Aerobic Granular Sludge Reactor Research Articles

Nitrogen removal pathways and microbial community structure in the aerobic granular sludge reactor with co-existence of granules and flocs

Granules and floc in the aerobic granular sludge (AGS) reactor undertook different roles in the nitrogen removal process and displayed significant disparities in microbial communities. This study quantified the contribution of different nitrogen removal pathways and analyzed the nitrogen removal mechanisms inside granules and flocs by metagenomics sequencing. The results showed that the AGS reactor performed excellent nitrogen removal efficiency and the removal efficiencies of NH4+-N and total nitrogen were more than 90.0 % and 60.0 %, respectively, with the main nitrogen form of NO2--N in the effluent. Batch experiments indicated that the nitrogen removal pathway was principally shortcut nitrification and denitrification (SCND), accounting for 51.6 %, followed by anaerobic ammonium oxidation (Anammox) accounting for 26.1 % and the remaining 22.3 % was simultaneous nitrification and denitrification (SND). The metagenomic sequencing results pointed out that the abundance of SCND-related genes in the granules was 0.18 %, and the abundance of Anammox-related genes was 0.02 %, which indicated that the granules mainly reduced nitrogen via SCND, and minor nitrogen was removed through Anammox. Inside the flocs, nitrogen was mainly reduced via the unconventional SND pathway (HAND). The in-depth analysis of the nitrogen removal process in the reactor with granules and flocs provides an important reference for further exploring the nitrogen removal mechanism, and supplies foundation for expanding the application of AGS.

Aerobic granular sludge for swine wastewater treatment: Implications for antibiotic and antibiotic resistance gene elimination

Swine wastewater (SW) contains high levels of traditional pollutants, antibiotics, and antibiotic resistance genes (ARGs), necessitating effective elimination. Two parallel aerobic granular sludge (AGS) reactors, R1 and R2, were constructed and optimized for treating SW from two pig farms, identified as SW1 and SW2. R2 showed higher antibiotic removal efficiency, particularly in the removal of sulfonamides, while fluoroquinolones tended to adsorb onto the sludge. Process optimization by introducing an additional anoxic phase enhanced denitrification and reduced effluent ARG levels, also aiding in the improved removal of fluoroquinolones. The nitrite-oxidizing bacteria (NOB) Nitrospira accumulated after the treatment process, reaching 12.8 % in R1 and 14.1 % in R2, respectively. Mantel’s test revealed that pH, NH4+-N, and Mg significantly affected ARGs and microbial community. Sulfadiazine and sulfamethazine were found to significantly impact ARGs and the microbial communities. This study provides innovative insights into the application of AGS for the treatment of real SW.

Influence of residual anaerobic granular sludge (AnGS) from anaerobically digested molasses wastewater in aerobic granular sludge reactor

This study investigated the impact of residual anaerobic granular sludge (AnGS) from anaerobic digesters treating molasses wastewater on ammonium reduction in a downstream aerobic granular sludge (AGS) reactor. Two conditions were tested: raw (high AnGS concentration) and settled (low AnGS concentration) anaerobically digested molasses wastewaters were fed into the AGS reactor. With the introduction of raw wastewater, enhanced nitrite accumulation at 30 % and improved total inorganic nitrogen (TIN) removal at 11 % were observed compared to 1 % nitrite accumulation and 8 % TIN removal with the introduction of settled wastewater. However, AnGS adversely affected other aspects of reactor performance, increasing effluent solid content and decreasing soluble chemical oxygen demand removal efficiency from 20 % in the low AnGS condition to 11 % in the high AnGS condition. Despite the observed retention of AnGS in the reactor, no significant bioaugmentation effects on the microbial community of the AGS were observed. Aerobic granular sludge was consistently observed in both conditions. The study suggests that AnGS may act as a nucleus for granule formation, helping to maintain granule stability in a disturbed environment. This study offers a systematic understanding of the impact of AnGS on subsequent nitrogen removal process using AGS, aiding in the decision making in the treatment of high solid anaerobic digestate.

