Organic media-based two-stage traditional and electrode-integrated tidal flow wetlands to treat landfill leachate: Influence of aeration strategy and plants.

Landfill leachate treatment employing normal and electrode-integrated constructed wetlands is difficult due to the presence of significant amounts of organic compounds, which frequently impede the progression of microbial-based aerobic pollutant removal pathways. As a result, this study examines the effect of supplementary air availability via intermittent and continuous aeration strategies in improving organic, nutrient, and coliform removals of the unplanted, planted (normal and electrode-integrated) two-stage tidal flow constructed wetlands designed to treat landfill leachate. The constructed wetlands were filled with coal and biochar media and planted with Canna indica. Mean chemical oxygen demand (COD), total nitrogen (TN), total phosphorus (TP), and coliform removal percentages of the externally aerated two-stage unplanted, only planted, planted-microbial fuel cell integrated constructed wetland systems ranged between 96 and 99%, 82 and 93%, 91 and 98%, 86 and 96%, respectively, throughout the experimental campaign. External aeration inhibited the development of a dominant anaerobic environment within the media of the wetland systems and improved overall pollutant removal. The electrode-integrated planted tidal flow wetlands produced better effluent quality than the unplanted or only planted tidal flow systems without electrode assistance. The first stages of the three wetland systems achieved an additional 5-7% COD, 7-12% TN, and 15-22% coliform removal during the continuous aeration period compared to the corresponding performance of the intermittent aeration phase. The pollutant removal performance of the second-stage wetlands decreased during the continuous aeration phase. The media composition supported electrochemically active and inactive microbial-based pollutant removal routes and the chemical adsorption of pollutants. Nitrogen and phosphorus accumulation percentage in plant tissues was low, i.e., 0.4-2.2% and 0.04-0.8%, respectively. During the continuous aeration period, the electrode-integrated tidal flow constructed wetlands achieved higher power density production, i.e., between 859 and 1432 mW (mW)/meter3(m3). This study demonstrates that external aeration might improve pollutant removal performance of the normal, electrodes integrated tidal flow-based constructed wetlands when employed for high organic-strength wastewater treatment such as landfill leachate.

Contaminant Discharge From Outfalls and Subsequent Aquatic Ecological Risks in the River Systems in Dhaka City: Extent of Waste Load Contribution in Pollution.

Dhaka, the capital city, which is the nerve center of Bangladesh, is crisscrossed by six different rivers. A network of peripheral rivers connects the city and functions as a natural drainage system for a massive amount of wastewater and sewage by the increased number of inhabitants impacting the overall environmental soundness and human health. This study intended to identify and characterize the outfalls along the peripheral rivers of Dhaka city with the assessment of different pollution indices such as comprehensive pollution index (CPI), organic pollution index (OPI), and ecological risk indices (ERI). The study evaluated the status of the pollution in the aquatic system in terms of ambient water quality parameters along the peripheral rivers due to discharge from outfalls with a particular focus on waste load contribution. Among the identified outfalls, the majority are industrial discharge (60%), and some are originated from municipal (30%), or domestic sewers (10%). Water quality parameters such as suspended solids (SS), 5-day biochemical oxygen demand (BOD5), and Ammoniacal Nitrogen (NH3-N) for most of the peripheral rivers deviated by as much as 40-50% from industrial discharge standards by the environment conservation rules, Bangladesh, 1997. Based on the CPI, the rivers Buriganga, Dhaleshwari, and Turag could be termed as severely polluted (CPI > 2.0), while the OPI indicated heavy organic pollutant (OPI > 4) contamination in the Dhaleshwari and Buriganga rivers. The associated pollution indices demonstrate a trend for each subsequent peripheral river with significant pollution toward the downstream areas. The demonstrated waste loading map from the outfalls identified sources of significant environmental contaminants in different rivers leading to subsequent ecological risks. The study outcomes emphasize the necessity of systematic investigation and monitoring while controlling the point and non-point urban pollution sources discharging into the peripheral rivers of Dhaka city.

