Effect of iron ions (Fe<Superscript>2+</Superscript>, Fe<Superscript>3+</Superscript>) on the formation and structure of aerobic granular sludge

Aerobic granulation is a promising technology for wastewater treatment, but problems regarding its formation and stability need to be solved. Divalent metal ions, especially Ca²⁺, Mg²⁺ and Mn²⁺, have been demonstrated to play an important role in the process of aerobic granulation. Here, we studied whether iron ions can affect aerobic granulation. Granular sludge formed without iron ion addition (<0.02 mg Fe²⁺ L^?1) was fluffy and had a finger-type structure and filamentous out-growth. The addition of iron ions to concentrations of 1 and 10 mg Fe²⁺ L^?1 repressed the finger-type structure and filamentous out-growth. The results show that chemical precipitation in the granules with iron ion addition was higher than that in the granules without ferrous addition. The amount of precipitates was higher inside the granules than outside. This study demonstrates that iron ions (Fe²⁺/Fe³⁺) increase the size and stability of aerobic granular sludge but do not affect the granulation time, which is the time that the first granular sludge is observed. The study shows that aerobic granular sludge technology can be confidently applied to actual wastewater containing a high concentration of iron compounds.     

Treatment of old landfill leachate with high ammonium content using aerobic granular sludge

Background

Aerobic granular sludge has become an attractive alternative to the conventional activated sludge due to its high settling velocity, compact structure, and higher tolerance to toxic substances and adverse conditions. Aerobic granular sludge process has been studied intensively in the treatment of municipal and industrial wastewater. However, information on leachate treatment using aerobic granular sludge is very limited.

Methods

This study investigated the treatment performance of old landfill leachate with different levels of ammonium using two aerobic sequencing batch reactors (SBR): an activated sludge SBR (ASBR) and a granular sludge SBR (GSBR). Aerobic granules were successfully developed using old leachate with low ammonium concentration (136 mg L^?1 NH₄ ⁺-N).

Results

The GSBR obtained a stable chemical oxygen demand (COD) removal of 70% after 15 days of operation; while the ASBR required a start-up of at least 30 days and obtained unstable COD removal varying from 38 to 70%. Ammonium concentration was gradually increased in both reactors. Increasing influent ammonium concentration to 225 mg L^?1 N, the GSBR removed 73 ± 8% of COD; while COD removal of the ASBR was 59 ± 9%. The GSBR was also more efficient than the ASBR for nitrogen removal. The granular sludge could adapt to the increasing concentrations of ammonium, achieving 95 ± 7% removal efficiency at a maximum influent concentration of 465 mg L^?1 N. Ammonium removal of 96 ± 5% was obtained by the ASBR when it was fed with a maximum of 217 mg L^?1 NH₄ ⁺-N. However, the ASBR was partially inhibited by free-ammonia and nitrite accumulation rate increased up to 85%. Free-nitrous acid and the low biodegradability of organic carbon were likely the main factors affecting phosphorus removal.

Conclusion

The results from this research suggested that aerobic granular sludge have advantage over activated sludge in leachate treatment.

     

Kinetic analysis on the two-step processes of AOB and NOB in aerobic nitrifying granules

Complete granulation of nitrifying sludge was achieved in a sequencing batch reactor. For the granular sludge, batch experiments were conducted to characterize the kinetic features of ammonia oxidizers (AOB) and nitrite oxidizers (NOB) in the granules using the respirometric method. A two-step nitrification model was established to determine the kinetic parameters of both AOB and NOB. In addition to nitrification reactions, the new model also took into account biomass maintenance and mass transfer through the granules. The yield coefficient, maximum specific growth rate, and affinity constant for ammonium for AOB were 0.21 g chemical oxygen demand (COD) g⁻¹ N, 0.09 h⁻¹, and 9.1 mg N L⁻¹, respectively, whereas the corresponding values for NOB were 0.05 g COD g⁻¹ N, 0.11 h⁻¹, and 4.85 mg N L⁻¹, respectively. The model developed in this study performed well in simulating the oxygen uptake rate and nitrogen conversion kinetics and in predicting the oxygen consumption of the AOB and NOB in aerobic granules.     

