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- 2-Fluorophenol degradation by aerobic granules in a sequencing batch reactorPublication . Duque, A. F.; Bessa, V. S.; Carvalho, M. F.; De Kreuk, M. K.; Loosdrecht, M. C. M. van; Castro, P. M. L.
- Variability in the composition of extracellular polymeric substances from a full-scale aerobic granular sludge reactor treating urban wastewaterPublication . Oliveira, Ana S.; Amorim, Catarina L.; Ramos, Miguel A.; Mesquita, Daniela P.; Inocêncio, Paulo; Ferreira, Eugénio C.; Loosdrecht, Mark van; Castro, Paula M. L.Within the framework of the circular economy, there is a need for waste management alternatives that promote the reuse of materials produced in wastewater treatment plants (WWTP). An interesting option is the recovery of extracellular substances from sludge. The variability of characteristics of potential recovered bioproducts has to be assessed in full scale operational settings. In this study, aerobic granular sludge (AGS) from a full-scale WWTP treating urban wastewater was regularly collected for 4 months to assess variability in extracellular polymeric substances (EPS) composition and in granular morphology. Variations in the EPS composition occurred with time. Proteins and humic substances were the main EPS components (329–494 and 259−316 mg/g VSS of AGS, respectively), with polysaccharides and DNA representing minor components. The application of an extra purification step after extraction to obtain a purer EPS led to a decrease in the yield of each EPS component, particularly pronounced for the polysaccharides. The final product had a rather constant composition for the monthly samples. The granules showed morphological stability throughout the sampling period and the yield of EPS was correlated to the size of the granules, higher when there was a higher content of small granules (Deq<150 μm) comparing to intermediate (150 ≤ Deq<1500 μm) or large granules (Deq≥1500 μm). This is the first time that a potential valorization strategy for surplus AGS biomass is studied in a full-scale environment. Knowledge on yield and product homogeneity is important as these features are essential for downstream application of the recovered EPS.
- Intermittent load of 2-fluorophenol in saline wastewater shapes aerobic granular sludge microbiome and reactor performancePublication . Oliveira, Ana S.; Amorim, Catarina L.; Loosdrecht, Mark C. M. van; Castro, PaulaIndustrial effluents often contain organic pollutants and variable salinity levels, making their treatment challenging. Aerobic granular sludge (AGS) is an innovative and compact wastewater treatment technology for the treatment of domestic and industrial water streams. The high content of extracellular polymeric substances (EPS) in AGS composition is thought to protect , to some extent, the microbial communities from stressful conditions in external environment. This work aimed to evaluate the robustness of AGS systems in terms of nutrient removal performance and to unravel the protective role of EPS towards transient feeding with a toxic pollutant (2-fluorophenol - 2-FP) in saline wastewater. Moreover, the taxonomic and functional patterns of the AGS microbiome were characterized and linked with nutrient removal performance and EPS production. In order to mimic transient states of composition typical of industrial effluents, the reactor inlet medium periodically varied regarding to 2-FP presence and salt concentration.
- Removal of fluoxetine and its effects in the performance of an aerobic granular sludge sequential batch reactorPublication . Moreira, Irina S.; Amorim, Catarina L.; Ribeiro, Ana R.; Mesquita, Raquel B. R.; Rangel, António O.S.S.; Loosdrecht, Mark C.M. van; Tiritan, Maria E.; Castro, Paula M. L.Fluoxetine (FLX) is a chiral fluorinated pharmaceutical mainly indicated for treatment of depression and is one of the most distributed drugs. There is a clear evidence of environmental contamination with this drug. Aerobic granular sludge sequencing batch reactors constitute a promising technology for wastewater treatment; however the removal of carbon and nutrients can be affected by micropollutants. In this study, the fate and effect of FLX on reactor performance and on microbial population were investigated. FLX adsorption/desorption to the aerobic granules was observed. FLX shock loads (<= 4 mu M) did not show a significant effect on the COD removal. Ammonium removal efficiency decreased in the beginning of first shock load, but after 20 days, ammonia oxidizing bacteria became adapted. The nitrite concentration in the effluent was practically null indicating that nitrite oxidizing bacteria was not inhibited, whereas, nitrate was accumulated in the effluent, indicating that denitrification was affected. Phosphate removal was affected at the beginning showing a gradual adaptation, and the effluent concentration was <0.04 mM after 70 days. A shift in microbial community occurred probably due to FIX exposure, which induced adaptation/restructuration of the microbial population. This contributed to the robustness of the reactor, which was able to adapt to the FLX load.
