Excess Extracellular Polymeric Substances Research Articles

Elucidating biofouling development and succession in membrane distillation using treated effluent

Biofouling in membrane distillation (MD) has several repercussions, including reduced efficiency of the MD process and limiting membrane life. Additionally, the evaluation of MD biofouling using treated effluents from wastewater treatment plants remains an unexplored area. Thus, biofouling formation and development in a long term MD process (15 days) using treated effluent from a wastewater treatment plant was explored in this study. The results revealed that flux decline occurred in four phases: i) initial decline (0–1 d), ii) gradual decline (1–5 d), iii) progressive decline (5–10 d), and iv) rapid decline (10–15 d). Liquid Chromatography-Organic Carbon Detection (LC-OCD) analysis demonstrated that the treated effluent contained humic-like substances, which deposited on the membrane surface in phase 1. Whereas biopolymers development on the membrane surface in phase 2 and 3 was linked to biofouling. Microbial community analysis revealed that the initial colonisers were predominantly thermophilic bacteria, which were different from the microbial community of the treated effluent. The biofilm-forming bacteria included Schlegelella, Meiothermus, and Vulcaniibacterium. These microorganisms proliferate and release excessive extracellular polymeric substances (EPS), leading to the development of mature biofilm on membrane surface. This helped in the deposition of organics and inorganics from the bulk feed, which led to microbial community succession in phase 4 with the emergence of the Kallotenue genus. The results suggested that organic substances and microbial communities on membrane surface at different stages in a long-term MD process had a significant influence on MD performance for high-quality wastewater reuse.

The role of fungi in the formation and instability of halophilic aerobic granular sludge in a continuous flow reactor

The role of fungi in aerobic granular sludge remains unknown. This study explored the formation and instability of halophilic aerobic granular sludge (HAGS) induced by fungi in a continuous flow reactor (CFR) figured with an aeration tank and a two-zone sedimentation tank. HAGS with a particle size of 420.10 ± 1.13 μm was successfully cultivated by increased hydraulic shear force (HSF). Increasing HSF stimulated microorganisms such as unclassified_f_Rhodobacteraceae, Muricauda, and Raineyella to secrete more extracellular polymeric substances (EPS) with a high protein (PN) content (81.03 ± 2.65 %). The peeled filamentous fungal fragments could act as colonization carriers to connect microorganisms. The mature HAGS had excellent removal efficiency of chemical oxygen demand (COD; 96.20 ± 1.19 %) and ammonia (92.13 ± 5.36 %) from 20 g/L saline wastewater. However, HAGS was disintegrated due to the fast growth of fungi, mainly unclassified_f_Nectriaceae, resulting in the deterioration of the sludge settling performance (SVI5 = 499.05 ± 7.80 mL/g). Filamentous fungi can use EPS secreted by bacteria under anaerobic and aerobic conditions. The excess EPS consumption finally destroyed the EPS and bacteria network. This study revealed the role of fungi in the formation and instability of HAGS in CFR, providing the basis for maintaining long-term HAGS stability.

Synergistic effects of trace sulfadiazine and corrosion scales on disinfection by-product formation in bulk water of cast iron pipe

The effects of trace sulfadiazine (SDZ) and cast-iron corrosion scales on the disinfection by-product (DBP) formation in drinking water distribution systems (DWDSs) were investigated. The results show that under the synergistic effect of trace SDZ (10 μg/L) and magnetite (Fe3O4), higher DBP concentration occurred in the bulk water with the transmission and distribution of the drinking water. Microbial metabolism-related substances, one of the important DBP precursors, increased under the SDZ/Fe3O4 condition. It was found that Fe3O4 induced a faster microbial extracellular electron transport (EET) pathway, resulting in a higher microbial regrowth activity. On the other hand, the rate of chlorine consumption was quite high, and the enhanced microbial EET based on Fe3O4 eliminated the need for microorganisms to secrete excessive extracellular polymeric substances (EPS). More importantly, EPS could be continuously secreted due to the higher microbial activity. Finally, high reactivity between EPS and chlorine disinfectant resulted in the continuous formation of DBPs, higher chlorine consumption, and lower EPS content. Therefore, more attention should be paid to the trace antibiotics polluted water sources and cast-iron corrosion scale composition in the future. This study reveals the synergistic effects of trace antibiotics and corrosion scales on the DBP formation in DWDSs, which has important theoretical significance for the DBP control of tap water.

