonium (NH4 +) and nitrite (NO2 −) were observed under NH4 +:NO2 − ratios ranging from 1:1 to 1:2 in both types of SBRs. The specific rates of total nitrogen removal in SBR-S and SBR-A were 0.52 mg N/mg VSS-d and 0.44 mg N/mg VSS-d, respectively, at an NH4 +:NO2 − ratio of 1:2. N2O production by the enriched anammox culture in both SBR-S and SBR-A was significantly higher at NH4 +:NO2 − ratio of 1:2 than at NH4 +:NO2 − ratios of 1:1 and 1:1.32. In addition, N2O production was higher at a pH of 6.8 than at pH 7.3, 7.8, and 8.3 in both SBR-S and SBR-A. The results of this investigation demonstrate that the anammox process may avoid N2O emission by maintaining an NH4 +:NO2 − ratio of less than 1:2 and pH higher than 6.8.

Real-time process dynamics monitoring in Anammox reactors.

Process dynamics in Anammox systems were evaluated through continuous monitoring of pH, oxidation reduction potential (ORP) and conductivity in two separate newly started-up sequencing batch reactors, one seeded with an enriched Anammox sludge and the other seeded with mixed activated sludge. The pH and ORP profiles exhibited characteristic patterns depending on the process dynamics during early start-up, start-up and enrichment phases of the operational period of 410 days. The simultaneously continuing processes of the start-up period showed apparent indicative trend lines in pH and ORP profiles. Conductivity profiles were consistent with the process dynamics in all phases. During the enrichment phase, conductivity decreases could quantitatively be related to process removal efficiencies and all real-time profiles exhibited specific break-points which coincided with the end of Anammox in each cycle. The end of Anammox was observed as an 'apex' on pH profiles and a 'valley' on ORP profiles. The 'apex' and 'valley' points exactly coincided with the end point of the linear decrease in the conductivity profiles. The overall findings suggested a great potential in using real-time pH, ORP and conductivity measurements for quick and reliable monitoring of Anammox systems during start-up and enrichment periods.

Shotgun metagenomic data reveals significant abundance but low diversity of “Candidatus Scalindua” marine anammox bacteria in the Arabian Sea oxygen minimum zone

Anaerobic ammonium oxidizing (anammox) bacteria are responsible for a significant portion of the loss of fixed nitrogen from the oceans, making them important players in the global nitrogen cycle. To date, marine anammox bacteria found in both water columns and sediments worldwide belong almost exclusively to “Candidatus Scalindua” species. Recently the genome assembly of a marine anammox enrichment culture dominated by “Candidatus Scalindua profunda” became available and can now be used as a template to study metagenome data obtained from various oxygen minimum zones (OMZs). Here, we sequenced genomic DNA from suspended particulate matter recovered at the upper (170 m deep) and center (600 m) area of the OMZ in the Arabian Sea by SOLiD and Ion Torrent technology. The genome of “Candidatus Scalindua profunda” served as a template to collect reads. Based on the mapped reads marine anammox Abundance was estimated to be at least 0.4% in the upper and 1.7% in the center area. Single nucleotide variation (SNV) analysis was performed to assess diversity of the “Candidatus Scalindua” populations. Most highly covered were the two diagnostic anammox genes hydrazine synthase (scal_01318c, hzsA) and hydrazine dehydrogenase (scal_03295, hdh), while other genes involved in anammox metabolism (narGH, nirS, amtB, focA, and ACS) had a lower coverage but could still be assembled and analyzed. The results show that “Candidatus Scalindua” is abundantly present in the Arabian Sea OMZ, but that the diversity within the ecosystem is relatively low.

