Effect of hydraulic regime on sulfur-packed bed performance: Denitrification and disproportionation.

Sulfur-packed beds (SPBs) have been increasingly incorporated into constructed wetland systems to overcome limitations in achieving satisfactory nitrate removal efficiency. However, the underlying impact of hydraulic regimes on SPB performance remains understudied. This study investigated the performance of a pilot-scale SPB, encompassing sulfur autotrophic denitrification (SAD) and sulfur disproportionation (SDP) processes, under various horizontal flow (HF) and vertical flow (VF) regimes. The HF regime exhibited superior SAD efficiency, achieving 3.1-4.4 mg-N/L of nitrate removal compared to 0.9-2.8 mg-N/L under VF regimes. However, greater sulfide production of 3.8-5.6 mg/L was observed, in contrast to only 1.5-2.3 mg/L under VF regimes when SDP occurred. Employing current computational fluid dynamics simulations could predict general regimes but lacked precision in detailing sulfur layer dynamics. In contrast, determining the spatial distribution of SAD substrates and SDP products offered a viable solution, revealing stagnate, short-circuit, and back flows. Moreover, the feasibility of an aeration approach to reduce sulfide emissions below 0.5 mg/L in case of accidental SDP occurrence was confirmed. This study offers a method for assessing detailed hydraulic regimes within SPBs. Additionally, it provides guidance on optimizing the packing of sulfur-based materials when implementing SPBs in constructed wetland systems and presents a strategy for mitigating excessive sulfide emissions.

Biotransformation of 6:2 Fluorotelomer Thioether Amido Sulfonate in Aqueous Film-Forming Foams under Nitrate-Reducing Conditions.

Despite the prevalence of nitrate reduction in groundwater, the biotransformation of per- and polyfluoroalkyl substances (PFAS) under nitrate-reducing conditions remains mostly unknown compared with aerobic or strong reducing conditions. We constructed microcosms under nitrate-reducing conditions to simulate the biotransformation occurring at groundwater sites impacted by aqueous film-forming foams (AFFFs). We investigated the biotransformation of 6:2 fluorotelomer thioether amido sulfonate (6:2 FtTAoS), a principal PFAS constituent of several AFFF formulations using both quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) and qualitative high-resolution mass spectrometry analyses. Our results reveal that the biotransformation rates of 6:2 FtTAoS under nitrate-reducing conditions were about 10 times slower than under aerobic conditions, but about 2.7 times faster than under sulfate-reducing conditions. Although minimal production of 6:2 fluorotelomer sulfonate and the terminal perfluoroalkyl carboxylate, perfluorohexanoate was observed, fluorotelomer thioether and sulfinyl compounds were identified in the aqueous samples. Evidence for the formation of volatile PFAS was obtained by mass balance analysis using the total oxidizable precursor assay and detection of 6:2 fluorotelomer thiol by gas chromatography-mass spectrometry. Our results underscore the complexity of PFAS biotransformation and the interactions between redox conditions and microbial biotransformation activities, contributing to the better elucidation of PFAS environmental fate and impact.

The co-existence of anammox genera in an expanded granular sludge bed reactor with biomass carriers for nitrogen removal.

Anaerobic ammonium oxidation (anammox)-based nitrogen removal saves aeration energy and organic carbon costs, attributed to its anaerobic and autotrophic nature. However, due to the slow growth of anaerobic ammonium oxidation bacteria (AnAOB), drawbacks including long startup time and sensitivity to toxins still hamper the application of anammox-based processes. To cope with the slow growth of AnAOB, various bioreactor configurations have been investigated for the capability of retaining anammox biomass, among which, the expanded granular sludge bed (EGSB) reactor is a promising option. In this study, two laboratory-scale EGSB reactors were used to gain insights of microbial population and their response to amending biofilm-carriers, aiming to enhance the biomass retention of AnAOB. The respective ammonium and nitrite removal efficiencies were up to over 90%, and the overall nitrogen removal efficiency (NRE) was stable at over 70%, in the EGSB reactor amended with carriers (CEGSB). Compared to the control EGSB, CEGSB's observed performance was more stable during the 236-day operational period. The abundance of AnAOB reached 22% in the EGSB and 49% in the CEGSB. It was also observed that Ca. Brocadia (14.25%) and Asahi BRW2 (33.19%) coexisted in the CEGSB. The dynamics of major metabolisms and functional genes involved in nitrogen conversion were further observed by FAPROTAX based on the taxonomic data, providing more insights into the functions of the microbial communities.

