Bacterial community and filamentous population of industrial wastewater treatment plants in Belgium.

The discharge of industrial water requires the removal of its pollutants, where biological wastewater treatment plants (WWTPs) are the most used systems. Biological WWTPs make use of activated sludge (AS), where bacteria are responsible for the removal of pollutants. However, our knowledge of the microbial communities of industrial plants is limited. Understanding the microbial population is essential to provide solutions to industrial problems such as bulking. The aim of this study was to identify at a high taxonomic resolution the bacterial population of 29 industrial WWTPs using 16S rRNA amplicon sequencing. Our results revealed that the main functional groups were dominated by Thauera and Zoogloea within denitrifiers, Dechloromonas in phosphate-accumulating organisms, and Defluviicoccus in glycogen-accumulating organisms. The activated sludge characterization indicated that 59% of the industrial plants suffered from bulking sludge, with DSVI values of up to 448 mL g-1. From the bulking cases, 72% corresponded to filamentous bulking with Thiothrix as the most abundant filament; meanwhile, the other 28% corresponded to viscous bulking sludge in which Zoogloea was the most abundant genus. Furthermore, the bacterial population did not share a core of taxa across all industrial plants. However, 20 genera were present in at least 50% of the plants comprising the general core, including Thauera, Ca. Competibacter, and several undescribed microorganisms. Moreover, statistical analysis revealed that wastewater salinity strongly affected the microbial richness of the industrial plants. The bacterial population across industrial plants differed considerably from each other, resulting in unique microbial communities that are attributed to the specificity of their wastewaters. KEY POINTS: • The general core taxa of industrial plants were mostly made up of undescribed bacterial genera. • Filamentous bacteria constituted on average 4.1% read abundance of the industrial WWTPs. • Viscous bulking remains a significant type of bulking within industrial WWTPs.

Impact of low and high temperatures on aerobic granular sludge treatment of industrial wastewater.

The goal of this study was to unravel the impact of high and low temperatures (T) on glycogen-accumulating microorganisms (GAOs) which were stimulated in an aerobic granular sludge plant fed with industrial wastewater, which is derived from the cleaning of trucks transporting chocolate and beer. Among GAOs, Candidatus Competibacter (Ca. Competibacter) was the most abundant. The long-term impact on (1) anaerobic dissolved organic carbon (DOC) uptake, (2) sludge morphology, and (3) microbial community composition was investigated. In addition, the short-term impact of T changes on the anaerobic uptake rate was evaluated. High T (above 38 °C) and low T (below 11 °C) had a negative impact on the relative read abundance of Ca. Competibacter and the anaerobic DOC uptake. Nevertheless, the carbon removal efficiency and the settleability of the biomass were not affected. Denitrifiers such as Thauera and Zoogloea were promoted over Ca. Competibacter under high T and low T, respectively, indicating their positive contribution to granulation maintenance.

Full dynamic control of dairy wastewater treatment by aerobic granular sludge using electric conductivity and oxygen uptake rate.

The objective of the current study was to determine the applicability of a sensor-based dynamic control strategy for the treatment of real variable dairy wastewater by aerobic granular sludge (AGS) performing enhanced biological phosphorus removal (EBPR). Two parallel sequencing batch reactors (SBRs) were set up that used only an anaerobic feast/aerobic famine microbial selection strategy to successfully obtain sludge granulation. SBR-STA used a fixed cycle length, while the duration of the reaction steps in SBR-DYN was variable. The control strategy was based solely on (derived) signals from low-cost and common sensors. The profile of the electric conductivity during the anaerobic reaction step was related to the microbial release of phosphate (PO4-P) and the associated uptake of dissolved organic carbon (DOC) by polyphosphate-accumulating organisms (PAOs). Control of the aerobic reaction step was based on the oxygen uptake rate (OUR). This resulted in a dynamic reactor operation with significant efficiency gains, such as 32% shorter cycle times and 42% higher sludge loading rates without impairing the effluent quality. These results extend the existing potential of indirect control strategies to full biological nutrient removal processes, which may be of great assistance to the operators and designers of industrial installations.

Application of partial ozonation on tank truck cleaning concentrate and the influence on biodegradability and ecotoxicity: a pilot-scale study.