Effect of ultrasound on the stability of partial nitrification: Under the interference of aeration rate

The fluctuation of dissolved oxygen is one of the primary cause of disruptions to the consistent operation of partial nitrification, and the level of dissolved oxygen is mainly controlled by the aeration rate. This study investigated the influence of ultrasonic treatment on the stability of partial nitrification of activated sludge under different aeration conditions. After being treated with ultrasound (energy density = 0.20 W·mL−1, treatment time = 10 min), partial nitrification process operated stably for 67 days, with the nitrite accumulation rate above 83.89 %. The effluent contained 42.50 mg·L−1 of nitrite, much higher than the control reactor (0.30 mg·L−1). The gap between the specific ammonia and nitrite oxidation rates widened continuously as the aeration rate increased, and nitrite-oxidizing bacteria activity did not recover even under conditions with a very high oxygen content. Further analysis showed that ultrasonic treatment had obvious stripping effect on excess extracellular polymeric substances (EPS), especially loosely bound EPS and protein. Additionally, long-term ultrasonic treatment promoted the enrichment of Nitrosomonas and strongly inhibited Nitrotoga. Based on these findings, it appears that under conditions of high aeration rate, ultrasound effectively suppress the recovery of Nitrotoga activity and improve the stability of partial nitrification.

The influence of different nitrate concentrations on aerobic sludge granulation and the role of extracellular polymeric substances

This study investigated the influence of nitrate on aerobic granular sludge (AGS) granulation. The introduction of nitrate at 5, 15 and 20 mg L−1 promoted AGS granulation, and the promoting effect was positively correlated with nitrate concentrations. Meanwhile, exogenous nitrate significantly increased denitrification rate in the AGS system. However, granular disintegration appeared at a long-term addition of nitrate. An in-deep analysis showed that nitrate stimulated the secretion of extracellular polymeric substances (EPS), especially the content of proteins, which might be the main reason for the AGS granulation. However, the rapid and excessive increase in EPS might cause granular disintegration, as excessive EPS blocked the transmission of substrates, leading to the increase of dead cells in the granules. Besides, nitrate also altered the hydrophobicity of EPS and the content of α-helix, 3-turned helix and polymeric chain that favored aggregation, which also affected AGS granulation. From the microbial community level, nitrate induced the enrichment of denitrifying bacteria, including those that also functioned as EPS producers, such as Micropruina and Flavobacterium, resulting in the rapid increase of functional enzymes associated with amino acid synthesis, thereby promoting the secretion of proteins in EPS. Conversely, disintegration caused by mass transfer blockage might lead to the loss of EPS producing bacteria and subsequent decrease in EPS content, further accelerating granular disintegration.

The response of nitrifying activated sludge to chlorophenols: Insights from metabolism and redox homeostasis