Elucidating the role of organic loading rate on the performance and microbial dynamics of a novel continuous-flow aerobic granular sludge reactor

Aerobic granular sludge technology operated in continuous-flow reactors enables the upgrading of existing conventional wastewater treatment plants. The viability of applying this technology at full-scale increases with the simplicity of the bioreactor design. Hence, in contrast to previous research carried out in complex configurations, this study is based on evaluating the capacity of a novel single-chamber continuous-flow bioreactor to operate at different loads found in domestic sewage, highlighting that this configuration combines and joins the advantages of activated sludge and granular sludge technologies. At organic loading rates ranging from 0.45 to 1.85 kg COD•m−3•d−1, the reactors achieved high performance, removing >80 % of the chemical oxygen demand, regardless influent characteristics. The trend of ammonium removal rates was risen over operational time. Moreover, the granular size and biomass concentration increased with the organic loading rate. Changes in the influent did not compromise the structure of the granules. Microbiota studies showed a high diversity in communities at lower organic loading rates, whereas a strong microbial competition was detected at high loading rates, linking with lower evenness. Phylotypes belonging to Hypocreales played a key role in mature granules.

Treatment of high ammonia anaerobically digested molasses wastewater using aerobic granular sludge reactor

This study addressed the treatment of high ammonia, low biodegradable chemical oxygen demand (bCOD) anaerobically digested molasses wastewater, utilizing an aerobic granular sludge (AGS) reactor. The AGS achieved 99 % ammonia removal regardless of the bCOD supplementation. By adding low ammonia (<60 mg/L), high bCOD raw molasses wastewater (before anaerobic digestion) as a carbon source, enhanced nitrogen removal, increasing from 10 % to 97 %, and improved sludge settleability via bio-induced calcite precipitation were observed. Functional genes prediction suggested two potential denitrification pathways, including heterotrophic denitrification by Paracoccus and Thauera, and autotrophic denitrification, specifically sulfide-oxidizing autotrophic denitrification by Thiobacillus. An increase in the relative abundance of microorganisms involved in heterotrophic denitrification was observed with the addition of high bCOD raw molasses wastewater. Consequently, incorporating raw molasses wastewater into the AGS presents a sustainable approach to achieve mixotrophic denitrification, maintain stable granular sludge and ensure stable treatment performance when treating anaerobically digested molasses wastewater.

Operational variables and microbial community dynamics affect granulation stability in continuous flow aerobic granular sludge reactors

Retrofitting wastewater treatment plants with continuous aerobic granular sludge reactors is a promising alternative to enhance treatment capacities and reduce footprint. This study investigates the main variables influencing granulation and microbial dynamics in two reactor configurations (25 L): stirred tanks in series (R1) and a plug-flow-like system (R2). Granule formation was achieved by increasing the organic loading rate (OLR) from 0.7 to 4.1 kg COD/(m3·d) and the up-flow velocity in the biomass selector from 1.4 to 6.9 m/h. However, irreversible granule destabilization occurred at day 68 for R1 and day 108 for R2. Principal component analysis and examination of food-to-microorganisms (F/M) ratio medians identified the F/M ratio as the primary variable associated with instability. Microbial analysis revealed that a high F/M ratio induced significant increases in the abundance of specific genera such as Arcobacter, Cloacibacterium, Rikenella, Aquaspirillum and Sphaerotillus, whose overgrowth may negatively impact granule stability. Based on these findings, maximum F/M ratio thresholds were obtained to establish operational conditions allowing the maintenance of stable aerobic granules on continuous flow reactor configurations.