This study investigates the pilot-scale ozone treatment of reverse osmosis concentrate (ROC), originating from variable tank truck cleaning wastewater. The influence of ozonation on short- and long-term biodegradation potential was examined through respirometry and Zahn-Wellens, respectively. Ecotoxicity was also examined for several concentrate batches and ozonation steps. Chemical oxidation through ozone had a beneficial effect on chemical oxygen demand removal, with a removal efficiency up to 56%. Formation of short-term biochemical oxygen demand (BODst) was induced for several, but not all batches, showing the potential of subsequent biological treatment of ozonated ROC. An increase in the inherent biodegradability through Zahn-Wellens was observed for all tested samples after ozonation, rising to a maximum of 68% after 3 hours of ozonation, highlighting the importance of sludge adaptation. Ecotoxicity, tested with Artemia franciscana and the saltwater algae P. tricornutum, showed initial decreases in algae inhibition after short ozonation periods. An increase in algae inhibition was, however, seen after prolonged ozonation for all tested ROC samples, pointing to the formation of ecotoxic by-products. Artemia showed no significant toxicity effects. When applying biological treatment through Zahn-Wellens, a decrease in ecotoxicity was observed for several samples, likely through biological oxidation of the produced degradation products.

Implementation of an anaerobic selector step for the densification of activated sludge treating high-salinity petrochemical wastewater.

Sludge bulking is a common challenge in industrial biological wastewater treatment. Leading to difficulties such as bad sludge settling and washout, which is a problem also encountered in the petrochemical industry. Anaerobic feeding strategies can be used to induce the growth of storage-capable organisms, such as glycogen-accumulating organisms (GAO), leading to denser sludge flocs and better settling. In this study, the implementation of an anaerobic feeding strategy was investigated for high-salinity petrochemical wastewater (±35 g salts·L-1), using a sequencing batch reactor. Influent, effluent and sludge characteristics were analyzed throughout the operational period, which can be divided into three stages: I (normal operation), II (increased influent volume) and III (longer anaerobic mixing). Good effluent quality was observed during all stages with effluent chemical oxygen demand (COD) < 100 mgO2·L-1 and removal efficiencies of 95%. After 140 days, the sludge volume index decreased below 100 mL·g-1 reaching the threshold of good settling sludge. Sludge morphology clearly improved, with dense sludge flocs and less filaments being present. A maximum anaerobic dissolved oxygen carbon (DOC) uptake was achieved on day 80 with 74% during stage III. 16S rRNA amplicon sequencing showed the presence of GAOs, with increasing relative read abundance over time from 1 to 3.5%.

Performance of an aerobic granular sludge membrane filtration in a full-scale industrial plant.

This study quantifies the hydraulic performance of a pilot-scale ultrafiltration system integrated into a full-scale industrial aerobic granular sludge (AGS) plant. The treatment plant consisted of parallel AGS reactors, Bio1 and Bio2, with similar initial granular sludge properties. During the 3-month filtration test, a chemical oxygen demand (COD) overloading episode took place, affecting the settling properties, morphology, and microbial community composition in both reactors. The impact on Bio2 was more severe than on Bio1, with higher maximal sludge volume index values, a complete loss of granulation, and the excessive appearance of filamentous bacteria extending from the flocs. The membrane filtration properties of both sludges, with these different sludge qualities, were compared. The permeability in Bio1 varied between 190.8 ± 23.3 and 158.9 ± 19.2 L·m-2·h-1·bar-1, which was 50% higher than in Bio2 (89.9 ± 5.8 L·m-2·h-1·bar-1). A lab-scale filtration experiment using a flux-step protocol showed a lower fouling rate for Bio1 in comparison with Bio2. The membrane resistance due to pore blocking was three times higher in Bio2 than in Bio1. This study shows the positive impact of granular biomass on the long-term membrane filtration properties and stresses the importance of granular sludge stability during reactor operation.

Is building up substrate during anaerobic feeding necessary for granulation?