The specialized wastewater treatment plants for the chemical industry are rapidly developed in China and many other countries. But there is a common bottleneck in that the toxic pollutants in chemical wastewater often cause shock impacts on biological nitrogen removal systems, which directly affects the stability and cost of operation. As the research on nitrification inhibition characteristics is not sufficient till now, there is a great lack of theoretical guidance on the control of the inhibition. This study investigated the response of nitrifying activated sludge to chlorophenols (CPs) inhibition in terms of metabolism disorder and oxidative stress. At the initial stage of reaction (i.e., 1 h), reactive oxygen species (ROS)-induced membrane damage which might account for declining nitrification performance. Simultaneously excessive extracellular polymeric substances (EPS) were secreted to alleviate oxidative stress injury and protected microorganisms to some extent. In particular tyrosine-like substances in LB-EPS with a Fmax increase of 242.30% were confirmed to efficiently resist phenols inhibition. Thus, as the inhibition proceeded, metabolism disorder replaced oxidative stress as the main cause of nitrification inhibition. The affected metabolic processes include weakened enzyme catalysis, restricted electron transport and lessened energy generation. At 4 h, nitrifying production of sludge amended with 5 mg/L chlorophenols was 89.27 ± 9.51%–98.15 ± 9.60% lower than blank, the inhibition could be attributed to comprehensively affected metabolism. The structural equation modeling indicated that phenols restricted nitrification enzymes and bacterial electron transport efficiency which was critical to nitrification performance. Moreover, the lessened energy generation weakens enzyme activity to further suppress nitrification. These findings enriched our knowledge of nitrifiers’ responses to CPs inhibition and provided the basis for addressing nitrification inhibition.

Performance and stability enhancement of methanogenic granular sludge process: Feed pre-acidification and magnetite-embedded granule formation

Recently, inducing the formation of conductive magnetite-embedded granules (MEGs) by self-embedding submicron magnetite particles into anaerobic granules has been demonstrated to be effective in enhancing methanation and improving sludge stability in upflow anaerobic sludge blanket (UASB) reactors. However, MEGs still suffer structural and functional destabilization at high organic loading rates (OLRs) due to the excessive secretion of extracellular polymeric substances (EPS), resulting in a significant washout of active microbes. This is particularly true when treating carbohydrate-rich wastewater that is prone to producing excessive EPS. In order to address this issue, this study explored the potential of combining MEG-UASB process with feed pre-acidification as a means of improving the methanogenic treatment performance and stability of whey, a carbohydrate-rich dairy effluent. Two UASB reactors, one with MEGs and one without, were operated to treat pre-acidified whey at increasing OLRs of 2–11 g chemical oxygen demand/L·d. Feed pre-acidification was effective in preventing granule destabilization and maintaining stable UASB performance with or without MEGs. Meanwhile, the reactor with MEGs (JM) showed significantly better performance and stability than the one without MEGs (JC), especially at high OLRs, which was likely because MEGs improved sludge retention and promoted electro-syntrophic methanogenic activity. Accordingly, JM sludge exhibited greater electron transfer capability and a more pronounced enrichment of electroactive microbes than JC sludge. External voltage application (0.6 V) did not relieve but rather aggravated the process upset under overloading conditions for both reactors. The overall results suggest that the MEG-UASB process coupled with pre-acidification holds promise for efficient and stable treatment of carbohydrate-rich wastewater.

The Role of Extracellular Polymeric Substances in the Toxicity Response of Anaerobic Granule Sludge to Different Metal Oxide Nanoparticles.

Wastewater treatment plants (WWTP) are regarded as the last barriers for the release of incompletely separated and recycled nanoparticles (NPs) into the environment. Despite the importance and ubiquity of microbial extracellular polymeric substances (EPSs) in the complex wastewater matrix, the interaction between NPs and EPSs of anaerobic microflora involved in wastewater treatment and the resultant impact on the biomass metabolomics are unclear. Thus, the impacts of different metal oxide (TiO2, ZnO, and CuO) NPs on functional bacteria in anaerobic granular sludge (AGS) and the possible toxicity mechanisms were investigated. In particular, the binding quality, enhanced resistance mechanism, and chemical fractional contribution of EPSs from AGS against the nanotoxicity of different NPs was assessed. The results showed that CuO NPs caused the most severe inhibition towards acetoclastic and hydrogenotrophic methanogens, followed by ZnO NPs, whereas TiO2 NPs caused no inhibition to methanogenesis. Excessive EPS production, especially the protein-like substances, was an effective strategy for reducing certain NPs’ toxicity by immobilizing NPs away from AGS cells, whereas the metabolism restriction on inner microorganisms of AGS induced by CuO NPs can deteriorate the protective role of EPS, indicating that the roles of EPS may not be amenable to generalizations. Further investigations with lactate dehydrogenase (LDH) and reactive oxygen species (ROS) assays indicated that there are greatly essential differences between the toxicity mechanisms of metal NPs to AGS, which varied depending on the NPs’ type and dosage. In addition, dynamic changes in the responses of EPS content to different NPs can result in a significant shift in methanogenic and acidogenic microbial communities. Thus, the production and composition of EPSs will be a key factor in determining the fate and potential effect of NPs in the complex biological matrix. In conclusion, this study broadens the understanding of the inhibition mechanisms of metal oxide NPs on the AGS process, and the influence of EPSs on the fate, behavior, and toxicity of NPs.