Correlation between aeration time in aerobic granular sludge reactors with the production of bioactive polysaccharides and microbial communities

Bioactive polysaccharides (PSs) as useful resources have been extracted from floc sludge, but there is limited knowledge about those from granular sludge. To address this gap, three aerobic granular sludge (AGS) reactors (R1, R2 and R3) with various operating conditions were set up for PS production (R1PS, R2PS and R3PS). Reactor operated under medium aeration time of 117 min (R2) led to the highest yield of PS (10.97 ± 0.34 %, g/g). The carbohydrate, protein and sulphate contents for all PSs ranged between 43.2–46.4 %, 0.82–1.52 %, and 2.23–24.12 %, respectively. All PSs were mainly heterogenous heteroglycans with different degree of sulphation. Four antioxidative activities were evaluated with R1PS having the most prominent effect in scavenging DPPH, ABTS and hydroxy radicals, while R2PS had the strongest reducing power. Major microbial communities found in the reactors at the phylum level were Proteobacteria, Bacteroidota, and Firmicutes. Correlation analysis revealed the relationships among aeration time, bacteria, and polysaccharide properties. Person correlation method was used to construct two complete linked pathways (aeration time->microbiota->sludge/effluent wastewater qualities and aeration time->microbiota->PS composition->PS antioxidant property) with two cutoffs (p-value < 0.05 and R2 > 0.8), and 35 microbial genera, 3 wastewater and sludge properties (sludge particle diameter, nitrite and nitrate) and 6 polysaccharide properties (glucose, galactose, fructose, sulphate content, hydroxyl scavenging radical and reducing power) were retained. These results facilitate the understanding of the relationship between the cultivation conditions in the AGS reactors, sludge granulation, PS properties and bacterial profiles for the optimal production of PS and their application as resource of bioactive substances.

Response of extracellular polymeric substances to high load shock in an aerobic granular sludge reactor performing simultaneous heterotrophic nitrification-aerobic denitrification

Response of extracellular polymeric substances to high load shock in an aerobic granular sludge reactor performing simultaneous heterotrophic nitrification-aerobic denitrification

Tire materials disturb transformations of nitrogen compounds and affect the structure of biomass in aerobic granular sludge reactors

Tire materials (TMs) present a notable hazard due to their potential to release harmful chemicals and microplastics into the environment. They can infiltrate wastewater treatment plants, where their effects remain inadequately understood, raising concerns regarding their influence on treatment procedures. Thus, this study investigated the impact of TMs in wastewater (10, 25, 50 mg/L) on wastewater treatment efficiency, biomass morphology, and microbial composition in aerobic granular sludge (AGS) reactors. TM dosage negatively correlated with nitrification and denitrification efficiencies, reducing overall nitrogen removal, but did not affect the efficiency of chemical-oxygen-demand removal. The presence of TMs increased the diameter of the granules due to TM incorporation into the biomass. The most frequently leached additives from TMs were N-(1,3-dimethylbutyl)-N′-phenyl-1,4-phenylenediamine, benzothiazole (BTH), and 2-hydroxybenzothiazole. In the treated wastewater, only BTH and aniline were detected in higher concentrations, which indicates that tire additives were biodegraded by AGS. The microbial community within the AGS adapted to TMs and their chemicals, highlighting the potential for efficient degradation of tire additives by bacteria belonging to the genera Rubrivivax, Ferruginibacter, and Xanthomonas. Additionally, our research underscores AGS's ability to incorporate TMs into biomass and effectively biodegrade tire additives, offering a promising solution for addressing environmental concerns related to TMs.

Granular stability, nitrogen and phosphorus removal pathways of aerobic granular sludge treating real municipal wastewater at different temperatures