A comparative landfill leachate treatment performance in normal and electrodes integrated hybrid constructed wetlands under unstable pollutant loadings.

This study provides a comparative pollutant removal performance assessment between organic or construction materials-based four hybrid wetland systems that received landfill leachate. The hybrid systems included vertical flow (VF) followed by horizontal flow (HF)-based unplanted and planted systems, and planted electrodes incorporated microbial fuel cell (MFC) integrated hybrid wetlands systems. All the systems were run in free-draining mode. Overall mean chemical oxygen demand (COD), total nitrogen (TN), and total phosphorus (TP) removal percentage of the hybrid systems ranged between 81 and 99%, 82 and 96%, 74 and 99%, respectively, under unstable input pollutant loading conditions. Additionally, up to 27% organic and up to 14% nitrogen removal improvement was observed in electrodes integrated free-draining VF wetlands. Free-draining and additional oxygen availability from atmospheric diffusion, rootzone improved the removal performance of MFC-based VF wetlands. Input load increment decreased organic, nutrient removals in second stage HF units due to saturated media. The chemical composition of the employed media supported biotic, abiotic organic, nutrient removal pathways. Nutrient accumulation percentage in plants tissue was very low, i.e., ≤3%. Bioenergy production across the MFC-based VF-HF wetlands decreased with input pollutant load increment. The single anode electrode-based VF wetland achieved maximum power density production, i.e., 294 mW/m2.. The electrodes integrated hybrid systems achieved comparatively stable removal performance despite input pollutant/hydraulic load variation.

Phenotypic dynamics in polyphosphate and glycogen accumulating organisms in response to varying influent C/P ratios in EBPR systems.

This study employed molecular tools and single cell Raman micro-spectroscopy techniques to reveal the single cell- and population-level phenotypic dynamics and changes in functionally relevant organisms, namely polyphosphate accumulating organisms (PAOs) and glycogen accumulating organisms (GAOs), in response to influent loading readily biodegradable carbon to phosphorus ratio (C/P) changes in enhanced biological phosphorus removal (EBPR) systems. The results, for the first time, provided direct and cellular evidence confirming the adaptive anaerobic metabolic pathway shifts in PAOs in response to influent loading variations. Increase in influent readily biodegradable carbon to phosphorus (C/P) ratio from 20 to 50 led to nearly 50% decline in polyphosphate content and drastic rise of intracellular polyßhydroxybutyrate (PHB) to polyphosphate (polyP) ratio by nearly 6 times in PAOs, indicating corresponding diminishing reliance on polyP hydrolysis for energy as P becomes limiting. Influent carbon availability surge also impacted the intracellular carbon polymers in GAOs, with significant increase in the mean PHB content level but no observed changes in the intracellular glycogen level. Furthermore, the Raman-based quantification of differentiated intracellular polymer content associated with PAOs and GAOs, revealed new insights into the quantitative shift in intracellular carbon storage distribution between the two populations and their variations between the two carbon polymers (PHB, Glycogen). In summary, this investigation revealed high-resolution cellular level information regarding the metabolic flexibility in PAOs, phenotypic stoichiometry changes and carbon flux and distribution among PAOs and GAOs, in response to influent loading conditions. The new information will contribute to improvement in mechanistic EBPR modeling and design.

Advances in single cell Raman spectroscopy technologies for biological and environmental applications.

The increasing sophistication of single cell Raman spectroscopy (SCRS) via its integrations with other advanced analytical techniques and modern data analytics, enable unprecedented exploration of complex biological and environmental samples with significantly improved specificity, sensitivity, and resolution. Because of the merits of being high-resolution, label-free, non-invasive, molecular-specific, culture-independent, and suitable for in situ, in vitro or in vivo analysis, the SCRS-derived techniques offer abilities superior to conventional bulk measurements for environmental and biological studies. Here, we provide a comprehensive and critical review of the most recent advances in the development and application of SCRS-enabled technologies, with focus on those biomolecular and cellular high-resolution applications in environmental and biological fields. The basic principles, unique advantages, and suitable applications, as well as recognized limitations for each technology are recapitulated. The remaining challenges, research needs and future outlook are discussed. We predict that SCRS-enabled technologies are earning its place as a routine and powerful tool in many and rapidly expanding applications across disciplines.