For a successful granulation process in activated sludge systems, the stimulation of slow growing organisms such as glycogen accumulating microorganisms (GAOs) is a key factor. Here we show that the introduction of an anaerobic feast followed by an aerobic famine phase successfully transforms bulking sludge, caused by the abundance of genus Kouleothrix, to a hybrid floccular-granular sludge. Two sequencing batch reactors (SBRs) were operated for 228 days treating the same industrial wastewater derived from the cleaning of trucks transporting liquid food (the cargo consists of approximately 70% chocolate and 30% beer). By respectively applying a fast and slow feeding in two parallel SBRs, different degrees of substrate build-up were achieved in the two reactors during the feast phase. The F/M ratio over the feeding time was 1.41 ± 0.48 and 0.57 ± 0.16 kg COD·(kg VSS*d)-1 for the fast-fed and the slow-fed SBR respectively. Our results demonstrate that substrate build-up during the anaerobic selection step is not necessary to obtain well-settling granular-like sludge.

Pilot-scale evaluation of ozone as a polishing step for the removal of nonylphenol from tank truck cleaning wastewater.

The presence of nonylphenol (NP) in the wastewater of the tank truck cleaning industry is a major concern because of its endocrine disruptive properties. In this paper, the use of ozone for degrading NP from tank truck cleaning wastewater was investigated by operating a pilot-scale biological wastewater treatment in combination with an ozonation unit. The impact of the added ozonation step on the removal of NP, soluble chemical oxygen demand (sCOD) and total organic carbon (TOC) was monitored over one year. sCOD and TOC removal were not significantly enhanced, but the NP peak concentrations in the effluent were significantly lower than those obtained after biological treatment only: a relatively low NP concentration was observed, even when peak loads were present in the influent of the pilot-scale biological wastewater treatment plant (influentbio). Contrariwise, the effluent of the sole biological treatment follows the peak load trends of the influentbio. During the ozonation period, the average NP concentration in the combined biological-ozone unit was 0.29 µg/L, compared to 1.89 µg/L for the effluent obtained after a sole biological treatment, resulting in an improved average removal efficiency of 32%.

A dynamically controlled anaerobic/aerobic granular sludge reactor efficiently treats brewery/bottling wastewater.

This study investigated the application of a dynamic control strategy in an aerobic granular sludge (AGS) reactor treating real variable brewery/bottling wastewater. For 482 days, the anaerobic and aerobic reaction steps in a lab-scale AGS system were controlled dynamically. A pH-based control was used for the anaerobic step, and an oxygen uptake rate (OUR) based control for the aerobic step. Additionally, the effect of an elongated aerobic step, and the effect of the removal of the suspended solids from the influent, on AGS formation were also investigated. In comparison to a static operation, the dynamic operation resulted in similar reactor performance, related to effluent quality and the anaerobic dissolved organic carbon (DOC) uptake efficiency, while the organic loading rate was significantly higher. The removal of suspended solids from the influent by chemical coagulation with FeCl3 turned hybrid floccular-granular sludge into fully granular sludge. The granulation coincided with a significant increase in the abundance of the glycogen-accumulating Candidatus Competibacter and an increase in the content of gel-forming EPS to respectively around 14% and 30%. In conclusion, this study showed the successful application of a dynamic control strategy based on common and low-cost sensors for AGS treatment of industrial wastewater.

Fast liquid chromatography-tandem mass spectrometry methodology for the analysis of alkylphenols and their ethoxylates in wastewater samples from the tank truck cleaning industry.

A fast methodology to quantify 4-tert-octylphenol (4-t-OP) and 4-nonylphenol (4-NP) and their mono- and di-ethoxylates was developed, validated, and applied to real wastewater samples. Dispersive liquid-liquid microextraction was employed as a sample preparation step, leading to a pre-concentration factor of roughly 30. Analysis was carried out by liquid chromatography-tandem mass spectrometry with electrospray ionisation in multiple reaction monitoring mode. Average recoveries were generally between 80 and 120% for both the alkylphenols and their mono- and di-ethoxylates in influent and effluent wastewater. A minimum of 5 concentration levels per compound, ranging between 1 and 500 ng/mL, were prepared to construct calibration curves making use of isotopically labelled internal standards. The method presented good linearity and repeatability over the whole range of concentrations. Taking into account the concentration factor, and the recovery of the compounds, lower limits of quantification obtained in effluent wastewater were 0.04 ng/mL for 4-t-OP and 0.14 ng/mL for 4-NP, complying with European regulations, and between 0.03 ng/mL and 0.39 ng/mL for the ethoxylates. In influent wastewater, these limits were slightly higher. The total run time of 5 min for the alkylphenols and 8 min for the ethoxylates ensured high throughput. The developed method was applied to determine 4-t-OP and 4-NP and their mono- and di-ethoxylates in wastewater from several tank truck cleaning companies, which was subjected to ozonation and/or biological treatment. It was demonstrated that ozonation was best applied after the biological treatment, since in this case, the biological treatment could degrade most of the biodegradable organic matter, after which ozone could react directly with the recalcitrant organic pollutants. In this case, the concentrations of the target compounds in the wastewater of the investigated company decreased below the legally allowed concentration of the European water legislation.