Variations in temperature (T) can affect the activity of microorganisms in aerobic granular sludge (AGS) and biological wastewater treatment. Seasonal or diurnal fluctuations specifically low T can affect the overall wastewater treatment performance. Here, the short-term response of AGS to variation in T was investigated in terms of granular stability, sewage treatment and biological nutrient removal (BNR) pathways. AGS cultivated under tropical climate at T ≥ 30 ℃ was incubated at 15–25 ℃ by decreasing either in 10–15 °C steps or in 5 °C steps. Ammonium (74 % at 15 ℃; 94 % at 20 ℃), total inorganic nitrogen (46 % at 15 ℃; 50 % at 20 ℃), and phosphorus (38 % at 15 ℃; 51 % at 20 ℃) removals were significantly lower at 15 ℃ and 20 ℃ than at 30 ℃ (ammonium: >99 %; total inorganic nitrogen: 99 %; phosphorus: 76 %). The effect of lower temperature on BNR was less prominent when T was gradually decreased in 5 ℃ steps from 30 to 15 ℃. Moreover, the impact of lower temperature was quickly recovered upon reverting to 30 ℃. Ammonium was removed by partial nitrification and denitritation pathway. While, phosphorus was removed through enhanced biological phosphorus removal (EBPR) via polyphosphate accumulating organisms (PAOs). About 2–4-fold difference in the gene copies of ammonium oxidizing bacteria (AOB), denitrifying bacteria, and PAOs between 5 ℃ and 10 ℃−15 ℃ changes, respectively, explained operation of partial nitrification-denitritation and EBPR mechanisms. This study shows that a gradual decrease in T from 30° to 15°C in 5 ℃ steps can avoid inhibition of BNR pathways in AGS reactors. The granules were found to be stable with excellent settling properties. However, the sludge was dominated by smaller granules at lower temperatures of 15 ℃ and 20 ℃ as compared to 30 ℃ warranting long-term studies for evaluating the potential impact of T on AGS process.

Efficient removal of nitrogen and organic matter strategy from landfill leachate under high seasonal substrate variations

Landfill leachate is commonly used as a co-substrate in aerobic granular sludge (AGS) systems due to its high nitrogen concentration and variable organic matter content, which have a substantial impact on the removal efficiency. Typically, the nutrient content and biodegradability of leachate are categorized based on landfill age, often overlooking important factors such as seasonal variations. To address this limitation, a two-step process consisting of an anaerobic digestion (AD) reactor followed by an AGS reactor was evaluated over 431 days. Landfill leachate from the winter and spring seasons was chosen for its favorable biodegradability characteristics.Raw landfill leachate assays revealed a biodegradability range of 25%–92% within a single year, with higher values observed during winter compared to summer. To effectively handle the nutritional variations of landfill leachate, the hydraulic retention time (HRT) emerged as a crucial parameter influencing the performance of the reactors. When the AD reactor was operated in a hydrolytic configuration with an OLR of 68.4 g COD L−1 d−1, the proposed system showed 76.9% and 78.3% of organic matter and nitrogen removals, respectively. Best performance was obtained with COD/N 6.30 in the influent, ensuring required conditions for anoxic reactions (denitrification) at the center of the AGS.The proposed system offers valuable design tools for effectively removing nitrogen and organic matter from landfill leachates, despite significant seasonal nutrient variations. This approach can also be applied to wastewater with organic and nitrogen variations to face organic matter fluctuations in the AD and nitrogen fluctuations in the AGS.

Effect of influent ammonia nitrogen concentration on the phosphorus removal process in the aerobic granular sludge reactor

The effects of different influent ammonia nitrogen (NH3-N) concentrations (stage I to IV: 20, 40, 60 and 80 mg/L) on the phosphorus (P) removal process were studied in an aerobic granular sludge (AGS) reactor under anaerobic/aerobic operating conditions. The phosphorous removal performance kept stable at first (stage I and II) and then significantly deteriorated at further increase of influent NH3-N concentration (stage III and IV). On the process of microbial metabolism, further increase of NH3-N aggravated the competition between phosphate accumulating organisms (PAOs) and glycogen accumulating organisms (GAOs) for carbon sources in the anaerobic phase, and weakened the P absorption capacity of PAOs in the aerobic phase. Denitrifying phosphate accumulating organisms (DPAOs) showed higher resistance to the increase of NH3-N concentration than traditional PAOs. On the functional structure of the system, microorganisms related to phosphorus removal in the system were mainly enriched in the granules, while microorganisms with heterotrophic denitrifying function dominated the sludge flocs to obtain higher growth advantage at high ammonia nitrogen concentration. The results of this study provide more insight into the biological stability of AGS systems.

