Polyhydroxyalkanoate accumulation in Streptomyces coelicolor affected by SCO7613 gene region.

Polyhydroxyalkanoate (PHA) is stored as an important carbon and energy source in bacterial cells. For biomedical applications, gram-positive bacteria can be better sources of PHAs, since they lack outer membrane lipopolysaccharide. Although gram-positive Streptomyces coelicolor A3(2) has been indicated as a high potential PHA producer, pha C gene that encodes the key enzyme PHA synthase in the metabolic pathway is not determined in its genome. BLAST search results of the GenBank database argued that SCO7613 could specify a putative polyhydroxyalkanoate synthase (PhaC) responsible for PHA biosynthesis. Deduced amino acid sequence of SCO7613 showed the presence of conserved lipase box like sequence, 555GASAG559, in which serine residue was present as the active nucleophile. Present study describes deletion of putative S. coelicolor pha C gene via PCR dependent method. We showed that SCO7613 is not an essential gene in S. coelicolor and its deletion affected PHA accumulation negatively although it is not ceased. Transcomplementation abolished the mutant phenotype, demonstrating that the decrease in PHA resulted from the deletion of SCO7613.

Determination of the potential of pickle wastewater as feedstock for biopolymer production.

Food industry wastewater (FIWW) streams with high organic content are among the most suitable and inexpensive candidates for polyhydroxyalkanoate (PHA) biopolymer production. Due to its high organic acid content, pickle industry wastewater (PIWW), can be considered as one of the prospective alternatives to petroleum-based polymers for PHA production. In this context, this study aimed to investigate the production of PHA with enriched microbial culture using PIWW. Two laboratory scale sequencing batch reactors (SBRs) were operated under aerobic dynamic feeding conditions at a sludge retention time of 8 days, with a total cycle duration of 24 hours. SBRs were fed with peptone mixture and PIWW. In-cycle analysis and batch respirometric tests were performed to evaluate PHA storage together with biodegradation kinetics. In-cycle analysis showed that maximum PHA content was 1,820 mgCOD/L, corresponding to 44% in the biomass (ratio of chemical oxygen demand (COD) to volatile suspended solids) for PIWW. Experimental results were also confirmed with activated sludge model simulations. As for the PHA composition, hydroxybutyrate was the major fraction. Model simulations proposed a unique conversion-degradation-storage pathway for the organic acid mixture. This paper presents a novel insight for better understanding of PHA biopolymer production using high saline FIWW.

Microbial community shifts in the oxic-settling-anoxic process in response to changes to sludge interchange ratio.

This particular study set out to demonstrate alterations on the microbial community of the oxic-settling-anaerobic/anoxic (OSA) process treating real domestic wastewater by changing interchange ratios (IRs). The sludge yield of systems operated at different IRs (1/13, 1/17 and 1/20) to assess sludge reduction was used to analyze microbial community composition variations. The highest IR (1/13) resulted in the highest sludge reduction (52.1%), while the OSA systems with IR of 1/17 and 1/20 reduced sludge production by 37.4% and 35.5%, respectively, in comparison to conventional systems. 16S rRNA gene amplicon sequencing analysis showed that the bacterial communities were composed of similar phylogenetic groups, Proteobacteria, Acidobacteria, and Bacteroidetes being dominant. The relative abundances differed due to the applied IRs. The highest abundance of Actinobacteria was determined at the highest IR (1/13) and increasing of the HRT to 1/20 caused a significant reduction in Actinobacteria species and the lowest abundance (6%) was determined in the OSA systems. The abundant of Thiothrix species that are boosted in the OSA trials may have a vital role in OSA systems, where its abundance was below the detection limits in the seed sludge sample. Therefore, they could be used as bioindicators in the OSA system.

Potential for simultaneous nitrogen removal and sludge reduction of the oxic-settling-anaerobic process operated as a dual fed sequencing batch reactor.

The objective of the study was to explore the impact of different operation strategies on the performance of the Oxic-Settling-Anaerobic (OSA) process. A sequencing batch reactor was selected as the aerobic (oxic) unit of the OSA system and it was operated as a dual mixing phase with step feeding in order to optimize simultaneous nitrogen removal and sludge minimization. For this purpose, the effect of COD/N ratio, filling pattern and the fraction of anaerobic period was investigated on the performance of a laboratory-scale OSA system fed with synthetic wastewater (peptone mixture) approximating the characteristics of sewage. In nine consecutive experimental stages, each sustaining different metabolic processes and biochemical reactions, the sludge reduction potential of the OSA system was not impaired, maintaining an average observed yield value of 0.18 g VSS/g COD, which was approximately half the level observed in the reference reactor. Similarly, the OSA scheme of operation did not affect or alter the fundamental mechanisms of biological nitrogen removal, where system performance could be modified and optimized the same way as in a conventional activated sludge configuration. Furthermore, the OSA system maintained a higher level of biomass concentration compared with the reference reactor. This observation confirmed the previous analyses that the continuous biomass inflow from the side-stream anaerobic bioreactor was the reason to establish the microbial mechanism that resulted in a much lower sludge generation, regardless of the biochemical processes taking place in the reactor.