Pollutant removal from landfill leachate employing two-stage constructed wetland mesocosms: co-treatment with municipal sewage.

Constructed wetlands are low-cost, natural technologies that are often employed for the treatment of different types of wastewater. In this study, landfill leachate and municipal wastewater were co-treated by the three parallel two-stage Phragmites- or Vetiver-based constructed wetland mesocosms. Two-stage wetland mesocosms included vertical flow (VF) units as the first stage, followed by horizontal flow (HF)/surface flow (SF)/floating treatment (FT) units. VF and HF wetland mesocosms were filled with gravel, steel slag, concrete block, and intermittent carbon-saturated ceramic filters as substrates. Mean input nitrogen, organics, and phosphorus load across first stages were 75 g N/m2 day, 283 g COD/m2 day, 88 g BOD/m2 day, and 10 g P/m2 day, respectively. N and P accumulation rate was not substantial (< 10%) with respect to total removal in most wetland mesocosms. Gravel-based VF wetland mesocosm achieved better NH4-N and BOD removal (55-59%) during landfill leachate treatment phase, when compared with co-treatment periods (12-52%). Slag-concrete- and ceramic filter-based VF wetland mesocosms maintained stable NH4-N and BOD removals; the former wetland mesocosm was the most efficient VF unit (than other two wetland mesocosms) due to media characteristics. Media-based adsorption accelerated P removal (93%) in slag-concrete-based VF wetland mesocosm. Carbon scarcity limited denitrification in all VF wetland mesocosms; removal of TN was < 32%. Second stage wetland mesocosms achieved higher nitrogen (85-92%), organics (66-90%), and phosphorus (97-100%) removals regardless of operational variations; low input load, long retention time, media, and rhizosphere enhanced removal performances, particularly in HF and FT wetland mesocosms. In general, this study demonstrates potential application of two-stage wetland mesocosms for landfill leachate treatment or co-treatment with municipal sewage.

Impact of solid residence time (SRT) on functionally relevant microbial populations and performance in full-scale enhanced biological phosphorus removal (EBPR) systems.

Investigations of the impact of solid residence time (SRT) on microbial ecology and performance of enhanced biological phosphorus removal (EBPR) process in full-scale systems have been scarce due to the challenges in isolating and examining the SRT from other complex plant-specific factors. This study performed a comprehensive evaluation of the influence of SRT on polyphosphate-accumulating organisms (PAOs) and glycogen-accumulating organisms (GAOs) dynamics and on P removal performance at Clark County Water Reclamation District Facility in Las Vegas, USA. Five parallel treatment trains with separated clarifiers were operated with five different SRTs ranging from 6 to 40 days. Microbial community analysis using multiple molecular and Raman techniques suggested that the relative abundances and diversity of PAOs and GAOs in EBPR systems are highly affected by the SRT. The resultant EBPR system stability and performance can be potentially controlled and optimized by manipulating the system SRT, and shorter SRT (<10 days) seems to be preferred. PRACTITIONER POINTS: Phosphorus removal performance and kinetics are highly affected by the operational solid residence time (SRT), with lower and more stable effluent P level achieved at SRT < 10 days. Excessive long SRTs above that needed for nitrification may harm EBPR performance; additionally, excessive long SRT may favor GAOs to dominate over PAOs and thus further reducing efficient use of rbCOD for EBPR. Microbial population abundance and diversity, especially those functionally relevant PAOs and GAOs, can impact the P removal performances, and they are highly dependent on the operational solid residence time. EBPR performance can be potentially controlled and optimized by manipulating the system SRT, and shorter SRT (≤10 days) seems to be preferred at the influent rbCOD/P ratio and environmental conditions as in the plant studied.