Pilot-scale aerobic granular sludge reactors with granular activated carbon for effective nitrogen and phosphorus removal from domestic wastewater

Aerobic granular sludge (AGS) is a breakthrough biotechnology of 21st century and an innovative alternative to activated sludge for treating wastewater. Concerns on long-start up periods for development of AGS and stability of granules are impeding its widespread implementation for treating low-strength domestic wastewater especially in tropical climate conditions. Addition of nucleating agents have been shown to improve development of AGS while treating low-strength wastewaters. There are no previous studies on AGS development and biological nutrient removal (BNR) in the presence of nucleating agents during treatment of real domestic wastewater. This study investigated AGS formation and BNR pathways while treating real domestic wastewater in a 2 m3 pilot-scale granular sequencing batch reactor (gSBR) operated without and with granular activated carbon (GAC) particles. The gSBRs were operated under tropical climate (T ≈ 30 °C) for >4-years to evaluate the effect of GAC addition on granulation, granular stability and BNR at pilot-scale. Formation of granules was observed within 3 months. MLSS values of 4 and 8 g/L were recorded within 6 months in gSBRs without and with GAC particles, respectively. The granules had an average size of 1.2 mm and SVI5 of 22 mL/g. Ammonium was mainly removed through nitrate formation in the gSBR without GAC. But, ammonium was removed by short-cut nitrification via nitrite due to washout of nitrite oxidizing bacteria in the presence of GAC. Phosphorus removal was much higher in gSBR with GAC due to the establishment of enhanced biological phosphorus removal (EBPR) pathway. After 3 months, the phosphorus removal efficiencies were at 15 % and 75 %, respectively, without and with GAC particles. The addition of GAC led to moderation in bacterial community and enrichment of polyphosphate-accumulating organisms. This is the first ever report on pilot-scale demonstration of AGS technology in the Indian sub-continent and GAC addition on BNR pathways.

Coupling physical selection with biological selection for the startup of a pilot-scale, continuous flow, aerobic granular sludge reactor without treatment interruption

This study removes two technical constraints for transitioning full-scale activated sludge infrastructure to continuous flow, aerobic granular sludge (AGS) facilities. The first of these is the loss of treatment capacity as a result of the rapid washout of flocculent sludge inventory and in turn the potential loss of nitrification during initial AGS reactor startup. The second is the physical selector design which currently is limited to either the complex sequencing batch reactor selection or sidestream hydrocyclones. Briefly, real wastewater data collected from this study suggested that by increasing the surface overflow rate (SOR) of an upflow clarifier to 10 m h −1, the clarifier can be taken advantage of as a physical selector to separate flocculant sludge from AGS. Redirecting the physical selector underflow and overflow sludge to the feast and famine zones of a treatment train, respectively, can create a biological selection that not only promotes AGS formation but also safeguards the effluent quality throughout the AGS reactor startup period. This study provides a novel concept for economically implementing continuous flow AGS within existing full-scale, continuous flow treatment trains.

Description of new single-chamber continuous-flow reactors of aerobic granular sludge: Technical and biological study

Aerobic granular sludge reactors usually operate in sequential batch mode, although this configuration limits the treatment of large volumes of wastewater, and they require a storage system. To implement this technology at full-scale, it would be necessary to design a simple and compact continuous-flow bioreactor able to treat larger volumes of wastewater. In this study, four aerobic granular sludge single-chamber continuous-flow reactors (R1, R2, R3 and R4) were designed and operated at the lab scale to evaluate and select a bioreactor configuration that achieves high organic matter removal performance while maintaining a stable granulation for long-term operation, promoting the washout of filamentous microorganisms. Results confirmed that the bioreactor including a lateral decanter (R1), was able to work in a steady state without loss of granular biomass and reached 95% of organic matter removal performance. Its granules had excellent compaction, with settling velocity values above 100 m·h−1. The R1 bioreactor also allowed rapid biomass adaptation and therefore a fast start-up (11 days). Results of this preliminary study at the lab scale suggested that the new and simple bioreactor design could be promising for its implementation at full scale. For that, future research is required to optimise the current model and to determine the most suitable operational parameters to treat domestic wastewater at full scale.