Evaluation of sludge reduction in an oxic-settling-anoxic system operated with step feeding regime for nutrient removal and fed with real domestic wastewater.

In the present study, the effect of sludge age was evaluated for simultaneous sludge reduction and nitrogen removal in an oxic-settling-anoxic (OSA) system fed with real domestic wastewater. Three laboratory-scale systems utilizing aerobic and anoxic zones and step feeding regime were operated for sludge age of 13, 17 and 20 days in the main reactors. A significant influence of sludge age on the sludge reduction was observed compared to conventional activated sludge systems (CAS). The greater corresponding sludge reduction was achieved as 58% operated at interchange ratio of 7.7% (1/13) in the side-stream reactor, while others revealed 37% and 35%, where interchange ratios were 5.9 (1/17) and 5.0% (1/20), respectively. In both CAS and OSA systems, high removal efficiencies of organic matter and nitrogen were achieved using real domestic wastewater. The results indicate that intermittently aerated OSA systems could accomplish less sludge production and higher nitrogen removal (up to 85%) simultaneously. Thus, it is suggested that interchange ratio of around 8% is more optimized level, which is a little lower than that of proposed and applied in most studies in the literature, which would possibly be more cost-effective.

Determination and evaluation of kinetic parameters of activated sludge biomass from a sludge reduction system treating real sewage by respirometry testing.

Oxic-settling-anoxic (OSA) process is one of the promising variants that produces lower amounts of sludge and has been applied to aerobic and nutrient removal systems. The only consequence on this modification is that supplementary research is advisable to fully understand the mechanism, which eventually leads to the development of a more realistic model. This study evaluated the characteristics of an OSA process as a sludge reduction system by calibration of kinetic coefficients of ASM1 model with some modifications. A series of respirometric tests were designed for the assessment of microbial kinetics and for further clarification of sludge reduction mechanism. The calibration results depicted that the decay rate is the most variable kinetic parameter depending on the system configuration. It was determined that this kinetic coefficient increased significantly after the system was modified into OSA configuration while other model parameters were almost kept constant. This may be referred to a change either in the microbial population or in the metabolism of the community. The active biomass ratio in the CAS reactor was found to be around 75%, while it was almost 2 times lower in the side-stream reactor. All results led to a conclusion that OSA process is encouraging endogenous decay and consequently lowers biomass viability in the reactor and achieves excess sludge reduction in the system. All experimental results confirmed that side-stream reactor accelerates decay rate of the community and causes the introduction of sludge with low viability to the main reactor.

The effect of iron dosing on reducing waste activated sludge in the oxic-settling-anoxic process.

This study evaluates the biological solid reduction in a conventional activated sludge system with an anoxic/anaerobic side stream reactor receiving 1/10 of return sludge mass. Influent iron concentrations and feeding modes were changed to explore the consistency between the influent iron concentration and yield values and to assess the impact of feeding pattern. The results indicated that sludge reduction occurs during alternately exposure of sludge to aerobic and anoxic/anaerobic conditions in a range of 38-87%. The sludge reduction values reached a maximum level with the higher iron concentrations. Thus, it is concluded that this configuration is more applicable for plants receiving high iron concentrations in the wastewaters.

Effect of acetate to biomass ratio on simultaneous polyhydroxybutyrate generation and direct microbial growth in fast growing microbial culture.

The study investigated the effect of variations in the acetate to biomass ratio on substrate storage potential, and the kinetics of substrate utilization. A series of batch experiments were conducted with biomass taken from the fill and draw reactor operated at a sludge age of 2 d. One of the batch reactors duplicated the substrate loading in the main reactor. The others were started with different initial acetate to biomass ratios both in lower and higher ranges. Increasing available acetate did not totally divert excess substrate to storage; the microbial culture adjusted the kinetics of the metabolic reactions to a higher growth rate so that more substrate could be utilized for direct growth at high acetate levels. Conversely, storage rate was increased, utilizing a higher substrate fraction for polyhydroxybutyrate generation when acetate concentration was lowered. The physiological and molecular bases of storage at low substrate levels were discussed.

Characteristics of mixed microbial culture at different sludge ages: effect on variable kinetics for substrate utilization.