Biological responses to P‐limitation in indigenous bacteria isolated from drinking water

AbstractBiofilm formation in direct biofiltration causes operational issues such as accelerated head loss accumulation rate resulting in short filter runtime. Applying coagulant in low P level source water may impose bacteria to the P‐limited condition. It is hypothesized that the nutrient limitation, specifically P, as an essential macronutrient for bacteria causes stress and triggers excess extracellular polymeric substances (EPS). Two of the most biofilm‐producing bacteria were isolated from a full‐scale biofilter, provided the opportunity to study the indigenous biofilter bacteria. The isolated bacteria were identified using full‐length 16S rRNA and characterized. The biological behavior of the species was studied under different P‐limited conditions in a nutrient‐limited medium simulating freshwater nutrient availability. Carbohydrate and Protein‐EPS were increased when decreasing the available P in the medium, suggesting that lack of P can trigger higher EPS.Article Impact StatementThis study demonstrated that the lower P levels are directly related to excessive extracellular polymeric substances production and consequently the biofilter performance.

Effects of cations on biofilm formation and characteristics in integrated fixed film activated sludge process at different carbon and nitrogen loadings

Both divalent cations including calcium and magnesium play important roles for microbial aggregates in binding to negatively charged functional groups on bacterial surfaces, in extracellular polymeric substances (EPS), and on inorganic materials in flocs and biofilms. Monovalent cations such as sodium and potassium deteriorate the floc structure and physical properties. The Integrated Fixed Activated Sludge (IFAS) process employs fixed film media in the aerobic zone; therefore, both monovalent and divalent cations are involved in the process performances. In this study, the effects of cations indicated as the monovalent to divalent cations (M/D) ratio on the biofilm formation and characteristics, and on the IFAS performances for carbon and ammonium removals were evaluated. The experiments were conducted in three IFAS systems feeding with the same wastewater but different M/D ratios and two carbon and nitrogen loadings. The findings revealed that high monovalent with low divalent cations at the M/D ratios higher than 2.0 produced excessive polysaccharides in EPS resulting in high viscosity of activated sludge flocs causing viscous bulking with high SVI values, decreasing the biofilm formation, and increasing the biofilm sloughing. Increasing of both monovalent and carbon loading increased more polysaccharides in the EPS leading to the failures of IFAS system. Nitrification failed at higher M/D ratios because of less nitrifiers in flocs and biofilm. The M/D ratio less than 2.0 is suggested to minimize the excessive EPS production in the IFAS system, especially at high organic loading.

Excessive extracellular polymeric substances induced by organic shocks accelerate electron transfer of oxygen reducing biocathode

Electrotrophic bacteria on cathodes are promising substitutes to precious metals as oxygen reduction reaction catalysts in bioelectrochemical systems (BESs). Leading the anodic effluent to the biocathode has additional benefits of neutralizing pH and removing residual pollutants. However, the overflow of excessive organic pollutants inhibits the activity of autotrophic biocathodes. Adding glucose as an organic shock, we confirm that the startup time of biocathodes is initially prolonged by 1.2 times with a decrease in current. However, the currents inversely surpass the control in glucose-added BESs when the biofilm is mature, and the maximum current density increase by 5.5 times with a relatively stable performance. This increase is mainly attributed to the production of agglomerates dominated by polysaccharides and proteins as extracellular polymeric substances. These agglomerates wrap additional redox shuttles that accelerated the electron transfer between electrotrophic bacteria and the cathode. This study demonstrates for the first time that organic shocks enhance the electroactivity of autotrophic biocathodes and provides insights into the feedback mechanisms of electrotrophic microbial community to environmental changes.