Influence of mixed feeding rate in a conventional SBR on biological P-removal and granule stability while treating different industrial effluents.

In this study, the influence of the anaerobic mixed feeding rate on granule stability and reactor performance in a conventional sequencing batch reactor (C-SBR) was investigated while treating various industrial wastewaters. A laboratory-scale SBR fed with malting wastewater rich in phosphorus was operated for approximately 250 days, which was divided into two periods: (I) mixed pulse feed and (II) prolonged mixed feed. Initially, no bio-P activity was observed. However, by lowering the feeding rate biological P-removal was rapidly established and no effect on the aerobic granular sludge (AGS) characteristics was observed. Additionally, to investigate the effect of the mixed feeding rate when treating an industrial effluent with low phosphorus content, i.e. brewery wastewater, a laboratory-scale reactor was operated for approximately 400 days applying different mixed feeding rates. Morphological and molecular analysis indicated that a low substrate concentration promoted the enrichment of anaerobic carbon storing filaments when fed with brewery wastewater. Findings suggest that a prolonged mixed feeding regime can be used as a tool to easily establish bio-P removal in a C-SBR system for the treatment of phosphorus-rich wastewaters. It should however be considered that under P-limiting conditions, enrichment of poly-P storing filaments may occur, possibly due to their higher substrate affinity under anaerobic conditions.

Formation of aerobic granular sludge and the influence of the pH on sludge characteristics in a SBR fed with brewery/bottling plant wastewater.

A laboratory-scale sequencing batch reactor (SBR) was operated for 450 days to assess aerobic granule formation when treating brewery/bottling plant wastewater by consistent application of a feast/famine regime. The experiment was divided into three major periods according to the different operational conditions: (I) no pH control and strong fluctuations in organic loading rate (OLR) (1.18 ± 0.25 kgCOD·(m3·day)-1), (II) pH control and aeration control strategy to reduce OLR fluctuations (1.45 ± 0.65 kgCOD·(m3·day)-1) and (III) no pH control and stable OLR (1.42 ± 0.18 kgCOD·(m3·day)-1). Aerobic granule formation was successful after 80 days and maintained during the subsequent 380 days. The aerobic granular sludge was characterized by SVI5 and SVI30 values below 60 mL.g-1 and dominated by granular, dense structures. An oxygen uptake rate based aeration control strategy insured endogenous respiration at the end of the aerobic phase, resulting in stable SBR operation when the influent composition fluctuated. The quantitative polymerase chain reaction results show no significant enrichment of Accumulibacter or Competibacter during the granulation process. The 16S rRNA sequencing results indicate enrichment of other, possibly important species during aerobic granule formation while treating brewery wastewaters.

Aeration control strategies to stimulate simultaneous nitrification-denitrification via nitrite during the formation of aerobic granular sludge.

In this study, a sequencing batch reactor (SBR), treating synthetic wastewater (COD/N = 5), was operated in two stages. During stage I, an aeration control strategy based on oxygen uptake rate (OUR) was applied, to accomplish nitrogen removal via nitrite >80%. In stage II, the development of aerobic granular sludge (AGS) was examined while two aeration control strategies (OUR and pH slope) maintained the nitrite pathway and optimized the simultaneous nitrification-denitrification (SND) performance. Stimulation of slow-growing organisms, (denitrifying) polyphosphate-accumulating organisms (D)PAO and (denitrifying) glycogen-accumulating organisms (D)GAO leads to full granulation (at day 200, SVI10 = 47.0 mL/g and SVI30 = 43.1 mL/g). The average biological nutrient removal efficiencies, for nitrogen and phosphorus, were 94.6 and 83.7%, respectively. Furthermore, the benefits of an increased dissolved oxygen concentration (1.0-2.0 mg O2/L) were shown as biomass concentrations increased with approximately 2 g/L, and specific ammonium removal rate and phosphorus uptake rate increased with 33 and 44%, respectively. It was shown that the combination of both aeration phase-length control strategies provided an innovative method to achieve SND via nitrite in AGS.