The study focused on variable kinetics for substrate utilization, primarily addressing the following issue: Is variable process kinetics observed under different operating conditions and culture history (sludge ages), the result of changes inflicted on the metabolic machinery of the same microbial culture? Or, is this the result of a different microbial population selected under different operating conditions? For this purpose, the study mainly emphasized to assess the microbial population composition sustained at different sludge ages. It explored the relationship between observed process kinetics and microbial population structure using respirometric modeling and high-throughput sequencing of 16S rRNA gene. Experimental results indicated a significant change in the composition of the microbial community fed with the same organic substrate, when the culture history was changed, lower sludge age selecting a different and faster growing microbial community.

COD fractionation and denitrification potential of sonicated waste activated sludge liquids.

This study characterized sonicated waste activated sludge (WAS) liquids as a possible carbon source for nitrogen removal. In this context, the effect of sonication density on chemical oxygen demand (COD) and nitrogen release was determined by particle size distribution (PSD) analysis and anoxic batch experiments. The increase in ultrasonic density from 0.8 W/mL to 1.6 W/mL had a slight impact on the soluble COD/total COD ratio. The high ultrasonic energy input increased the solubilization of nitrogenous organic substances and resulted in a low COD/TKN (total Kjeldahl nitrogen) ratio, which is not appropriate for nutrient removal systems. The change in ultrasonic power had a significant effect on COD fractionation of sonicated WAS liquid. The COD fraction at the size ranges higher than 1600 nm decreased from 44% to 3% as the energy input increased. The increase in specific energy raised the COD fraction, at the size ranges of <2 nm, from 11% to 23%.The PSD-based COD fractionation showed that increasing the sonication density markedly changed the size distribution. The anoxic batch tests indicated that the specific denitrification rate of sonicated WAS liquid was in the range of that reported for the slowly biodegradable fraction of the domestic wastewater and higher than those reported for agro-food wastewater.

Effect of nitrogen limitation on enrichment of activated sludge for PHA production.

Polyhydroxyalkanoates (PHA) are good candidates to plastics because of their material properties similar to conventional plastics and complete biodegradability. The use of activated sludge can be a cheaper alternative to pure cultures for PHA production. In this study, effect of nitrogen limitation during acclimatization period of biomass on production of polyhydroxyalkanoate was investigated. Activated sludge was selected in two sequencing batch reactors operated with and without nitrogen limitation. Batch tests were performed to examine polymer productions of activated sludges acclimatized to different nitrogen regimes. Responses of biomass to different organic loading rates, organic acids, and carbon to nitrogen (C/N) ratios were studied by determining specific polymer storage rate, polymer storage yield, and sludge polymer content of biomasses. Results obtained from batch experiments showed that concentrations of polymer accumulated by two different sludges increased directly with initial substrate concentration. Observed highest polymer yields for the biomasses enriched with and without nitrogen deficiency were 0.69 g COD PHA g(-1) COD S and 0.51 g COD PHA g(-1) COD S, and corresponding polymer contents of biomasses were 43.3% (g COD PHA g(-1) COD X) and 38.3% (g COD PHA g(-1) COD X), respectively. Polymer yields for both biomasses decreased with substrate shift however, biomass enriched with nitrogen deficiency adapted well to acetate-propionate mixture. The results presented in this study showed that polymer storage ability of biomass was improved more under dynamic conditions with nitrogen deficiency when compared to that without nitrogen deficiency. Limiting ammonia availability during batch experiments also caused higher polymer production by suppressing growth, as well as during enrichment of biomass.

The investigation and assessment of characteristics of waste activated sludge after ultrasound pretreatment.

In this study, the characteristics of sonicated waste activated sludge (WAS) originating from a nutrient-removal wastewater treatment plant were investigated and evaluated. Different combinations of power inputs, sonication durations and volumes were used for optimization of the sonication conditions. Ultrasound density levels ranged between 0.32 and 3.2 W/mL. Optimal conditions based on soluble COD concentrations (SCOD) after sonication at different ultrasound density levels and sonication durations (1, 5, 10 and 30 min) were determined. An ultrasonic density of 1.6 W/mL and a sonication time of 30 min were identified as optimal sonication conditions in terms of SCOD concentration. The sludge sonicated under these optimal conditions was further investigated to determine the effect of ultrasound treatment on solubilization of organic matter and volatile solids, particle size and biodegradability. Disruption of the sludge structure was confirmed by the increase in the SCOD by 35.5%, reduction in volatile suspended solids by 26% and reduction in particle size by 85%. The availability of COD fractions of sonicated sludge was also tested with oxygen uptake rate and phosphate release data indicating that using sonicated sludge for both subsequent biological treatment and sludge treatment is promising.

Mechanism and design of intermittent aeration activated sludge process for nitrogen removal.