Influence of ibuprofen and its biotransformation products on different biological sludge systems and ecosystem

Ibuprofen (IBU) is one of the frequently detected non-steroidal anti-inflammatory drugs (NSAIDs) in wastewater treatment plants (WWTPs) and aquatic environment. However, little is known about the effect of IBU and its biotransformation products (TPs) on different biological sludge systems and aquatic environment. The effects and toxicity of IBU and TPs on three biological sludge systems (i.e., activated sludge (AS), sulfate-reducing bacteria (SRB)-enriched sludge and anaerobic methanogenic (AnM) sludge systems) and aquatic environment were comprehensively evaluated through a long-term operation of three bioreactors and a series of batch experiments. Both of the SRB-enriched sludge and AnM sludge systems were not affected under a long-term exposure to IBU, based on removing organic carbon and sulfur and producing methane. This could be attributed to the high tolerance of functional microbes in the SRB-enriched sludge (e.g., genus Desulfobacter) and AnM sludge systems (e.g., genus Candidatus Methanomethylicus) for IBU. In contrast, IBU had some apparently inhibitory effects on the AS system, such as reduced organic removal efficiency and poor sludge settling. The analysis on microbial community revealed that IBU significantly inhibited the genera involved in organic degradation (e.g., genus Candidatus Competibacter) and also stimulated those genera (e.g., genus Brachymonas) to secret excess extracellular polymeric substances (EPS), which thus caused sludge bulking in the AS system. The toxicity of IBU and its TPs in the effluent of the AS system was also investigated with Vibrio fischeri bioluminescence inhibition tests and quantitative structure activity relationship (QSAR) analysis by ecological structure-activity relationship (ECOSAR) program. The results indicated that the AS system could effectively eliminate the acute toxicity of both IBU and TPs, but a potential chronic toxicity of IBU could still existed, which could be more harmful to aquatic organisms than that of its TPs. These findings provide an insight into the toxic effects of IBU and its TPs on biological sludge systems and ecosystem.

Nitrogen removal mechanism of marine anammox bacteria treating nitrogen-laden saline wastewater in response to ultraviolet (UV) irradiation: High UV tolerance and microbial community shift

Salt stress can be naturally overcome by marine anammox bacteria (MAB), while their low growth rate and sensitivity to operational conditions are still challenges for the application of anammox. To enhance the enrichment of MAB and decipher the effects of ultraviolet (UV) irradiation on MAB, UV was introduced in the nitrogen removal of MAB treating nitrogen-laden saline wastewater for the first time. The results indicated that MAB were resistant to a fairly high UV-C dose, 12000 mJ/cm2. Their relative abundance was enhanced by 1.2 folds under 12000 mJ/cm2 UV-C. However, the relative abundance of Actinobacteria, Acidobacteria, Chloroflexi and Marinicella were greatly dropped with enhanced UV-C dose. The tolerance mechanism was diversified, e.g. excessive extracellular polymeric substances, special structure of MAB and interspecific competition/cooperation. Although further study was still needed, the findings shed a light on MAB enrichment and exploited great potentials of MAB in nitrogen-laden saline wastewater treatment.