Two-stage constructed wetland systems for polluted surface water treatment.

Two pilot scale wetland systems were studied for the removal of organics, nitrogen, phosphorus and coliform from polluted surface water. Each system consisted of two units: a vertical flow (VF) wetland packed with construction materials gravel, brick or organic sugarcane bagasse, followed by a surface flow (SF) or floating treatment (FT) wetland. All wetland units were planted with Phragmites. The wetland systems were operated under constant and shock hydraulic load (HL) periods. Input COD, N, P loadings ranged between 61 and 2181, 7-1040, 2-194 g/m2d, respectively across first stages of each system. Mean removal percentages ranged between 39 and 97, 11-83, 20-100% and 4-85, 16-86, 1.4-100% across first and second stage wetlands, respectively. Mass balance analyses revealed ≤7% N and ≤14% P accumulation in plants; as such, microbial and adsorption kinetics controlled removal dynamics. Nitrification was the limiting nitrogen removal factor in first stage wetlands; organic carbon was supplied by the employed media. Aerobic organics removal and nitrification were diminished during initial stage of shock load periods. In contrast, second stage SF and FT wetlands showed stable removal performances under similar conditions. Resuspension of settled particles decreased removal performance in second stage wetlands, as shock periods progressed toward final stage. Coliform mortality was increased in second stage wetlands. Physico-chemical properties of brick materials in construction material based VF wetland and hanging root volume inside the water column of FT wetland supplemented removal performance. In general, this study provides evidence on potential application of constructed wetlands for polluted surface water treatment.

Consumption of heavy metal contaminated foods and associated risks in Bangladesh.

This study investigated the magnitude of heavy metal contamination and determined the carcinogenic as well as non-carcinogenic risks associated with selected food consumption in Bangladesh. Commonly consumed varieties of rice, vegetables, and fish samples were analyzed to measure the concentrations of heavy metals such as cadmium, chromium, lead, arsenic, manganese, nickel, and zinc. These staple food items showed the greatest probabilities of heavy metal contamination in different phases of their production and marketing. Wide variations of metal concentrations were observed. Specifically, estimated daily intakes of arsenic and cadmium exceeded allowable daily intakes in all three food items. Toxicity scores of the metals were evaluated, and a comprehensive risk assessment was conducted to quantify the risks associated with the daily food consumption. Except for cadmium and lead in vegetables, all the contaminants present in each food item posed significant levels of carcinogenic risks up to 2.99 × 10-3 compared to the EPA recommended carcinogenic risk level of 1.0 × 10-6. Cadmium and arsenic intake due to rice consumption also posed unsafe levels of non-carcinogenic risks of 4.587 and 6.648, respectively, compared to the EPA recommended non-carcinogenic risk level of 1.0. Finally, a revised set of permissible limits was proposed for the heavy metals detected in the food items. Those permissible limits would ensure the risks associated with food consumption below the allowable carcinogenic and non-carcinogenic risk levels. Thus, this comprehensive approach would provide guidelines to formulate adequate control measures and regulatory limits of toxic metals in foods produced and marketed in Bangladesh.

Advances in techniques for phosphorus analysis in biological sources.

In general, conventional P analysis methods suffer from not only the fastidious extraction and pre-treatment procedures required but also the generally low specificity and poor resolution regarding the P composition and its temporal and spatial dynamics. More powerful yet feasible P analysis tools are in demand to help elucidating the biochemistry nature, roles and dynamics of various phosphorus-containing molecules in vitro and in vivo. Recent advances in analytical chemistry, especially in molecular and atomic spectrometry such as NMR, Raman and X-ray techniques, have enabled unique capability of P analysis relevant to submicron scale biochemical processes in individual cell and in natural samples without introducing too complex and invasive pretreatment steps. Great potential still remains to be explored in wider and more combined and integrated requests of these techniques to allow for new possibilities and more powerful P analysis in biological systems. This review provides a comprehensive summary of the available methods and recent developments in analytical techniques and their applications for characterization and quantification of various forms of phosphorus, particularly polyphosphate, in different biological sources.