Zero‐dimensional modelling and bacterial characterization of an aerobic granular sludge reactor

AbstractBackgroundAerobic granular sludge (AGS) has emerged as a novel wastewater treatment technology and as a suppressing alternative to conventional activated sludge. The development of mathematical models of the AGS process, which can be used for wastewater treatment plant (WWTP) design and optimization, is therefore necessary to support successful implementation of AGS technology. This study aims to develop a zero‐dimensional (0D) AGS model that can be used in engineering practice for the above‐mentioned purposes.ResultsA laboratory AGS sequencing batch reactor (SBR), removing soluble organic substrate, nitrogen and phosphorus, was fed with artificial wastewater and modelled using a 0D approach. Model development was supported by bacterial characterization using 16S rRNA gene amplicon high‐throughput sequencing. The mathematical model was based on the activated sludge model no. 2d (ASM2d) and extended with both two‐step nitrification as implemented in the wastewater treatment plant simulator Sumo19 (Dynamita SARL) and ammonia nitrogen adsorption and desorption according to the Langmuir model. The variations of ammonia nitrogen, nitrite nitrogen, nitrate nitrogen, orthophosphate phosphorus and dissolved oxygen concentrations during one cycle of the SBR were successfully reproduced by the model (all Pearson's r values ≥0.93 and all r2 ≥ 0.86).ConclusionsThe 0D modelling approach was proven to be applicable to AGS. It is possible that the 0D approach can fill the gap that has developed between engineering and research in the biofilm modelling community. The 0D modelling approach therefore merits further exploration using reactors fed with real wastewater, as well as on pilot and full‐scale WWTPs. © 2022 Society of Chemical Industry (SCI).

Responses of various carbon to nitrogen ratios to microbial communities, kinetics, and nitrogen metabolic pathways in aerobic granular sludge reactor

The role of different ammonia concentrations (mg N/L) (of 100 (carbon to nitrogen ratio (C/N) = 12; Stage I), 200 (C/N = 6; Stage II), 400 (C/N = 3; Stage III) and 200 (C/N = 6; Stage IV)) in nitrogen metabolic pathways, microbial community, and specific microbial activity were investigated in an aerobic granular sludge reactor. Heterotrophic ammonia oxidizing bacteria (HAOB) showed higher ammonia oxidation rates (AORs) than autotrophic ammonia oxidizing bacteria (AAOB) at higher C/N conditions (Stages I and II). Paracoccus was the dominant HAOB. AAOB, with only 0.2–0.3 % in relative abundance, showed 2.7-fold higher AORs than HAOB at elevated ammonia and free ammonia (FA) concentrations with C/N at 3. Nitrosomonas and a genus in Nitrosomondaceae family were the major AAOB. This study proposed that FA inhibition on heterotrophic bacteria might be the mechanism that contributes to the development of the autotrophic ammonia oxidation pathway and enrichment of AAOB.

Oxygen transfer efficiency in an aerobic granular sludge reactor: Dynamics and influencing factors of alpha

In the pursuit of reducing carbon footprint and in view of increasing energy prices, energy efficiency is more important than ever before. Batch-wise operated aerobic granular sludge reactors consume up to 50% less energy compared to conventional activated sludge systems because pumping energy is reduced and mixing equipment is not needed. Further energy reduction efforts should therefore target aeration energy requirements. The alpha factor is an important factor influencing the oxygen transfer efficiency, however the dynamic behaviour of alpha has hardly been investigated in general and never for an aerobic granular sludge reactor. This study showed that alpha increases during the aeration phase of a cycle due to the influence of different process parameters. Through a data analysis study of 175 batch cycles of the Prototype Nereda® installation in Utrecht over the summer and winter period of 2020–2021, the exchange ratio and temperature were identified as the main influencing factors on the rate of increase of alpha in a batch cycle. A higher exchange ratio was related to a slower increase in alpha over the aeration phase, while a higher temperature was related to a faster increase in alpha. Moreover, alpha was characterized by a same minimal value at the beginning of every aeration phase, which could be explained by the adsorption of soluble biodegradable organic carbon described by a Langmuir adsorption model. Two mathematical models, a decreasing exponential and a first order model, were set up to unravel the dynamic behaviour of alpha. Both models were discussed in view of their practical implications for the design and performance optimization of aerobic granular sludge reactors and other batch-wise operated aerobic wastewater treatment systems.