The paper provided a comprehensive evaluation of the mechanism and design of intermittent aeration activated sludge process for nitrogen removal. Based on the specific character of the process the total cycle time, (T(C)), the aerated fraction, (AF), and the cycle time ratio, (CTR) were defined as major design parameters, aside from the sludge age of the system. Their impact on system performance was evaluated by means of process simulation. A rational design procedure was developed on the basis of basic stochiometry and mass balance related to the oxidation and removal of nitrogen under aerobic and anoxic conditions, which enabled selected of operation parameters of optimum performance. The simulation results indicated that the total nitrogen level could be reduced to a minimum level by appropriate manipulation of the aerated fraction and cycle time ratio. They also showed that the effluent total nitrogen could be lowered to around 4.0 mgN/L by adjusting the dissolved oxygen set-point to 0.5 mg/L, a level which promotes simultaneous nitrification and denitrification.

A new interpretation of ASM2d for modeling of SBR performance for enhanced biological phosphorus removal under different P/HAc ratios.

This study evaluated the prediction capability of Activated Sludge Model No. 2d (ASM2d), for the enhanced biological phosphorus removal (EBPR) performance of a sequencing batch reactor (SBR) receiving variable influent phosphate load. For this purpose, a laboratory-scale SBR was operated with a synthetic feed containing acetate as the sole carbon source. The experiments were conducted in four different Runs to ensure a range of different phosphate/acetate ratios in the influent. Model evaluations were carried out using concentration profiles measured throughout a representative cycle at steady state. An iterative calibration methodology was developed based on sensitivity analysis and applied to four different sets of experimental data on relevant model parameters reflecting SBR performance. ASM2d was able to predict the steady state behavior of the SBR system receiving variable influent phosphate loads only with the recalibrated parameter set. The regular changing pattern of the coefficients could be interpreted with the ability of the SBR system to sustain glycogen accumulating microorganisms, GAOs, which can store substrate under anaerobic conditions without polyphosphate energy, but deriving energy from the degradation of glycogen. Thus they are capable of prevailing at lower P/Ac ratios. The results indicate the need to include glycogen and GAOs as model components for processes involving both phosphate accumulating organisms, (PAOs) and GAOs, in order to obtain a better prediction of X(PHA) and oxygen uptake rate (OUR) profiles in the system.

Design of sequencing batch reactors for biological nitrogen removal from high strength wastewaters.

This paper covers an evaluation of more than twenty full-scale industrial wastewater treatment plants employing sequencing batch reactor (SBR) process mainly for carbon removal and a pilot-scale SBR designed for carbon and nitrogen removal from tannery effluent. The study highlights the major features of the SBR technology and proposes a rational dimensioning approach for carbon and nitrogen removal SBRs treating high strength industrial wastewaters based on scientific information on process stoichiometry and modeling, also emphasizing practical constraints in design and operation.

Metabolic model for acetate uptake by a mixed culture of phosphate- and glycogen-accumulating organisms under anaerobic conditions.

This paper proposes a new metabolic model for acetate uptake by a mixed culture of phosphate- and glycogen-accumulating organisms (PAOs and GAOs) under anaerobic conditions. The model uses variable overall stoichiometry based on the assumption that PAOs may have the ability of using the glyoxylate pathway to produce the required reducing power for polyhydroxyalkonate (PHA) synthesis. The proposed model was tested and verified by experimental results. A sequencing batch reactor system was operated for enhanced biological phosphorus removal (EBPR) with acetate as the sole carbon source at different influent acetate/phosphate ratios. The resulting experimental data supported the validity of the proposed model, indicating the presence of GAOs for all tested HAc/P ratios, especially under P-limiting conditions. Strong agreement is observed between experimental values and model predictions for all model components, namely, PHB production, PHA composition, glycogen utilization, and P release.

The effect of nitrate and different substrates on enhanced biological phosphorus removal in sequencing batch reactors.

Enhanced biological phosphorus removal (EBPR) requires an anaerobic-aerobic sequence and short chain fatty acids, namely acetate. It is also known that the presence of nitrate in the anaerobic phase inhibits EBPR. This study describes a lab-scale experimentation carried out to study the effect of different substrates on EBPR and behaviour of PAOs under anoxic conditions in a sequencing batch reactor operated using synthetic wastewater. Experimental data show that the EBPR performance is significantly affected by glucose rich influent. Low COD/TKN ratios caused lower phosphorus removal performance since nitrate entering the anaerobic zone consumes substrate for denitrification. The results also show that anoxic phosphate uptake took place together with nitrate reduction when there was no external substrate. However, the uptake rate under anoxic conditions was lower than that under aerobic conditions.