Concomitant Leaching and Electrochemical Extraction of Rare Earth Elements from Monazite.

Rare earth elements (REEs) have become increasingly important in modern day technologies. Unfortunately, their recycling is currently limited, and the conventional technologies for their extraction and purification are exceedingly energy and chemical intensive. New sustainable technologies for REE extraction from both primary and secondary resources would be extremely beneficial. This research investigated a two-stage recovery strategy focused on the recovery of neodymium (Nd) and lanthanum (La) from monazite ore that combines microbially based leaching (using citric acid and spent fungal supernatant) with electrochemical extraction. Pretreating the phosphate-based monazite rock (via roasting) dramatically increased the microbial REE leaching efficiency. Batch experiments demonstrated the effective and continued leaching of REEs by recycled citric acid, with up to 392 mg of Nd L-1 and 281 mg of La L-1 leached during seven consecutive 24 h cycles. Neodymium was further extracted in the catholyte of a three-compartment electrochemical system, with up to 880 mg of Nd L-1 achieved within 4 days (at 40 A m-2). Meanwhile, the radioactive element thorium and counterions phosphate and citrate were separated effectively from the REEs in the anolyte, favoring REE extraction and allowing sustainable reuse of the leaching agent. This study shows a promising technology that is suitable for primary ores and can further be optimized for secondary resources.

Incomplete Wood-Ljungdahl pathway facilitates one-carbon metabolism in organohalide-respiring Dehalococcoides mccartyi.

The acetyl-CoA "Wood-Ljungdahl" pathway couples the folate-mediated one-carbon (C1) metabolism to either CO2 reduction or acetate oxidation via acetyl-CoA. This pathway is distributed in diverse anaerobes and is used for both energy conservation and assimilation of C1 compounds. Genome annotations for all sequenced strains of Dehalococcoides mccartyi, an important bacterium involved in the bioremediation of chlorinated solvents, reveal homologous genes encoding an incomplete Wood-Ljungdahl pathway. Because this pathway lacks key enzymes for both C1 metabolism and CO2 reduction, its cellular functions remain elusive. Here we used D. mccartyi strain 195 as a model organism to investigate the metabolic function of this pathway and its impacts on the growth of strain 195. Surprisingly, this pathway cleaves acetyl-CoA to donate a methyl group for production of methyl-tetrahydrofolate (CH3-THF) for methionine biosynthesis, representing an unconventional strategy for generating CH3-THF in organisms without methylene-tetrahydrofolate reductase. Carbon monoxide (CO) was found to accumulate as an obligate by-product from the acetyl-CoA cleavage because of the lack of a CO dehydrogenase in strain 195. CO accumulation inhibits the sustainable growth and dechlorination of strain 195 maintained in pure cultures, but can be prevented by CO-metabolizing anaerobes that coexist with D. mccartyi, resulting in an unusual syntrophic association. We also found that this pathway incorporates exogenous formate to support serine biosynthesis. This study of the incomplete Wood-Ljungdahl pathway in D. mccartyi indicates a unique bacterial C1 metabolism that is critical for D. mccartyi growth and interactions in dechlorinating communities and may play a role in other anaerobic communities.

Bioleaching of rare earth elements from monazite sand.

Three fungal strains were found to be capable of bioleaching rare earth elements from monazite, a rare earth phosphate mineral, utilizing the monazite as a phosphate source and releasing rare earth cations into solution. These organisms include one known phosphate solubilizing fungus, Aspergillus niger ATCC 1015, as well as two newly isolated fungi: an Aspergillus terreus strain ML3-1 and a Paecilomyces spp. strain WE3-F. Although monazite also contains the radioactive element Thorium, bioleaching by these fungi preferentially solubilized rare earth elements over Thorium, leaving the Thorium in the solid residual. Adjustments in growth media composition improved bioleaching performance measured as rare earth release. Cell-free spent medium generated during growth of A. terreus strain ML3-1 and Paecilomyces spp. strain WE3-F in the presence of monazite leached rare earths to concentrations 1.7-3.8 times those of HCl solutions of comparable pH, indicating that compounds exogenously released by these organisms contribute substantially to leaching. Organic acids released by the organisms included acetic, citric, gluconic, itaconic, oxalic, and succinic acids. Abiotic leaching with laboratory prepared solutions of these acids was not as effective as bioleaching or leaching with cell-free spent medium at releasing rare earths from monazite, indicating that compounds other than the identified organic acids contribute to leaching performance.