A comprehensive comparison between non-bulking and bulking aerobic granular sludge in microbial communities

Filamentous sludge bulking poses great threats to operational stability of aerobic granular sludge. Exploration of the microbial community aids knowledge of the causative factors to sludge bulking and guides directions for corresponding actions for prevention and controlling. Detailed changes of bacterial community within the non-bulking and bulking were performed and compared with a non-specific method through 1‰ (v/v) hydrogen peroxide (H2O2) addition. Results revealed that non-bulking/bulking granules maintained effective carbon and nitrogen removal, while bulking completely deteriorated enhanced biological phosphorus removal (EBPR). Excess extracellular polymeric substances (EPS) especially polysaccharide (PS) were directly linked with sludge bulking and abundant PS contributed to subsequent granular re-stability. Filamentous bulking dramatically altered the bacterial populations and 1‰ H2O2 effectively controlled bulking by eliminating causative filaments Singulisphaera and Thiothrix. Together, this study provides new insights into the non-bulking/bulking granules and could direct the prevention and control of filamentous bulking in aerobic granules.

Enhanced aerobic granulation by inoculating dewatered activated sludge under short settling time in a sequencing batch reactor

The effect of dewatered activated sludge on aerobic granulation was investigated in a sequencing batch reactor (SBR) under short settling time. The results showed that dewatered sludge accelerated aerobic granulation and the granulation was completed within 5 days. On day 5, the aerobic granules were regular, compact, fast-settling and high granular strength and possessed excellent removal performance of carbon and nitrogen. The change trend of extracellular polymeric substances (EPS) was basically consistent with granular strength and granulation rate, indicating that EPS in granules played a vital function for granulation. Microbial community succession was investigated by pyrosequencing. In 5 days, microbial diversity was reduced and certain strains were rapidly enriched in the granules to become dominant species, serving on a crucial role in rapid granulation and pollutant removal as they could secrete excess EPS and possess the excellentability removing carbon and nitrogen pollutants.

Enhancement of the removal and settling performance for aerobic granular sludge under hypersaline stress

The aerobic granular sludge (AGS) dominated by halophilic microorganisms, was successfully cultivated in a lab-scale sequencing batch reactor (SBR) under varying salinity levels (from 0% to 6% (w/v)). Removal performance of AGS improved with the increase of salinity and increased up to 42.86 mg g−1 VSS h−1 at 6% salinity. Increased salinity resulted in better settling performance of AGS in terms of the sludge volume index (SVI), which was initially 148.80 mL/g at 0% salinity and gradually decreased to 59.1 mL/g at 6% salinity. The increase of salinity stimulated bacteria to secret excessive extracellular polymeric substances (EPS), with its highest production of 725.5 mg/(g·VSS) at 5% salinity. The total protein (PN) exhibited highly positive correlation with the total EPS (R = 0.951), indicating that selective secretion of some functional PN played a key constituent in resisting the external osmotic pressure and improving sludge performance. Salinicola, accounted for up to 91% relative abundance at 6% salinity, showed the high positive correlation (R = 0.953) with salinity. The enrichment of such halophilic or halotolerant microbial community assured both stable and improved removal performance in the AGS system. The enrichment of salt response pathways and altered metabolic processes for salt-tolerant bacteria indicated that the microbial community formed special metabolic pattern under long-term hypersaline stress to maintain favourable cellular activity and removal performance.

Enhancement of waste activated sludge dewaterability using calcium peroxide pre-oxidation and chemical re-flocculation

The effects of combined calcium peroxide (CaO2) oxidation with chemical re-flocculation on dewatering performance and physicochemical properties of waste activated sludge was investigated in this study. The evolutions of extracellular polymeric substances (EPS) distribution, composition and morphological properties were analyzed to unravel the sludge conditioning mechanism. It was found that sludge filtration performance was enhanced by calcium peroxide oxidation with the optimal dosage of 20 mg/gTSS. However, this enhancement was not observed at lower dosages due to the absence of oxidation and the performance deteriorated at higher dosages because of the release of excess EPS, mainly as protein-like substances. The variation in soluble EPS (SEPS) component can be fitted well with pseudo-zero-order kinetic model under CaO2 treatment. At the same time, extractable EPS content (SEPS and loosely bound EPS (LB-EPS)) were dramatically increased, indicating sludge flocs were effectively broken and their structure became looser after CaO2 addition. The sludge floc structure was reconstructed and sludge dewaterability was significantly enhanced using chemical re-flocculation (polyaluminium chloride (PACl), ferric iron (FeCl3) and polyacrylamide (PAM)). The inorganic coagulants performed better in improving sludge filtration dewatering performance and reducing cake moisture content than organic polymer, since they could act as skeleton builders and decrease the sludge compressibility.