Anaerobic ammonium oxidation in polyvinyl alcohol and sodium alginate immobilized biomass system: a potential tool to maintain anammox biomass in application

Anaerobic ammonium oxidation (anammox) has been proved to be a promising nitrogen removal method for treating ammonium-rich wastewater. However, because of the low-growth rate of anammox bacteria, maintenance of a sufficient amount of anammox biomass in reactor became a key factor in application. Gel immobilization is an efficient method to prevent biomass from being washed out and to promote hyper-concentrated cultures. This study focused on a nitrogen removal process by anammox enrichment culture immobilized in polyvinyl alcohol and sodium alginate (PVA-SA) gel beads. The rapid startup of reactor demonstrated that gel entrapment was supposed to be a highly effective technique for immobilizing anammox bacteria. The anammox bacteria present in the enrichment were identified to be Jettenia-like species (>98%). Moreover, the effect of hydraulic retention time (HRT), pH, and temperature on immobilized anammox processes were investigated. The effect of pH and temperature on the anammox process was evidently weakened in PVA-SA immobilized gel beads, however, the effect of HRT on the anammox reaction was enhanced. Therefore, a stable operated reactor could be obtained in an anaerobic sequencing batch reactor, which proved gel immobilization was an excellent method to maintain the biomass in anammox reactor for application.

Study on Active Recovery of Anaerobic Ammonia Oxidation and Start-Up of ASBR Reactor

In order to prevent the loss of microbes, added anaerobic ammonia oxidation to (ASBR) reactor plus membrane as the growth of anaerobic ammonia oxidation bacteria carrier. After 30d cultivate, the activity of anammox bacteria restore to the original level, and improve TN volume load train anammox bacteria. The whole process of NH4+-N and NO2--N removal rate has remained at more than 90%.After the activity recovery of anammox NH4+-N, NO2--N the ratio of consumption to the NO3--N generation finally tends to a certain value (1. 00b 1. 30 b 0. 25).In the process of cultivate, the sludge was gradually changed from black to red brown color .Results show that the ASBR is well suited for anammox enrichment culture of bacteria.

Enrichment of Anaerobic Ammonium Oxidation (Anammox) Bacteria for Biological Nitrogen Removal of Wastewater

Anaerobic ammonium oxidation (anammox) bacteria enrichment was explored for the potential application of ammonium rich wastewater removal. Samples of sludge from mature and young landfill leachate treatment plants were screened and used as inocula for anammox enrichment cultures. Enrichments were monitored for N-NH3, N-NO2- and N-NO3- to detect anammox potential activity. Six of the twelve enrichment cultures showed anammox activity after more than five months of enrichment period. All enrichment cultures that gave positive results were obtained from bottom part of sequencing batch reactor (SBR) lagoon indicating localization of anammox bacteria in anaerobic condition. Polymerase Chain Reaction (PCR) with specific primers targeting anammox and planctomycete were able to amplify the 16S rRNA sequence for anammox bacteria under PCR optimum condition. However, only three of six positive samples were successfully sequenced. DNA sequence analysis using NCBI (BLAST) and RDP showed that the anammox bacterial sequences of the investigated samples were identified as Candidatus Kuenenia stuttgartiensis with similarity of 100% (NCBI) and 99.3% (RDP).

Enrichment of an anammox bacterial community from a flooded paddy soil

This study describes the enrichment of anammox bacteria in a column simulating oxygen limited flooded paddy soils, which are important man-made ecosystems that receive substantial amounts of fixed nitrogen. The upper 50 cm of the paddy soil, containing a high amount of ammonium [1.6-10.4 mmol N kg (dry weight)(-1)], was selected as the inoculum for anammox enrichment. After 18 months of incubation with freshwater from the paddy soil ecosystem, the enrichment culture consumed approximately 4 mmol ammonium l(-1) day(-1) and 5 mmol nitrite l(-1) day(-1). The maximum specific anammox activity of the culture was 35.7 μmol N g (dry weight)(-1) h(-1). Fluorescence in situ hybridization indicated that anammox cells constituted 50% ± 10% of the enrichment culture. The phylogenetic analyses of 16S rRNA and the diagnostic hydrazine synthase (hzsA) genes showed that two dominant anammox species were enriched from paddy soil. The enriched Candidatus Anammoxoglobus-like organisms showed a 16S rRNA gene similarity of 97.5-99.2% to Candidatus Anammoxoglobus propionicus and the Candidatus Jettenia-like organisms showed 92.1-93.1% 16S rRNA gene identity to Candidatus Jettenia asiatica. Real-time quantitative PCR of hzsA gene suggested that up to 10(10) copies g (dry weight)(-1) of soil anammox bacteria were present in the enrichment culture.