BASALT refines binning from metagenomic data and increases resolution of genome-resolved metagenomic analysis.

Metagenomic binning is an essential technique for genome-resolved characterization of uncultured microorganisms in various ecosystems but hampered by the low efficiency of binning tools in adequately recovering metagenome-assembled genomes (MAGs). Here, we introduce BASALT (Binning Across a Series of Assemblies Toolkit) for binning and refinement of short- and long-read sequencing data. BASALT employs multiple binners with multiple thresholds to produce initial bins, then utilizes neural networks to identify core sequences to remove redundant bins and refine non-redundant bins. Using the same assemblies generated from Critical Assessment of Metagenome Interpretation (CAMI) datasets, BASALT produces up to twice as many MAGs as VAMB, DASTool, or metaWRAP. Processing assemblies from a lake sediment dataset, BASALT produces ~30% more MAGs than metaWRAP, including 21 unique class-level prokaryotic lineages. Functional annotations reveal that BASALT can retrieve 47.6% more non-redundant opening-reading frames than metaWRAP. These results highlight the robust handling of metagenomic sequencing data of BASALT.

Double-edged sword effects of dissimilatory nitrate reduction to ammonium (DNRA) bacteria on anammox bacteria performance in an MBR reactor.

Dissimilatory nitrate reduction to ammonium (DNRA) bacteria imposing double-edged sword effects on anammox bacteria were investigated in an anammox-membrane bioreactor (MBR) experiencing an induced crash-recovery event. During the experiment, the anammox-MBR was loaded with NH4+-N:NO2--N ratios (RatioNH4+-N: NO2--N) of 1.20-1.60. Initially, the anammox-MBR removed over 95% of 100 mg/L NH4+-N and 132 mg/L NO2--N (RatioNH4+-N: NO2--N = 0.76, the well accepted stoichiometric RatioNH4+-N: NO2--N for anammox) in the influent (Stage 0). Then, we induced a system crash-recovery event via nitrite shock loadings to better understand responses from different guilds of bacteria in anammox-MBR, loaded with 1.60 RatioNH4+-N: NO2--N with 100 mg/L NO2--N in the influent (Stage 1). Interestingly, the nitrogen removal by anammox bacteria was maintained for about 20 days before starting to decrease significantly. In Stage 2, we further increased influent nitrite concentration to 120 mg/L (1.33 RatioNH4+-N: NO2--N) to simulate a high nitrite toxicity scenario for a short period of time. As expected, nitrogen removal efficiency dropped to only 16.8%. After the induced system crash, anammox-MBR performance recovered steadily to 93.2% nitrogen removal with a 1.25 RatioNH4+-N:NO2--N and a low nitrite influent concentration of 80 mg/L NO2--N. Metagenomics analysis revealed that a probable causality of the decreasing nitrogen removal efficiency in Stage 1 was the overgrowth of DNRA-capable bacteria. The results showed that the members within the Ignavibacteriales order (21.7%) out competed anammox bacteria (17.0%) in the anammox-MBR with elevated nitrite concentrations in the effluent. High NO2--N loading (120 mg N/L) further caused the predominant Candidatus Kuenenia spp. were replaced by Candidatus Brocadia spp. Therefore, it was evident that DNRA bacteria posed negative effects on anammox with 1.60 RatioNH4+-N: NO2--N. Also, when 120 mg/L NO2--N fed to anammox-MBR (RatioNH4+-N: NO2--N = 1.33), canonical denitrification became the primary nitrogen sink with both DNRA and anammox activities decreased. They probably fed on lysed microbial cells of anammox and DNRA. In Stage 3, a low RatioNH4+-N: NO2--N (1.25) with 80 mg/L NO2--N was used to rescue the system, which effectively promoted DNRA-capable bacteria growth. Although anammox bacteria's abundance was only 7.7% during this stage, they could be responsible for about 90% of the total nitrogen removal during this stage.

The impact of perfluorooctanoic acid shock on hydrogen-driven nitrate and arsenate removal.