Antioxidative responses of Elodea nuttallii (Planch.) H. St. John to short-term iron exposure

Antioxidative responses of Elodea nuttallii (Planch.) H. St. John to short-term iron exposure were investigated in the study. Results showed that iron accumulation in E. nuttallii was concentration dependent. Growth of E. nuttallii was promoted by low iron concentration (1–10 mg L^?1 [Fe³⁺]), but growth inhibition was observed when iron concentration beyond 10 mg L^?1. The synthesis of protein and pigments increased within 1–10 mg L^?1 [Fe³⁺] range. The activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and glutathione-S-transferase (GST) were up to maximal values at 10 mg L^?1 [Fe³⁺]. High iron concentration inhibited the synthesis of protein and pigments as well as activities of antioxidative enzymes, and accelerated degradation of pigment and production of ROS. Low iron concentration had no significant influences on PSII maximal quantum yield, activity of PSII and relative electron transport rate though PSII. Malondialdehyde (MDA) and proline concentrations were highest at 100 and 1 mg L^?1 [Fe³⁺], respectively.     

Comparison of biological removal via nitrite with real-time control using aerobic granular sludge and flocculent activated sludge

The process of nitrification–denitrification via nitrite for nitrogen removal under real-time control mode was tested in two laboratory-scale sequencing batch reactors (SBRs) with flocculent activated sludge (R1) and aerobic granular sludge (R2) to compare operational performance and real-time control strategies. The results showed that the average ammonia nitrogen, total inorganic nitrogen (TIN), and chemical oxygen demand (COD) removal during aeration phase were 97.6%, 57.0%, and 90.1% in R2 compared with 98.6%, 48.7%, and 88.1% in R1. The TIN removed in both SBRs was partially due to the presence of simultaneous nitrification–denitrification via nitrite, especially in R2. The specific nitrification and denitrification rates in R2 were 0.0416 mgNH₄⁺–N/gSS-min and 0.1889 mgNO_X⁻–N/gSS-min, which were 1.48 times and 1.35 times that of R1. The higher rates for COD removal, nitrification, and denitrification were achieved in R2 than R1 with similar influent quality. Dissolved oxygen (DO), pH, and oxidization reduction potential, corresponding to nutrient variations, were used as diagnostic parameters to control the organic carbon degradation and nitrification–denitrification via nitrite processes in both SBRs. The online control strategy of granular SBR was similar to that of the SBR with flocculent activated sludge. However, a unique U-type pattern on the DO curve in granular SBR was different from SBR with flocculent activated sludge in aerobic phase.     

Long-term starvation and subsequent reactivation of a high-rate partial nitrification activated sludge pilot plant

The starvation process of a high-rate partial nitrification system during 30 days and its controlled recovery were studied in an activated sludge pilot plant. Four ammonium-starved reactors under anoxic, aerobic and two different alternating aerobic/anoxic conditions were evaluated. The highest and the lowest decay rates of ammonia oxidizing bacteria (AOB) were obtained under full aerobic (0.24 d⁻¹) and full anoxic (0.11 d⁻¹) conditions, respectively. The evolution of biomass activity correlated well with the AOB quantification using FISH technique. AOB fractions lower than 1% were measured in the four reactors after 23 days of starvation. The recovery of the system was achieved in only 5 days using a nitrogen loading rate (NLR) control loop, obtaining the same conditions than before the long-term starvation period with a NLR of 1.2 g N L⁻¹ d⁻¹ and 98% of nitrite accumulation in the effluent.     