The sequencing batch reactor as an excellent configuration to treat wastewater from the petrochemical industry.

In the present study, the influence of a changing feeding pattern from continuous to pulse feeding on the characteristics of activated sludge was investigated with a wastewater from the petrochemical industry from the harbour of Antwerp. Continuous seed sludge, adapted to the industrial wastewater, was used to start up three laboratory-scale sequencing batch reactors. After an adaptation period from the shift to pulse feeding, the effect of an increasing organic loading rate (OLR) and volume exchange ratio (VER) were investigated one after another. Remarkable changes of the specific oxygen uptake rate (sOUR), microscopic structure, sludge volume index (SVI), SVI30/SVI5 ratio, and settling rate were observed during adaptation. sOUR increased two to five times and treatment time decreased 43.9% in 15 days. Stabilization of the SVI occurred after a period of 20 days and improved significantly from 300 mL·g-1 to 80 mL·g-1. Triplication of the OLR and VER had no negative influence on sludge settling and effluent quality. Adaptation time of the microorganisms to a new feeding pattern, OLR and VER was relatively short and sludge characteristics related to aerobic granular sludge were obtained. This study indicates significant potential of the batch activated sludge system for the treatment of this industrial petrochemical wastewater.

Enhanced treatment of secondary municipal wastewater effluent: comparing (biological) filtration and ozonation in view of micropollutant removal, unselective effluent toxicity, and the potential for real-time control.

Ozonation and three (biological) filtration techniques (trickling filtration (TF), slow sand filtration (SSF) and biological activated carbon (BAC) filtration) have been evaluated in different combinations as tertiary treatment for municipal wastewater effluent. The removal of 18 multi-class pharmaceuticals, as model trace organic contaminants (TrOCs), has been studied. (Biological) activated carbon filtration could reduce the amount of TrOCs significantly (>99%) but is cost-intensive for full-scale applications. Filtration techniques mainly depending on biodegradation mechanisms (TF and SSF) are found to be inefficient for TrOCs removal as a stand alone technique. Ozonation resulted in 90% removal of the total amount of quantified TrOCs, but a post-ozonation step is needed to cope with an increased unselective toxicity. SSF following ozonation showed to be the only technique able to reduce the unselective toxicity to the same level as before ozonation. In view of process control, innovative correlation models developed for the monitoring and control of TrOC removal during ozonation, are verified for their applicability during ozonation in combination with TF, SSF or BAC. Particularly for the poorly ozone reactive TrOCs, statistically significant models were obtained that correlate TrOC removal and reduction in UVA254 as an online measured surrogate parameter.

Dynamic control of nutrient-removal from industrial wastewater in a sequencing batch reactor, using common and low-cost online sensors.

On-line control of the biological treatment process is an innovative tool to cope with variable concentrations of chemical oxygen demand and nutrients in industrial wastewater. In the present study we implemented a simple dynamic control strategy for nutrient-removal in a sequencing batch reactor (SBR) treating variable tank truck cleaning wastewater. The control system was based on derived signals from two low-cost and robust sensors that are very common in activated sludge plants, i.e. oxidation reduction potential (ORP) and dissolved oxygen. The amount of wastewater fed during anoxic filling phases, and the number of filling phases in the SBR cycle, were determined by the appearance of the 'nitrate knee' in the profile of the ORP. The phase length of the subsequent aerobic phases was controlled by the oxygen uptake rate measured online in the reactor. As a result, the sludge loading rate (F/M ratio), the volume exchange rate and the SBR cycle length adapted dynamically to the activity of the activated sludge and the actual characteristics of the wastewater, without affecting the final effluent quality.

Evaluation of acute ecotoxicity removal from industrial wastewater using a battery of rapid bioassays.