Perfluorooctanoic acid (PFOA) is a type of toxic per- and poly-fluoroalkyl substance (PFAS) commonly found in groundwater due to its use in firefighting and industrial applications. The main purpose of this study was to investigate the influence of PFOA shock on the biological performance of a hydrogen-driven bioreactor for nitrate and arsenate removal. Four hydrogen-driven removal reactors (HdBRs) used for the simultaneous removal of nitrate and arsenal were operated with concentrations of either 0, 1, 5, and 10 mg/L of PFOA to induce shock on the systems and examine the corresponding bacterial response. Our results showed that PFOA shock inhibited and decreased the maximum hydrogen-driven arsenate removal rate. Principal Component Analysis (PCA) confirmed that this performance decrease occurred due to a bacterial strike triggered by PFOA shock. PFOA toxicity also led to protein secretion and sludge density decreases. Bacterial analyses showed shifts in the community population due to PFOA shock. The dominant bacteria phylum Proteobacteria became more abundant, from 41.24% originally to 48.29% after exposure to 10 mg/L of PFOA. Other phyla, such as Euryarchaeota and Bacteroidetes, were more tolerant to PFOA shock. Although some of the predominant species within the sludge of each HdBR exhibited a decline, other species with similar functions persisted and assumed the functional responsibilities previously held by the dominant species.

Managing microbial sulfur disproportionation for optimal sulfur autotrophic denitrification in a pilot-scale elemental sulfur packed-bed bioreactor.

Elemental sulfur packed-bed (S0PB) bioreactors for autotrophic denitrification have gained more attention in wastewater treatment due to their organic carbon-free operation, low operating cost, and minimal carbon emissions. However, the rapid development of microbial S0-disproportionation (MS0D) in S0PB reactor during deep denitrification poses a significant drawback to this new technology. MS0D, the process in which sulfur is used as both an electron donor and acceptor by bacteria, plays a crucial role in the microbial-driven sulfur cycle but remains poorly understood in wastewater treatment setups. In this study, we induced MS0D in a pilot-scale S0PB reactor capable of denitrifying over 1000 m3/d nitrate-containing wastewater. Initially, the S0PB reactor stably removed 6.6 mg-NO3--N/L nitrate at an empty bed contact time (EBCT) of 20 mins, which was designated the S0-denitrification stage. To induce MS0D, we reduced the influent nitrate concentrations to allow deep nitrate removal, resulted in the production of large quantities of sulfate and sulfide (SO42-:S2- 3.2 w/w). Meanwhile, other sulfur-heterologous electron acceptors (SHEAs), e.g., nitrite and DO, were also kept at trace levels. The negative correlations between the SHEAs concentrations and the sulfide productions indicated that the absence of SHEAs was a primary inducing factor to MS0D. The microbial community drastically diverged in response to the depletion of SHEAs during the switch from S0-denitrification to S0-disproportionation. An evident enrichment of sulfur-disproportionating bacteria (SDBs) was found at the S0-disproportionation stage, accompanied by the decline of sulfur-oxidizing bacteria (SOBs). In the end, we discovered that shortening the EBCT and increasing the reflux ratio could inhibit sulfide production by reducing it from 43.9 mg/L to 3.2 mg/L or 25.5 mg/L. In conclusion, our study highlights the importance of considering MS0D when designing and optimizing S0PB reactors for sustainable autotrophic sulfur denitrification in real-life applications.

Dissimilatory sulfate reduction in an anaerobic biofilm reactor for tofu processing wastewater treatment: Bacterial community and their functional genes.

Dissimilatory sulfate reduction (DSR) is the key sulfur cycle that transforms sulfate to sulfide. This process leads to odour issues in wastewater treatment. However, few studies have focused on DSR during treating food processing wastewater with high sulfate. This study investigated DSR microbial population and functional genes in an anaerobic biofilm reactor (ABR) treating tofu processing wastewater. The tofu processing wastewater is a common food processing wastewater in Asia. The full-scale ABR was operated for over 120 days in a tofu and tofu-related products manufacturing factory. Mass balance calculations based on the reactor performance indicated that 79.6-85.1 % of the sulfate was transformed into sulfide irrelevant to dissolved oxygen supplementation. Metagenomic analysis revealed 21 metagenome-assembled genomes (MAGs) containing enzymes encoding DSR. The biofilm contained the complete functional genes of DSR pathway in the full-scale ABR, indicating that biofilm could process DSR independently. Comamonadaceae, Thiobacillus, Nitrosomonadales, Desulfatirhabdium butyrativorans, Desulfomonile tiedjei were the dominant DSR species in the ABR biofilm community. Dissolved oxygen supplementation directly inhibited DSR and mitigated HS- production. It was also found that Thiobacillus contained all the function genes encoding every necessary enzyme in DSR, and thus Thiobacillus distribution directly correlated to DSR and the ABR performance.