Identification of functionally relevant populations in enhanced biological phosphorus removal processes based on intracellular polymers profiles and insights into the metabolic diversity and heterogeneity.

This study proposed and demonstrated the application of a new Raman microscopy-based method for metabolic state-based identification and quantification of functionally relevant populations, namely polyphosphate accumulating organisms (PAOs) and glycogen accumulating organisms (GAOs), in enhanced biological phosphorus removal (EBPR) system via simultaneous detection of multiple intracellular polymers including polyphosphate (polyP), glycogen, and polyhydroxybutyrate (PHB). The unique Raman spectrum of different combinations of intracellular polymers within a cell at a given stage of the EBPR cycle allowed for its identification as PAO, GAO, or neither. The abundance of total PAOs and GAOs determined by Raman method were consistent with those obtained with polyP staining and fluorescence in situ hybridization (FISH). Different combinations and quantities of intracellular polymer inclusions observed in single cells revealed the distribution of different sub-PAOs groups among the total PAO populations, which exhibit phenotypic and metabolic heterogeneity and diversity. These results also provided evidence for the hypothesis that different PAOs may employ different extents of combination of glycolysis and TCA cycle pathways for anaerobic reducing power and energy generation and it is possible that some PAOs may rely on TCA cycle solely without glycolysis. Sum of cellular level quantification of the internal polymers associated with different population groups showed differentiated and distributed trends of glycogen and PHB level between PAOs and GAOs, which could not be elucidated before with conventional bulk measurements of EBPR mixed cultures.

Heterogeneity of intracellular polymer storage states in enhanced biological phosphorus removal (EBPR)--observation and modeling.

A number of agent-based models (ABMs) for biological wastewater treatment processes have been developed, but their skill in predicting heterogeneity of intracellular storage states has not been tested against observations due to the lack of analytical methods for measuring single-cell intracellular properties. Further, several mechanisms can produce and maintain heterogeneity (e.g., different histories, uneven division) and their relative importance has not been explored. This article presents an ABM for the enhanced biological phosphorus removal (EBPR) treatment process that resolves heterogeneity in three intracellular polymer storage compounds (i.e., polyphosphate, polyhydroxybutyrate, and glycogen) in three functional microbial populations (i.e., polyphosphate-accumulating, glycogen-accumulating, and ordinary heterotrophic organisms). Model predicted distributions were compared to those based on single-cell estimates obtained using a Raman microscopy method for a laboratory-scale sequencing batch reactor (SBR) system. The model can reproduce many features of the observed heterogeneity. Two methods for introducing heterogeneity were evaluated. First, biological variability in individual cell behavior was simulated by randomizing model parameters (e.g., maximum acetate uptake rate) at division. This method produced the best fit to the data. An optimization algorithm was used to determine the best variability (i.e., coefficient of variance) for each parameter, which suggests large variability in acetate uptake. Second, biological variability in individual cell states was simulated by randomizing state variables (e.g., internal nutrient) at division, which was not able to maintain heterogeneity because the memory in the internal states is too short. These results demonstrate the ability of ABM to predict heterogeneity and provide insights into the factors that contribute to it. Comparison of the ABM with an equivalent population-level model illustrates the effect of accounting for the heterogeneity in models.

Process optimization by decoupled control of key microbial populations: distribution of activity and abundance of polyphosphate-accumulating organisms and nitrifying populations in a full-scale IFAS-EBPR plant.