- Bioaugmentation of Aerobic Granular Sludge with specialized degrading granules treating 2-fluorophenol wastewaterPublication . Oliveira, Ana S.; Amorim, Catarina L.; Zlopasa, Jure; Lin, Yumei; Loosdrecht, Mark C. M. van; Castro, PaulaThe industry growth has been accompanied by an increase in the amount of industrial chemicals being released into the environment. Indigenous microbial communities in wastewater biotreatment processes are not always effective in removing xenobiotics. This work aimed to evaluate the efficiency of a new bioaugmentation strategy in an aerobic granular sludge sequencing batch reactor (AGS-SBR) system fed with 2-fluorophenol (2-FP). Bioreactor performance in terms of phosphate and ammonium removal, 2-FP degradation and chemical oxygen demand (COD) was evaluated. The new bioaugmentation strategy consisted in producing granules using extracellular polymeric substances (EPS) extracted from AGS as a carrying matrix and a 2-FP degrading strain, Rhodococcus sp. FP1. The produced granules were used for the bioaugmentation of a reactor fed with 2-FP. Shortly after bioaugmentation, the produced granules broke down into smaller fragments inside the bioreactor, but 2-FP degradation occurred. After 8 days of bioaugmentation, 2-FP concentration inside the reactor started to decrease, and stoichiometric fluorine release was observed 35 days later. Phosphate and ammonium removal also improved after bioaugmentation, increasing from 30% to 38% and from 20 to 27%, respectively. Complete ammonium removal was only achieved when 2-FP feeding stopped, and phosphate removal was not recovered during operation time. COD removal also improved after the addition of the produced granules. The persistence of Rhodococcus sp. FP1 in the reactor was followed by qPCR. Rhodococcus sp. FP1 was detected 1 day after in the AGS and up to 3 days after bioaugmentation at the effluent. Nevertheless, the 2-FP degradative ability remained thereafter in the granules. Horizontal gene transfer could have happened from the 2-FP degrading strain to indigenous microbiome as some bacteria isolated from the AGS, 3 months after bioaugmentation, were able to degrade 2-FP. This study presents a promising and feasible bioaugmentation strategy to introduce specialized bacteria into AGS systems treating recalcitrant pollutants in wastewater.
- Biological removal processes in aerobic granular sludge exposed to diclofenacPublication . Bessa, Vânia S.; Moreira, Irina S.; Loosdrecht, Mark C. M. van; Castro, Paula M. L.Diclofenac is a worldwide consumed drug included in the watch list of substances to be monitored according to the European Union Water Framework Directive (Directive 2013/39/EU). Aerobic granular sludge sequencing batch reactors (AGS-SBR) are increasingly used for wastewater treatment but there is scant information on the fate and effect of micropollutants to nutrient removal processes. An AGS-SBR fed with synthetic wastewater containing diclofenac was bioaugmented with a diclofenac degrading bacterial strain and performance and microbial community dynamics was analysed. Chemical oxygen demand, phosphate and ammonia removal were not affected by the micropollutant at 0.03 mM (9.54 mg L-1). The AGS was able to retain the degrading strain, which was detected in the sludge throughout after augmentation. Nevertheless, besides some adsorption to the biomass, diclofenac was not degraded by the augmented sludge given the short operating cycles and even if batch degradation assays confirmed that the bioaugmented AGS was able to biodegrade the compound. The exposure to the pharmaceutical affected the microbial community of the sludge, separating the two first phases of reactor operation (acclimatization and granulation) from subsequent phases. The AGS was able to keep the bioaugmented strain and to maintain the main functions of nutrient removal even through the long exposure to the pharmaceutical, but combined strategies are needed to reduce the spread of micropollutants in the environment.
- 2-Fluorophenol degradation by aerobic granular sludge in a sequencing batch reactorPublication . Duque, Anouk F.; Bessa, Vânia S.; Carvalho, Maria F.; Kreuk, Merle K. de; Loosdrecht, Mark C. M. van; Castro, Paula M. L.Aerobic granular sludge is extremely promising for the treatment of effluents containing toxic compounds, and it can economically compete with conventional activated sludge systems. A laboratory scale granular sequencing batch reactor (SBR) was established and operated during 444 days for the treatment of an aqueous stream containing a toxic compound, 2-fluorophenol (2-FP), in successive phases. Initially during ca. 3 months, the SBR was intermittently fed with 0.22 mM of 2-FP added to an acetate containing medium. No biodegradation of the target compound was observed. Bioaugmentation with a specialized bacterial strain able to degrade 2-FP was subsequently performed. The reactor was thereafter continuously fed with 0.22 and 0.44 mM of 2-FP and with 5.9 mM of acetate (used as co-substrate), for 15 months. Full degradation of the compound was reached with a stoichiometric fluoride release. The 2-FP degrading strain was successfully retained by aerobic granules, as shown through the recovering of the strain from the granular sludge at the end of the experiment. Overall, the granular SBR has shown to be robust, exhibiting a high performance after bioaugmentation with the 2-FP degrading strain. This study corroborates the fact that bioaugmentation is often needed in cases where biodegradation of highly recalcitrant compounds is targeted.