Glyoxylate metabolism is a key feature of the metabolic degradation of 1,4-dioxane by Pseudonocardia dioxanivorans strain CB1190.

The groundwater contaminant 1,4-dioxane (dioxane) is transformed by several monooxygenase-expressing microorganisms, but only a few of these, including Pseudonocardia dioxanivorans strain CB1190, can metabolize the compound as a sole carbon and energy source. However, nothing is yet known about the genetic basis of dioxane metabolism. In this study, we used a microarray to study differential expression of genes in strain CB1190 grown on dioxane, glycolate (a previously identified intermediate of dioxane degradation), or pyruvate. Of eight multicomponent monooxygenase gene clusters carried by the strain CB1190 genome, only the monooxygenase gene cluster located on plasmid pPSED02 was upregulated with dioxane relative to pyruvate. Plasmid-borne genes for putative aldehyde dehydrogenases, an aldehyde reductase, and an alcohol oxidoreductase were also induced during growth with dioxane. With both dioxane and glycolate, a chromosomal gene cluster encoding a putative glycolate oxidase was upregulated, as were chromosomal genes related to glyoxylate metabolism through the glyoxylate carboligase pathway. Glyoxylate carboligase activity in cell extracts from cells pregrown with dioxane and in Rhodococcus jostii strain RHA1 cells expressing the putative strain CB1190 glyoxylate carboligase gene further demonstrated the role of glyoxylate metabolism in the degradation of dioxane. Finally, we used (13)C-labeled dioxane amino acid isotopomer analysis to provide additional evidence that metabolites of dioxane enter central metabolism as three-carbon compounds, likely as phosphoglycerate. The routing of dioxane metabolites via the glyoxylate carboligase pathway helps to explain how dioxane is metabolized as a sole carbon and energy source for strain CB1190.

Dissimilatory sulfate reduction in the cake layer of a full-scale anaerobic dynamic membrane bioreactor for hotel laundry wastewater treatment: Bacterial community and functional genes.

Dissimilatory sulfate reduction (DSR) in cake layer of full-scale anaerobic dynamic membrane bioreactor for treating hotel laundry wastewater was studied. Change (Δ) of sulfate concentration (ΔSO42-) was positively correlated to dynamic cake layer (DCL) development, while ΔS2- was negatively correlated. ΔSO32- and ΔSorganic sulfur remained around 1.5-2.5 and 1.2-2.3 mg-S/L, respectively. Thus, DSR was the predominant sulfate reduction process in DCL. 33 binned genomes from DCL microbiome samples possessed one or more DSR functional genes. But only four binned genomes possess all functional genes, and thus can achieve complete DSR. However, no significant variations of these DSR bacteria was obseared during DCL development. Metagenomic analysis predicted that sulfate reduction in DCL was mainly carried out by collaborations between bacteria with incomplete DSR pathways. Among which, sulfite â†’ sulfide by dissimilatory-sulfite-reductase expression bacteria was the key process. Overall results suggested that controlling dissimilatory-sulfite-reductase activities could prevent sulfide buildup in the effluent.

Core carbon fixation pathways associated with cake layer development in an anoxic-oxic biofilm-membrane bioreactor treating textile wastewater.

Microbial carbon fixation pathways have not yet been adequately understood for their role in membrane case layer formation processes. Carbon fixation bacteria can play critical roles in either causing or enhancing cake layer formation in some autotrophic-prone anoxic conditions, such as sulfur-cycling conditions. Understanding the microbes capable of carbon fixation can potentially guide the design of membrane biofouling mitigation strategies in scientific ways. Thus, we used meta-omics methods to query carbon fixation pathways in the cake layers of a full-scale anoxic-oxic biofilm-MBR system treating textile wastewater in this study. Based on the wastewater constituents and other properties, such as anoxic conditions, sulfide-reducing and sulfur-oxidizing bacteria could co-exist in the membrane unit. In addition, low-light radiation conditions could also happen to the membrane unit. However, we could not quantify the light intensity or total energy input accurately because the whole experimental setup was a full-scale system. Potentially complete carbon fixation pathways in the cake layer included the Calvin-Benson-Bassham cycle, Wood-Ljungdahl pathway, and the 3-hydroxypropionate bicycle. We discovered that using aeration could effectively inhibit carbon fixation, which resulted in mitigating membrane cake layer development. However, the aeration resulted in the 3-hydroxypropionate bicycle pathway, presumably used by aerobic sulfur-oxidizing prokaryotes, to become a more abundant carbon fixation pathway in the cake layer under aerobic conditions.