Phosphorus removal characteristics of granular and flocculent sludge in SBR

Aerobic granulation technology has become a novel biotechnology for wastewater treatment. However, the distinct properties and characteristics of phosphorus removal between granules and flocculent sludge are still sparse in enhanced biological phosphorus removal process. Two identical sequencing batch reactors (SBRs) were operated to compare phosphorus removal performance with granular sludge (R1) and flocculate activated sludge (R2). Results indicated that the start-up period was shorter in R2 than R1 for phosphorus removal, which made R2 reach the steady-state condition on day 21, while R1 was on day 25, and R2 released and took up more phosphorus than R1. As a result, the phosphorus removal was around 90% in R2 while 80% in R1 at the steady-state system. The special phosphorus release rate and special phosphorus uptake rate were 8.818 mg P/g volatile suspended solids (VSS)/h and 9.921 mg P/g VSS/h in R2, which were consistently greater than those (0.999 and 3.016 mg P/g VSS/h) in R1. The chemical oxygen demand removal in two reactors was similar. The granular SBR had better solid-separation performance and higher removal efficiency of NH₄⁺–N than flocculent SBR. Denaturing gradient gel electrophoresis of PCR-amplified 16S rDNA fragment analysis revealed that the diversity and the level of phosphorus-accumulating bacteria in flocculent sludge were much more than those in the granular sludge.     

Aerobic granulation and nitrogen removal with the effluent of internal circulation reactor in start-up of a pilot-scale sequencing batch reactor

Aerobic granular sludge was successfully cultivated with the effluent of internal circulation (IC) reactor in a pilot-scale sequencing batch reactor (SBR) using activated sludge as seeding sludge. N removal was investigated in the start-up of aerobic granulation process. Initially, the phenomenon of partial nitrification was observed and nitrite accumulation rates (NO₂ ^?-N/NO _x ^? -N) were between 84.6 and 99.1?%. It was potentially caused by ammonium oxidizing bacteria (AOB) in the seeding activated sludge, high external environmental temperature (~32?°C) and free ammonia (FA) concentration. After 50?days’ running, the aerobic granules-based bioreactor demonstrated perfect performance in simultaneous removal of organic matter and ammonia nitrogen, and average removal efficiencies were maintained above 93 and 96?%, respectively. The maximum nitrogen removal efficiency of 83.1?% was achieved after the formation of aerobic granules. The average diameter of mature aerobic granular sludge mostly ranged from 0.5 to 1.0?mm. Furthermore, one typical cyclic test indicated that pH and DO profiles could be used as effective parameters for biological reactions occurring in the aerobic/anoxic process. The obtained results could provide further information on the cultivation of aerobic granular sludge with practical wastewater, especially with regard to nitrogen-rich industrial wastewater.     

A key cultivation technology for denitrifying granular sludge

Well-formed denitrifying granular sludge with a biomass concentration of 24.8 gVSS L^?1 and a specific nitrate removal rate of 0.19 gNO₃-N gVSS^?1 d^?1 was obtained in an upflow sludge blanket (USB) reactor by cultivating seeded aerobic flocculent sludge for 6–8 weeks. Regularity phenomena exist in the granulation including flotation of flocculent sludge, formation of fine granules, occurrence of channelling, and formation of mature granular sludge. The granulation is similar to crystal growth, that the non-denitrifying bacteria evolve into the carriers (fine granules), on the surface of which denitrifying bacteria proliferate and develop into mature granular sludge.There are several key parameters that must be considered when developing a good denitrifying granular sludge. First, the proper seed sludge must be chosen (VSS/SS at 0.65–0.75, SRT over 25 days) to accelerate the granulation process. Secondly, any floating sludge should be stirred, and the sludge loading rate should be within the range of 0.05–0.15 gNO₃-N gVSS^?1 d^?1 until fine granules emerge. Additionally, spontaneous gas agitation or interval air-blowing should be used to effectively eliminate channelling; Finally, the sludge loading rate should be less than 0.25 gNO₃-N gVSS^?1 d^?1 until dense, mature granular sludge appears. This study could support and promote the full-scale application of denitrifying granular sludge.     

Process for biological oxidation and control of dissolved iron in bioleach liquors