This study investigated the abundance and distribution of key functional microbial populations and their activities in a full-scale integrated fixed film activated sludge-enhanced biological phosphorus removal (IFAS-EBPR) process. Polyphosphate accumulating organisms (PAOs) including Accumulibacter and EBPR activities were predominately associated with the mixed liquor (>90%) whereas nitrifying populations and nitrification activity resided mostly (>70%) on the carrier media. Ammonia oxidizer bacteria (AOB) were members of the Nitrosomonas europaea/eutropha/halophila and the Nitrosomonas oligotropha lineages, while nitrite oxidizer bacteria (NOB) belonged to the Nitrospira genus. Addition of the carrier media in the hybrid activated sludge system increased the nitrification capacity and stability; this effect was much greater in the first IFAS stage than in the second one where the residual ammonia concentration becomes limiting. Our results show that IFAS-EBPR systems enable decoupling of solid residence time (SRT) control for nitrifiers and PAOs that require or prefer conflicting SRT values (e.g. >15 days required for nitrifiers and <5 days preferred for PAOs). Allowing the slow-growing nitrifiers to attach to the carrier media and the faster-growing phosphorus (P)-removing organisms (and other heterotrophs, e.g. denitrifiers) to be in the suspended mixed liquor (ML), the EBPR-IFAS system facilitates separate SRT controls and overall optimization for both N and P removal processes.

Application of Raman microscopy for simultaneous and quantitative evaluation of multiple intracellular polymers dynamics functionally relevant to enhanced biological phosphorus removal processes.

Polyphosphate (poly-P), polyhydroxyalkanoates (PHAs), and glycogen are the key functionally relevant intracellular polymers involved in the enhanced biological phosphorus removal (EBPR) process. Further understanding of the mechanisms of EBPR has been hampered by the lack of cellular level quantification tools to accurately measure the dynamics of these polymers during the EBPR process. In this study, we developed a novel Raman microscopy method for simultaneous identification and quantification of poly-P, PHB, and glycogen abundance in each individual cell and their distribution among the populations in EBPR. Validation of the method was demonstrated via a batch phosphorus uptake and release test, in which the total intracellular polymers abundance determined via Raman approach correlated well with those measured via conventional bulk chemical analysis (correlation coefficient r = 0.8 for poly-P, r = 0.94 for PHB, and r = 0.7 for glycogen). Raman results, for the first time, clearly showed the distributions of microbial cells containing different abundance levels of the three intracellular polymers under the same environmental conditions (at a given time point), indicating population heterogeneity exists. The results revealed the intracellular distribution and dynamics of the functionally relevant polymers in different metabolic stages of the EBPR process and elucidated the association of cellular metabolic state with the fate of these polymers during various substrates availability conditions.

Evaluation of intracellular polyphosphate dynamics in enhanced biological phosphorus removal process using Raman microscopy.

A Raman microscopy method was developed and successfully applied to evaluate the dynamics of intracellular polyphosphate in polyphosphate-accumulating organisms (PAOs) in enhanced biological phosphorus removal (EBPR) processes. Distinctive Raman spectra of polyphosphates allowed for both identification of PAOs and quantification of intracellular polyphosphate during various metabolic phases in a lab-scale EBPR process. Observation of polyphosphate at individual cell level indicated thatthere are distributed states of cells in terms of polyphosphate content at any given time, suggesting that agent-based distributive modeling would more accurately reflect the behavior of an EBPR process than the traditional average-state based modeling. The results, for the first time, showed that the polyphosphate depletion or replenishment observed at the overall population level were collective results from shifts/transition in the distribution of abundance of PAOs with different amounts of polyphosphate inclusions during EBPR. Imaging construction based on simultaneous quantification of intracellular polyphosphate and protein revealed the spatial distribution of polyphosphate inside cells and showed that the polyphosphates accumulate in smaller or larger aggregates, rather than being evenly distributed within the cytoplasm. The results demonstated that Raman microscopy will allow for detailed cellular-level evaluation of polyphosphate metabolism and dynamics in EBPR processes and revealed mechanism insights, which otherwise would not be obtained using a traditional bulk measurement-based approach.