Impact of seed sludge characteristics on granulation and performance of aerobic granular sludge process

Characteristics of seed sludge are of great importance in granulation and treatment performance of aerobic granular sludge process. Fast-settling floccular sludge has been shown to be affective on accelerating granulation. High-rate activated sludge process provides organic matter capturing from municipal wastewaters in order to harvest the chemical energy through adsorption rather than mineralization. Thus, waste sludge from the high-rate activated sludge process settles fast since the biosorption is the main mechanism in this technology. In this paper, impact of different seed sludge characteristics on granulation, and treatment performance of an aerobic granular sludge system was comparatively evaluated by seeding the system with solely conventional waste activated sludge (Stage 1) and a mixture of waste sludge from a conventional activated sludge process and a high-rate activated sludge process (Stage 2). Although the aerobic granular sludge reactor was seeded with a sludge that had a higher median particle size at Stage 2, after steady state conditions have been reached during the operation, average median particle size in the reactor at Stage 1 (124 ± 2.8 μm) was higher than that at Stage 2 (86 ± 2.7 μm). High ammonium nitrogen removal efficiencies (>98%) obtained in the study, however, total nitrogen (TN) and total phosphorus (TP) removal efficiencies obtained at Stage 2 (TN: 66.7 ± 2.2%; TP: 13.2 ± 2.9%) were lower those at Stage 1 (TN: 80.9 ± 2.2%; TP: 92.7 ± 4.4%). One of the reasons of incomplete denitrification and decreased phosphorus removal efficiencies observed at Stage 2 was smaller granules that was the cause of less anoxic/anaerobic volume. Another possible reason was the absence of denitrifiers, and polyphosphate-accumulating organisms in the high-rate activated sludge. Although high efficiencies were achieved in COD and ammonia removal by aerobic granular sludge process seeded with different types of sludge, this study showed that seed sludge affected anoxic/anaerobic volumes in the granules and thus affected denitrification and phosphate accumulation. Nitrogen and phosphorus loads resulting from wastewater treatment plants must be controlled and decreased before their discharge into the receiving water bodies. From this perspective, this study highlights the importance of seed sludge type on nitrogen and phosphorus removal performance of aerobic granular sludge process.

Hydrolysis capacity of different sized granules in a full-scale aerobic granular sludge (AGS) reactor

In aerobic granular sludge (AGS) reactors, granules of different sizes coexist in a single reactor. Their differences in settling behaviour cause stratification in the settled granule bed. In combination with substrate concentration gradients over the reactor height during the anaerobic plug-flow feeding regime, this can result in functional differences between granule sizes. In this study, we compared the hydrolytic activity in granules of 4 size ranges (between 0.5 and 4.8 mm diameter) collected from a full-scale AGS installation. Protease and amylase activities were quantified through fluorescent activity assays. To visualise where the hydrolytic active zones were located within the granules, the hydrolysis sites were visualized microscopically after incubating intact and sliced granules with fluorescent casein and starch. The microbial community was studied using fluorescent in situ hybridization (FISH) and sequencing. The results of these assays indicated that hydrolytic capacity was present throughout the granules, but the hydrolysis of bulk substrates was restricted to the outer 100 µm, approximately. Many of the microorganisms studied by FISH, such as polyphosphate and glycogen accumulating organisms (PAO and GAO), were abundant in the vicinity of the hydrolytically active sites. The biomass-specific hydrolysis rate depended mainly on the available granule surface area, suggesting that different sized granules are not differentiated in terms of hydrolytic capacity. Thus, the substrate concentration gradients that are present during the anaerobic feeding in AGS reactors do not seem to affect hydrolytic activity at the granule surfaces. In this paper, we discuss the possible reasons for this and reflect about the implications for AGS technology.