- Simultaneous nitrification and phosphate removal by bioaugmented aerobic granules treating a fluoroorganic compoundPublication . Duque, Anouk F.; Bessa, Vânia S.; Dongen, Udo van; Kreuk, Merle K. de; Mesquita, Raquel B. R.; Rangel, Antonio O. S. S.; Loosdrecht, Mark C. M. van; Castro, Paula M. L.The presence of toxic compounds in wastewater can cause problems for organic matter and nutrient removal. In this study, the long-term effect of a model xenobiotic, 2-fluorophenol (2-FP), on ammonia-oxidizing bacteria (AOB), nitrite oxidizing bacteria (NOB) and phosphate accumulating organisms (PAO) in aerobic granular sludge was investigated. Phosphate (P) and ammonium (N) removal efficiencies were high (>93%) and, after bioaugmentation with 2-FP degrading strain FP1, 2-FP was completely degraded. Neither N nor P removal were affected by 50 mg L−1 of 2-FP in the feed stream. Changes in the aerobic granule bacterial communities were followed. Numerical analysis of the denaturing gradient gel electrophoresis (DGGE) profiles showed low diversity for the ammonia monooxygenase (amoA) gene with an even distribution of species. PAOs, including denitrifying PAO (dPAO), and AOB were present in the 2-FP degrading granules, although dPAO population decreased throughout the 444 days reactor operation. The results demonstrated that the aerobic granules bioaugmented with FP1 strain successfully removed N, P and 2-FP simultaneously.
- Recovered granular sludge extracellular polymeric substances as carrier for bioaugmentation of granular sludge reactorPublication . Oliveira, Ana S.; Amorim, Catarina L.; Zlopasa, Jure; Loosdrecht, Mark van; Castro, Paula M. L.An increasing amount of industrial chemicals are being released into wastewater collection systems and indigenous microbial communities in treatment plants are not always effective for their removal. In this work, extracellular polymeric substances (EPS) recovered from aerobic granular sludge (AGS) were used as a natural carrier to immobilize a specific microbial strain, Rhodococcus sp. FP1, able to degrade 2-fluorophenol (2-FP). The produced EPS granules exhibited a 2-FP degrading ability of 100% in batch assays, retaining their original activity after up to 2-months storage. Furthermore, EPS granules were added to an AGS reactor intermittently fed with saline wastewater containing 2-FP. Degradation of 2-FP and stoichiometric fluorine release occurred 8 and 35 days after bioaugmentation, respectively. Chemical oxygen demand removal was not significantly impaired by 2-FP or salinity loads. Nutrients removal was impaired by 2-FP load, but after bioaugmentation, the phosphate and ammonium removal efficiency improved from 14 to 46% and from 25 to 42%, respectively. After 2-FP feeding ceased, at low/moderate salinity (0.6–6.0 g L−1 NaCl), ammonium removal was completely restored, and phosphate removal efficiency increased. After bioaugmentation, 11 bacteria isolated from AGS were able to degrade 2-FP, indicating that horizontal gene transfer could have occurred in the reactor. The improvement of bioreactor performance after bioaugmentation with EPS immobilized bacteria and the maintenance of cell viability through storage are the main advantages of the use of this natural microbial carrier for bioaugmentation, which can benefit wastewater treatment processes.
- Performance of aerobic granular sludge in a sequencing batch bioreactor exposed to ofloxacin, norfloxacin and ciprofloxacinPublication . Amorim, Catarina L.; Maia, Alexandra S.; Mesquita, Raquel B. R.; Rangel, António O.S.S.; Loosdrecht, Mark C.M. van; Tiritan, Maria Elizabeth; Castro, Paula M.L.A granular sludge sequencing batch reactor (SBR) was operated for 340 days for treating a synthetic wastewater containing fluoroquinolones (FQs), namely ofloxacin, norfloxacin and ciprofloxacin. The SBR was intermittently fed with FQs, at concentrations of 9 and 32 mM. No evidence of FQ biodegradation was observed but the pharmaceutical compounds adsorbed to the aerobic granular sludge, being gradually released into the medium in successive cycles after stopping the FQ feeding. Overall COD removal was not affected during the shock loadings. Activity of ammonia oxidizing bacteria and nitrite oxidizing bacteria did not seemto be inhibited by the presence of FQs (maximum of 0.03 and 0.01 mM for ammonium and nitrite in the effluent, respectively). However, during the FQs feeding, nitrate accumulation up to 1.7 mM was observed at the effluent suggesting that denitrificationwas inhibited. The activity of phosphate accumulating organismswas affected, as indicated by the decrease of P removal capacity during the aerobic phase. Exposure to the FQs also promoted disintegration of the granules leading to an increase of the effluent solid content, nevertheless the solid content at the bioreactor effluent returned to normal levelswithin ca. 1month after removing the FQs in the feed allowing recovery of the bedvolume. Denaturing gradient gel electrophoresis revealed a dynamic bacterial community with gradual changes due to FQs exposure. Bacterial isolates retrieved from the granules predominantly belonged to a- and g-branch of the Proteobacteria phylum.