A pilot-scale study on nitrogen removal from dry-spun acrylic fiber wastewater using anammox process

Dry-spun acrylic fiber wastewater (DAFW) is one kind of toxic refractory petrochemical wastewater. The results of GC–MS qualitative analysis demonstrated that the residual organic compounds in DAFW after the biotreatment are non-biodegradable and hardly available for traditional nitrification–denitrification processes. First, the batch test indicated that there was no obvious inhibition or toxicity of DAFW to anaerobic ammonium oxidizing (anammox) bacteria. A pilot test was then carried out to evaluate the feasibility and efficiency of biological nitrogen removal by the anammox process. A rapid start-up was achieved by inoculating enriched anammox sludge to a biofilm system with low inoculated sludge concentration. The pretreatment for anammox process was necessary because DAFW contains high SS which affected the nitrogen removal efficiency. By controlling the reactor temperature and SS content in influent, the removal efficiencies of ammonium and nitrite were 85% and 90%, respectively. The stoichiometric ratios of ammonium conversion, nitrite removal and nitrate production were 1:(1.29):(0.24) for synthetic wastewater and 1:(1.26):(0.22) for DAFW. After 180days of operation, there was a diverse change of the anammox consortium bacteria. Compared with the inoculated sludge (single Brocadia anammoxidans sp.), Kuenenia sp. became the predominant bacteria, with a small amount of Jettenia sp. and B. anammoxidans sp. The results demonstrated that ammonia in effluent of DAFW after two-stage biotreatment could be removed efficiently by the anammox process.

Anaerobic ammonium oxidation: from laboratory to full-scale application.

From discovery in the early 1990s to completion of full-scale anammox reactor, it took almost two decades to uncover the secret veil of anammox bacteria. There were three milestones during the commercialization of anammox: the development of the first enrichment culture medium, the completion of the first commercial anammox reactor, and the fast start-up of full-scale anammox plant. Till now, the culture of anammox bacteria experienced a big progress through two general strategies: (a) to start up a reactor from scratch and (b) to seed the reactor with enriched anammox sludge. The first full-scale anammox reactor took 3.5 years to realize full operation using the first approach due to several reasons besides the lack of anammox sludge. On the other hand, the first Asian anammox reactor started up in two months, thanks to the availability of anammox seed. Along with the implementation of anammox plants, anammox eventually becomes the priority choice for ammonium wastewater treatment.

Inhibition of anaerobic ammonium oxidizing (anammox) enrichment cultures by substrates, metabolites and common wastewater constituents

Anaerobic ammonium oxidation (anammox) is an emerging technology for nitrogen removal that provides a more environmentally sustainable and cost effective alternative compared to conventional biological treatment methods. The objective of this study was to investigate the inhibitory impact of anammox substrates, metabolites and common wastewater constituents on the microbial activity of two different anammox enrichment cultures (suspended and granular), both dominated by bacteria from the genus Brocadia. Inhibition was evaluated in batch assays by comparing the N2 production rates in the absence or presence of each compound supplied in a range of concentrations. The optimal pH was 7.5 and 7.3 for the suspended and granular enrichment cultures, respectively. Among the substrates or products, ammonium and nitrate caused low to moderate inhibition, whereas nitrite caused almost complete inhibition at concentrations higher than 15mM. The intermediate, hydrazine, either stimulated or caused low inhibition of anammox activity up to 3mM. Of the common constituents in wastewater, hydrogen sulfide was the most severe inhibitor, with 50% inhibitory concentrations (IC50) as low as 0.03mM undissociated H2S. Dissolved O2 showed moderate inhibition (IC50=2.3–3.8mgL−1). In contrast, phosphate and salinity (NaCl) posed very low inhibition. The suspended- and granular anammox enrichment cultures had similar patterns of response to the various inhibitory stresses with the exception of phosphate. The findings of this study provide comprehensive insights on the tolerance of the anammox process to a wide variety of potential inhibiting compounds.