Effect of extracellular polymeric substances isolated from Vibrio natriegens on corrosion of carbon steel in seawater

Extracellular polymeric substances (EPS) are isolated from marine Vibrio natriegens (V. natriegens) by a cation exchange resin method. The influences of EPS concentration and corrosion time on the corrosion of carbon steel in seawater are investigated. The results demonstrate that EPS are capable of forming a protective film on the surface of carbon steel in seawater, which retards the diffusion of dissolved oxygen and inhibits the reduction of oxygen on the cathode. In addition, the efficiency of corrosion inhibition increases with an increase in the amount of added EPS during the initial state (the first 3 days); however, excess EPS (exceeding 300 mg L−1) in seawater significantly accelerates the corrosion of carbon steel over the long term (10 days) due to the binding power of EPS with Fe ions.

Effect of wastewater COD/N ratio on aerobic nitrifying sludge granulation and microbial population shift

AbstractThe effect of COD/N ratio on the granulation process and microbial population succession was investigated. Four identical sequencing batch reactors, R1, R2, R3 and R4, were operated with various initial COD/N ratios ranging from 0/200 to 800/200 (m/m). Ethanol was fed as the source of COD. Aerobic granules were successfully cultivated in R2 and R3, operating with the COD/N ratio of 200/200 and 400/200, respectively. Scanning electron microscope observations indicated that short rod-shaped and spherical bacteria were dominant in R2, while granules produced in R3 were surrounded with a large amount of filamentous bacteria. The average specific nitritation rate in R2 and R3 were 0.019 and 0.008 mg N/(mg MLVSS hr), respectively. Fluorescence in situ hybridization results demonstrated that nitrifying bacteria population was enriched remarkably in R2. It indicated that nitrification ability and nitrifying bacteria population were enriched remarkably at low COD/N ratio. However, no granules were formed in R1 and R4 which might attribute to either limited or excessive extracellular polymeric substances production. This study contributed to a better understanding of the role of COD/N ratio in nitrifying sludge granulation.

ENVIRONMENTAL CONTROL OF STALK LENGTH IN THE BLOOM-FORMING, FRESHWATER BENTHIC DIATOM DIDYMOSPHENIA GEMINATA (BACILLARIOPHYCEAE)(1).

Blooms of the freshwater stalked diatom Didymosphenia geminata (Lyngb.) M. Schmidt in A. Schmidt typically occur in oligotrophic, unshaded streams and rivers. Observations that proliferations comprise primarily stalk material composed of extracellular polymeric substances (EPS) led us to ask whether or not the production of excessive EPS is favored under nutrient-limited, high-light conditions. We conducted experiments in outdoor flumes colonized by D. geminata using water from the oligotrophic, D. geminata-affected Waitaki River, South Island, New Zealand, to determine the relationship between D. geminata stalk length, cell division rates, and light intensity under ambient and nutrient-enriched conditions. Stalk lengths were measured in situ, and cell division rates were estimated as the frequency of dividing cells (FDC). FDC responded positively to increasing light intensity and to nutrient additions (N+P and P). Under ambient conditions, stalk length increased as light level increased except at low ambient light levels and temperature. Nutrient enrichment resulted in decreased stalk length and negative correlations with FDC, with this effect most evident under high light. Our results are consistent with the hypothesis that extensive stalk production in D. geminata occurs when cell division rates are nutrient limited and light levels are high. Thus, photosynthetically driven EPS production in the form of stalks, under nutrient-limited conditions, may explain the development of very high biomass in this species in oligotrophic rivers. The responses of FDC and stalk length under nutrient-replete conditions are also consistent with occurrences of D. geminata as a nondominant component of mixed periphyton communities in high-nutrient streams.