The present study compares conventional wastewater treatment technologies (coagulation-flocculation and activated sludge) and powdered activated carbon (PAC) treatment for the removal of acute ecotoxicity from wastewater generated by tank truck cleaning (TTC) processes. Ecotoxicity was assessed with a battery of four commercially available rapid biological toxicity testing systems, verified by the US Environmental Protection Agency. Chemical coagulation-flocculation of raw TTC wastewater had no impact on the inhibition of the bioluminescence by Vibrio fischeri (BioTox assay). Subsequent biological treatment with activated sludge without PAC resulted in BioTox inhibition-free effluent (<10% inhibition). In contrast, activated sludge treatment without PAC produced an effluent that significantly inhibited (>50%) (i) the bioluminescence by Photobacterium leiognathi (ToxScreen³ test kit), (ii) the photosynthesis by the green algae Chlorella vulgaris (LuminoTox SAPS test kit), and (iii) the particle ingestion by the crustacean Thamnocephalus platyurus (Rapidtoxkit test kit). The lowest inhibition was measured after activated sludge treatment with the highest PAC dose (400 mg/L), demonstrating the effectiveness of PAC treatment for ecotoxicity removal from TTC wastewater. In conclusion, the combination of bioassays applied in the present study represents a promising test battery for rapid ecotoxicty assessment in wastewater treatment.

Removal of ecotoxicity and COD from tank truck cleaning wastewater.

Tank truck cleaning (TTC) activities generate highly complex wastewater. In a previous study, we found that a significant ecotoxic effect was still present in biologically treated TTC wastewater. The aim of the present study was therefore to investigate the removal of acute toxicity from TTC wastewater by a sequence of technologies routinely applied for industrial wastewater. Acute toxicity was assayed with the widely applied and standardized Vibrio fischeri bioluminescence inhibition test. During a 5-month period, raw wastewater was grab-sampled from a full-scale TTC company and treated by the different unit operations on a laboratory scale. Chemical pretreatment of the wastewater by coagulation with FeCl3 removed approx. 38% of the influent chemical oxygen demand (COD) and reduced the bioluminescence inhibition by 8%. Biological treatment with activated sludge subsequently removed another 77% of the remaining COD. This treatment step also reduced the bioluminescence inhibition but the removal efficiency varied strongly from 5 to 92% for the different samples. Powdered activated carbon almost completely removed the remaining COD and inhibition in all samples. The results suggest that conventional technologies did not suffice for complete removal of toxicity from TTC wastewater, and that advanced wastewater treatment technologies such as activated carbon are required for a satisfactory detoxification.

Nitrous oxide formation during simultaneous phosphorus and nitrogen removal in aerobic granular sludge treating different carbon substrates.

The impact of different substrates on N2O dynamics and gene expression of marker enzymes (nirS, nirK and nosZ) involved in denitrifying enhanced biological phosphorus removal (d-EBPR) was investigated. Aerobic granular sludge fed with VFAs led to an anoxic P-uptake (27.7 ± 1.2 mg PO43--P.gVSS-1) and N2O emissions up to 80.7 ± 3.4% N2O-N. A decisive role of Accumulibacter in N2O formation was observed. Dosage of amino acids (12.0 ± 1.2 mg PO43--P.gVSS-1) and glucose (1.5 ± 0.9 mg PO43--P.gVSS-1) as sole substrate did not support d-EBPR activity. Presence of NO2- resulted in higher N2O formation in comparison to nitrate and a nosZ/(nirS + nirK) ratio lower than 0.3. A linear correlation (R2 > 0.95) between the nosZ/(nirS + nirK) ratio and the N2O reductase rate was found only when dosing the same type of substrate. This suggests an interplay between the microbial community composition and different polyhydroxyalkanoates derivatives, when dosing different substrates.

Impact of the substrate composition on enhanced biological phosphorus removal during formation of aerobic granular sludge.

Performance of enhanced biological phosphorus removal (EBPR) is often investigated with simple synthetic wastewater containing volatile fatty acids (VFAs). In this study, various (fermentable) substrates, individually and in mixtures, were examined during the application of a granulation strategy. In addition, the microbial community and N2O formation were monitored. Sludge densification was observed in all systems. Stable EBPR, associated with the presence of Accumulibacter and an anaerobic P-release up to 21.9 mgPO43--P.gVSS-1, was only obtained when VFAs were present as sole substrate or in mixture. Systems fed with VFAs were strongly related to the formation of N2O (maximum of 6.25% relative to the total available nitrogen). A moderate anaerobic dissolved organic carbon (DOC) uptake was observed when amino acids (64.27 ± 3.08%) and glucose (75.39 ± 5.79%) as sole carbon source were applied. The substrate/species-specific enrichment of Burkholderiaceae and Saccharimonadaceae respectively, resulted in unstable EBPR in those systems.