Improving Anammox start-up with bamboo charcoal

Three Upflow Anaerobic Sludge Blanket (UASB) reactors were compared for Anammox enrichment using synthetic wastewater with Spherical Plastic (SP) and Bamboo Charcoal (BC) addition, and without carrier (CK). After four months of operation, the Anammox activity occurred in all reactors allowing continuous removal of ammonium and nitrite. Ammonium and nitrite removal efficiencies were all higher than 98% in steady phase with the effluent concentrations below 1mgL−1. The start-up time could be shortened from 117 to 97d in CK and SP reactor to 85d in BC amendment reactor. Quantitative PCR (q-PCR) analyses indicated a significant increase in the number of Anammox bacteria in BC amended reactor as compared with CK and SP during the entire start-up periods. The copy numbers of Anammox of 16SrRNA gene in the reactor with BC amendment could reach up to 6×109copiesg−1 Volatile Suspended Solids, around 22.5times and 12.3times greater than that in CK and SP reactor, respectively. BC addition could accelerate the start-up of Anammox and significantly increase the Anammox bacteria number.

The metagenome of the marine anammox bacterium 'Candidatus Scalindua profunda' illustrates the versatility of this globally important nitrogen cycle bacterium.

Anaerobic ammonium-oxidizing (anammox) bacteria are responsible for a significant portion of the loss of fixed nitrogen from the oceans, making them important players in the global nitrogen cycle. To date, marine anammox bacteria found in marine water columns and sediments worldwide belong almost exclusively to the ‘Candidatus Scalindua’ species, but the molecular basis of their metabolism and competitive fitness is presently unknown. We applied community sequencing of a marine anammox enrichment culture dominated by ‘Candidatus Scalindua profunda’ to construct a genome assembly, which was subsequently used to analyse the most abundant gene transcripts and proteins. In the S. profunda assembly, 4756 genes were annotated, and only about half of them showed the highest identity to the only other anammox bacterium of which a metagenome assembly had been constructed so far, the freshwater ‘Candidatus Kuenenia stuttgartiensis’. In total, 2016 genes of S. profunda could not be matched to the K. stuttgartiensis metagenome assembly at all, and a similar number of genes in K. stuttgartiensis could not be found in S. profunda. Most of these genes did not have a known function but 98 expressed genes could be attributed to oligopeptide transport, amino acid metabolism, use of organic acids and electron transport. On the basis of the S. profunda metagenome, and environmental metagenome data, we observed pronounced differences in the gene organization and expression of important anammox enzymes, such as hydrazine synthase (HzsAB), nitrite reductase (NirS) and inorganic nitrogen transport proteins. Adaptations of Scalindua to the substrate limitation of the ocean may include highly expressed ammonium, nitrite and oligopeptide transport systems and pathways for the transport, oxidation, and assimilation of small organic compounds that may allow a more versatile lifestyle contributing to the competitive fitness of Scalindua in the marine realm.

Enrichment of anammox bacteria from mudflat sediments collected in Tokyo Bay

We established an enrichment culture of anammox bacteria by seeding a batch reactor with sediments from the Banzu mudflat in Tokyo Bay. The reactor was intermittently fed with an anammox substrate containing NH4Cl, NaNO2 and artificial seawater (3.03% salinity) for more than 800 days. An anammox enrichment culture capable of simultaneous reduction of approximately equivalent amounts of ammonium nitrogen and nitrite nitrogen was established. Batch assays of the enrichment culture revealed that anammox activity was similar for all batches at salinities ranging from 1.01 to 3.03%, with lower activity observed at a salinity of 0.02%. We identified three 16S rRNA genes in DNA fragments amplified from the enriched culture. One of these genes, mudflat sediment clone_K01 was highly homologous with Candidatus “Kuenenia stuttgartiensis” (identity 97%) being the most abundant (85 out of isolated 94 clones). In most of the studies conducted to date, anammox species belonging to the Candidatus “Scalindua” genus have been found to be the dominant species in enrichment cultures obtained from estuarine and marine environments. The results reported here are unusual, in that, a freshwater anammox bacterium, Candidatus K. stuttgartiensis, was the dominant bacterium in a sediment sample from a marine environment. Key words: Anammox, mudflat sediment, salinity, Candidatus “Kuenenia stuttgartiensis”,Candidatus “Scalindua wagneri”