Identification of dissimilatory nitrate reduction to ammonium (DNRA) and denitrification in the dynamic cake layer of a full-scale anoixc dynamic membrane bioreactor for treating hotel laundry wastewater.

Identification of dissimilatory nitrate reduction to ammonium (DNRA) and denitrification in the dynamic cake layer of a full-scale anoixc dynamic membrane bioreactor (AnDMBR) for treating hotel laundry wastewater was studied. A series of experiments were conducted to understand the contributions of DNRA and canonical denitrification activities in the dynamic cake layer of the AnDMBR. The dynamic cake layer developed included two phases - a steady transmembrane pressure (TMP) increase at 0.24 kPa/day followed by a sharp TMP jump at 1.26 kPa/day four to five days after the AnDMBR start-up. The nitrogen mass balance results showed that canonical denitrification was predominant during the development of the dynamic cake layer. However, DNRA activity and accumulation of bacteria equipped with a complete DNRA pathway showed a positive correlation to the development of the dynamic cake layer. Our metagenomic analysis identified an approximately 18% of the dynamic cake layer bacterial community has a complete DNRA pathway. Pannonibacter (1%), Thauera (0.8%) and Pseudomonas (3%) contained all genes encoding for funcional enzymes of both DNRA (nitrate reductase and DNRA nitrite reductase) and denitrification (nitrate reductase, nitrous oxide reductase and nitric oxide reductase). No other metagenome-assembled genomes (MAGs) possessed a complete cononical denitrification pathway, indicating food-chain-like interactions of denitrifiers in the dynamic cake layer. We found that COD loading rate could be used to control DNRA and canonical denitrification activities during the dynamic cake layer formation.

Carbon flow of heliobacteria is related more to clostridia than to the green sulfur bacteria.

The recently discovered heliobacteria are the only Gram-positive photosynthetic bacteria that have been cultured. One of the unique features of heliobacteria is that they have properties of both the photosynthetic green sulfur bacteria (containing the type I reaction center) and Clostridia (forming heat-resistant endospores). Most of the previous studies of heliobacteria, which are strict anaerobes and have the simplest known photosynthetic apparatus, have focused on energy and electron transfer processes. It has been assumed that like green sulfur bacteria, the major carbon flow in heliobacteria is through the (incomplete) reductive (reverse) tricarboxylic acid cycle, whereas the lack of CO(2)-enhanced growth has not been understood. Here, we report studies to fill the knowledge gap of heliobacterial carbon metabolism. We confirm that the CO(2)-anaplerotic pathway is active during phototrophic growth and that isoleucine is mainly synthesized from the citramalate pathway. Furthermore, to our surprise, our results suggest that the oxidative (forward) TCA cycle is operative and more active than the previously reported reductive (reverse) tricarboxylic acid cycle. Both isotopomer analysis and activity assays suggest that citrate is produced by a putative (Re)-citrate synthase and then enters the oxidative (forward) TCA cycle. Moreover, in contrast to (Si)-citrate synthase, (Re)-citrate synthase produces a different isomer of 2-fluorocitrate that is not expected to inhibit the activity of aconitase.

Selective utilization of exogenous amino acids by Dehalococcoides ethenogenes strain 195 and its effects on growth and dechlorination activity.

Bacteria of the genus Dehalococcoides are important members of bioremediation communities because of their ability to detoxify chloroethenes to the benign end product ethene. Genome-enabled studies conducted with Dehalococcoides ethenogenes 195 have revealed that two ATP-binding cassette (ABC)-type amino acid transporters are expressed during its exponential growth stages. In light of previous findings that Casamino Acids enhanced its dechlorination activity, we hypothesized that strain 195 is capable of importing amino acids from its environment to facilitate dechlorination and growth. To test this hypothesis, we applied isotopomer-based dilution analysis with (13)C-labeled acetate to differentiate the amino acids that were taken up by strain 195 from those synthesized de novo and to determine the physiological changes caused by the significantly incorporated amino acids. Our results showed that glutamate/glutamine and aspartate/asparagine were almost exclusively synthesized by strain 195, even when provided in excess in the medium. In contrast, phenylalanine, isoleucine, leucine, and methionine were identified as the four most highly incorporated amino acids, at levels >30% of respective proteinogenic amino acids. When either phenylalanine or all four highly incorporated amino acids were added to the defined mineral medium, the growth rates, dechlorination activities, and yields of strain 195 were enhanced to levels similar to those observed with supplementation with 20 amino acids. However, genes for the putative ABC-type amino acids transporters and phenylalanine biosynthesis exhibited insignificant regulation in response to the imported amino acids. This study also demonstrates that using isotopomer-based metabolite analysis can be an efficient strategy for optimizing nutritional conditions for slow-growing microorganisms.