Iron has a central role in bioleaching and biooxidation processes. Fe²⁺ produced in the dissolution of sulfidic minerals is re-oxidized to Fe³⁺ mostly by biological action in acid bioleaching processes. To control the concentration of iron in solution, it is important to precipitate the excess as part of the process circuit. In this study, a bioprocess was developed based on a fluidized-bed reactor (FBR) for Fe²⁺ oxidation coupled with a gravity settler for precipitative removal of ferric iron. Biological iron oxidation and partial removal of iron by precipitation from a barren heap leaching solution was optimized in relation to the performance and retention time (τ_FBR) of the FBR. The biofilm in the FBR was dominated by Leptospirillum ferriphilum and “Ferromicrobium acidiphilum.” The FBR was operated at pH 2.0 ± 0.2 and at 37 °C. The feed was a barren leach solution following metal recovery, with all iron in the ferrous form. 98–99% of the Fe²⁺ in the barren heap leaching solution was oxidized in the FBR at loading rates below 10 g Fe²⁺/L h (τ_FBR of 1 h). The optimal performance with the oxidation rate of 8.2 g Fe²⁺/L h was achieved at τ_FBR of 1 h. Below the τ_FBR of 1 h the oxygen mass transfer from air to liquid limited the iron oxidation rate. The precipitation of ferric iron ranged from 5% to 40%. The concurrent Fe²⁺ oxidation and partial precipitative iron removal was maximized at τ_FBR of 1.5 h, with Fe²⁺ oxidation rate of 5.1 g Fe²⁺/L h and Fe³⁺ precipitation rate of 25 mg Fe³⁺/L h, which corresponded to 37% iron removal. The precipitates had good settling properties as indicated by the sludge volume indices of 3–15 mL/g but this step needs additional characterization of the properties of the solids and optimization to maximize the precipitation and to manage sludge disposal.     

Aerobic granulation in sequencing batch reactors with different reactor height/diameter ratios

Effects of reactor height/diameter ratios ranged from 24 to 4 corresponding to reactor settling velocities from 12 to 2 m h^?1 on aerobic granulation were investigated. It was found that granules appeared after 1-week operation and granule volume percentages exceeded 50% after 2–3 weeks in four reactors. In addition, similar granule fraction of 94–96% was found at steady state in all four reactors. Sludge volume index (SVI), average sludge size, biomass density and granule settling velocity at steady state were around 50 ml g^?1, 1800 μm, 53 g l^?1 and 40 m h^?1, respectively, in four reactors. Extracellular polymeric substances (EPS) and specific oxygen uptake rate (SOUR) were around 38 mg g^?1 VSS and 40 mg O₂ g^?1 VSS h^?1, respectively. Denaturing gradient gel electrophoresis (DGGE) fingerprint of sludge in four reactors showed the same microbial population shift during the start-up period and same microbial community structure during steady-state period. These results recommended strongly that reactor height/diameter ratio or reactor setting velocity in the used range in this study did not affect granule formation, physical characteristics, microbial community structure of granules and stable operation of granular sludge reactor. Reactor height/diameter ratio thus can be very flexible in the practice, which is important for the application of aerobic granule technology.     

Kinetics of ferrous iron oxidation by batch and continuous cultures of thermoacidophilic Archaea at extremely low pH of 1.1–1.3

The extreme acid conditions required for scorodite (FeAsO₄·2H₂O) biomineralization (pH below 1.3) are suboptimal for growth of most thermoacidophilic Archaea. With the objective to develop a continuous process suitable for biomineral production, this research focuses on growth kinetics of thermoacidophilic Archaea at low pH conditions. Ferrous iron oxidation rates were determined in batch-cultures at pH 1.3 and a temperature of 75°C for Acidianus sulfidivorans, Metallosphaera prunea and a mixed Sulfolobus culture. Ferrous iron and CO₂ in air were added as sole energy and carbon source. The highest growth rate (0.066 h⁻¹) was found with the mixed Sulfolobus culture. Therefore, this culture was selected for further experiments. Growth was not stimulated by increase of the CO₂ concentration or by addition of sulphur as an additional energy source. In a CSTR operated at the suboptimal pH of 1.1, the maximum specific growth rate of the mixed culture was 0.022 h⁻¹, with ferrous iron oxidation rates of 1.5 g L⁻¹ d⁻¹. Compared to pH 1.3, growth rates were strongly reduced but the ferrous iron oxidation rate remained unaffected. Influent ferrous iron concentrations above 6 g L⁻¹ caused instability of Fe²⁺ oxidation, probably due to product (Fe³⁺) inhibition. Ferric-containing, nano-sized precipitates of K-jarosite were found on the cell surface. Continuous cultivation stimulated the formation of an exopolysaccharide-like substance. This indicates that biofilm formation may provide a means of biomass retention. Our findings showed that stable continuous cultivation of a mixed iron-oxidizing culture is feasible at the extreme conditions required for continuous biomineral formation.     

Aerobic granular sludge formation for high strength agro-based wastewater treatment

The present study investigates the formation of aerobic granular sludge in sequencing batch reactor (SBR) fed with palm oil mill effluent (POME). Stable granules were observed in the reactor with diameters between 2.0 and 4.0 mm at a chemical oxygen demand (COD) loading rate of 2.5 kg COD m⁻³ d⁻¹. The biomass concentration was 7600 mg L⁻¹ while the sludge volume index (SVI) was 31.3 mL g SS⁻¹ indicating good biomass accumulation in the reactor and good settling properties of granular sludge, respectively. COD and ammonia removals were achieved at a maximum of 91.1% and 97.6%, respectively while color removal averaged at only 38%. This study provides insights on the development and the capabilities of aerobic granular sludge in POME treatment.     

Control and optimization of nitrifying communities for nitritation from domestic wastewater at room temperatures

To achieve nitritation from complete-nitrification seed sludge at room temperature of 19 ± 1 °C, a lab-scale sequencing batch reactor (SBR) treating domestic wastewater with low C/N ratios was operated to investigate the control and optimization of nitrifying communities. Ammonia oxidizing bacteria (AOB) dominance was enhanced through the combination of low DO concentrations (<1.0 mg/L) and preset short-cycle control of aeration time. Nitritation was successfully established with NO₂^?-N/NO_x^?-N over 95%. To avoid the adverse impact of low DO concentrations on AOB activities, DO concentrations were increased to 1–2 mg/L. At the normal DO levels and temperatures, on-line control strategy of aerobic durations maintained the stability of nitritation with nitrite accumulation rate over 95% and ammonia removal above 97%. Fluorescence in-situ hybridization (FISH) analysis presented that the maximal percentage of AOB in biomass reached 10.9% and nitrite oxidizing bacteria (NOB) were washed out.     

Nitrifying granules cultivation in a sequencing batch reactor at a low organics-to-total nitrogen ratio in wastewater

It is possible to cultivate aerobic granular sludge at a low organic loading rate and organics-to-total nitrogen (COD/N) ratio in wastewater in the reactor with typical geometry (height/diameter = 2.1, superficial air velocity = 6 mm/s). The noted nitrification efficiency was very high (99%). At the highest applied ammonia load (0.3 ± 0.002 mg NH₄⁺–N g total suspended solids (TSS)⁻¹ day⁻¹, COD/N = 1), the dominating oxidized form of nitrogen was nitrite. Despite a constant aeration in the reactor, denitrification occurred in the structure of granules. Applied molecular techniques allowed the changes in the ammonia-oxidizing bacteria (AOB) community in granular sludge to be tracked. The major factor influencing AOB number and species composition was ammonia load. At the ammonia load of 0.3 ± 0.002 mg NH₄⁺–N g TSS⁻¹ day⁻¹, a highly diverse AOB community covering bacteria belonging to both the Nitrosospira and Nitrosomonas genera accounted for ca. 40% of the total bacteria in the biomass.     

Anaerobic granule development for removal of pentachlorophenol in an upflow anaerobic sludge blanket (UASB) reactor

The granulation process using synthetic wastewater containing pentachlorophenol (PCP) in four 1.1 l laboratory scale upflow anaerobic sludge blanket (UASB) reactors was studied, and the anaerobic biotransformation of PCP during the granulation process investigated. After 110 days granular sludge was developed and up to 160 and 180 mg/l of PCP was added into the reactors R1 and R2, respectively, when they were inoculated with acclimated anaerobic sludge from an anaerobic digester of a citric acid plant. The inoculum was predominately composed of bacilli and filamentous bacteria. Granulation did not occur in reactors R3 and R4 which were inoculated with acclimated anaerobic sludge from aerobic sludge of the municipal sewage treatment plant which consisted mainly of cocci. Despite similar bacilli in the granule, the filamentous bacteria from reactor R1 were thicker than those of reactor R2. The granular sludge had a maximum diameter of 2.5 and 2.2 mm, and SMA of 1.44 and 1.32 gCOD/gTVS per day for reactors R1 and R2, respectively. Over 98% chemical oxygen demand (COD) removal rate and 99% of PCP removal rate were achieved when reactors R1 and R2 were operated at PCP and COD loading rates of 150 and 7.5 g/l per day, respectively. H₂-producing acetogens were the dominant anaerobes in the granular sludge.     

Treatment of dairy effluents in an aerobic granular sludge sequencing batch reactor

Aerobic granular sludge can successfully be cultivated in a sequencing batch reactor (SBR) treating dairy wastewater. Attention has to be paid to the fact that suspended solids are always present in the effluent of aerobic granular sludge reactors, making a post-treatment step necessary. Sufficient post-treatment can be achieved through a sedimentation process with a hydraulic retention time of 15–30 min. After complete granulation and the separation of biomass from the effluent, removal efficiencies of 90% COD_total, 80% N_total and 67% P_total can be achieved at a volumetric exchange ratio of 50% and a cycle duration of 8 h. Effluent values stabilize at around 125 mg l^–1 COD_dissolved. The maximum applicable loading rate is nevertheless limited, as the stability of aerobic granules very much depends on the presence of distinct feast and famine conditions and the degradation of real wastewaters shows slower kinetics compared with synthetic wastewaters. As loading rate and volumetric exchange ratio are coupled in an SBR system, the potential of granular sludge for improving process efficiency is also limited.     

Change in ammonia-oxidizing microorganisms in enriched nitrifying activated sludge

In this study, sludge was taken from a municipal wastewater treatment plant that contained a nearly equal number of archaeal amoA genes (5.70 × 10⁶ ± 3.30 × 10⁵ copies mg sludge⁻¹) to bacterial amoA genes (8.60 × 10⁶ ± 7.64 × 10⁵ copies mg sludge⁻¹) and enriched in three continuous-flow reactors receiving an inorganic medium containing different ammonium concentrations: 2, 10, and 30 mM NH₄⁺–N (28, 140, and 420 mg N l⁻¹). The abundance and communities of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in enriched nitrifying activated sludge (NAS) were monitored at days 60 and 360 of the operation. Early on, between day 0 and day 60 of reactor operation, comparative abundance of AOA amoA genes to AOB amoA genes varied among the reactors depending on the ammonium levels found in the reactors. As compared to the seed sludge, the number of AOA amoA genes was unchanged in the reactor with lower ammonium level (0.06 ± 0.04 mgN l⁻¹), while in the reactors with higher ammonium levels (0.51 ± 0.33 and 0.25 ± 0.10 mgN l⁻¹), the numbers of AOA amoA genes were deteriorated. By day 360, AOA disappeared from the ammonia-oxidizing consortiums in all reactors. The majority of the AOA sequences from all NASs at each sampling period fell into a single AOA cluster, however, suggesting that the ammonium did not affect the AOA communities under this operational condition. This result is contradictory to the case of AOB, where the communities varied significantly among the NASs. AOB with a high affinity for ammonia were present in the reactors with lower ammonium levels, whereas AOB with a low affinity to ammonia existed in the reactors with higher ammonium levels.     

Causes and control of filamentous growth in aerobic granular sludge sequencing batch reactors

Poor long-term stability of aerobic granules developed in sequencing batch reactors (SBRs) remains a limitation to widespread use of aerobic granulation in treating wastewater. Filamentous growth has been commonly reported in aerobic granular sludge SBR. This review attempts to address the instability problem of aerobic granular sludge SBR from the perspective of filamentous growth in the system. The possible causes of filamentous growth are identified, including long retention times of solids, low substrate concentration in the liquid phase, high substrate gradient within the granule, dissolved oxygen deficiency in the granule, nutrient deficiency inside granule, temperature shift and flow patterns. Because of cyclic operation of aerobic granular sludge SBR and peculiarities of aerobic granules, various stresses can be present simultaneously and can result in progressive development of filamentous growth in aerobic granular sludge SBR. Overgrowth of filamentous bacteria under stress conditions appears to be a major cause of instability of aerobic granular sludge SBR. Specific recommendations are made for controlling filamentous growth.