Challenges in biodegradation of non-degradable thermoplastic waste: From environmental impact to operational readiness.

Non-degradable plastics such as polyethylene (PE), polypropylene (PP), polystyrene (PS), and polyethylene terephthalate (PET) are among the most generated plastic wastes in municipal and industrial waste streams. The mismanagement of abandoned plastics and toxic plastic additives have threatened marine and land fauna as well as human beings for several decades. The available thermal processes can degrade plastic at pilot- and commercial-scale. However, they are energy-intensive and can generate toxic gases. Degradation of plastic waste with the help of live microorganisms (biodegradation) is an eco- and environmentally friendly method for plastic degradation, although the slow processing time and low degradation rate still hinder its applications at pilot- and large-scale. In this review, the advantages and limitations of current plastic degradation methods, their technology readiness levels (TRL), biodegradation mechanisms and the associated challenges in biodegradation are assessed in detail. Based on this analysis, a path toward an efficient and greener way toward degradation of non-recyclable single-use PE, PP, PS and PET plastic is proposed.

Degradation of plastic waste using stimulated and naturally occurring microbial strains.

The capability of different strains derived from soil, activated sludge, farm sludge, and worms' excreta were investigated for biodegradation of high-density polyethylene, polystyrene foam, polypropylene and polyethylene terephthalate in unstimulated and stimulated conditions. Biodegradation using naturally occurring microbial strains examined in mixed (270 days) and individual (100 days) systems, while H2O2 stimulated strains were tested only in the mixed system (30 days). Penicillium raperi, Aspergillus flavus, Penicillium glaucoroseum and Pseudomonas sp. were isolated as the most plastic degrading microbes. Maximum weight loss was seen by incubation of polyethylene with Aspergillus flavus (5.5%) in unstimulated mix condition. Fourier Transform Infrared Spectroscopy (FT-IR) revealed formation of new functional groups as hydroxyl, carbonyl, alkene and alkoxy in the treated plastics. Visualisation of plastics by optical, atomic force (AFM) and electron microscopy (SEM) were also illustrated biodegradation. The derived by-products from microbial degradation was tested, and found no inhibition on microbial growth and performance.

Assessing effects of Ca2+ addition on membrane bioreactor performance and macro-floc sludge characteristics.

Calcium ions (Ca2+) can trigger coagulation-flocculation process to form macro-flocculated sludge (MFS). Thus, dosing Ca2+-containing reagents into membrane bioreactors (MBRs) is considered as a promising approach to mitigate membrane biofouling. However, a mechanistic understanding of Ca2+ addition to MBR performance remains elucidated, such as physicochemical characteristics of MFS and their functionality variations. Consequently, this study was sought to understand the interplays of Ca2+ addition and MBR performance with a focus on characterizing MFS in detail. Three parallel MBRs were amended with 82, 208 and 410 mg-Ca2+/L final concentrations. Particle size analyses revealed that MFS formation was overall enhanced by the Ca2+ addition and granular sludge with diameters of up to 900 µm was formed in the 410 mg-Ca2+/L scenario. We believed that cationic bridges facilitated by elevated Ca2+ concentrations in conjunction with coagulation-flocculation were primary mechanisms of the formation of large flocs. Moreover, significant portions of soluble proteins and polysaccharides were flocculated and precipitated by Ca2+, which demonstrated a negative correlation between extracellular polymeric substances (EPS) concentrations and the formation of MFS. Furthermore, the population abundancies of Thiotrichaceae, Sphingomonadales and Hyphomicrobiaceae decreased in the sludge with Ca2+ addition resulted in profound changes of the microbial communities in the MBRs. But MBR performance, such as chemical oxygen demand removal (over 90%), showed no variation during the MBR operation. On the contrary, total nitrogen removal was inhibited in the MBRs. It was because the enlarging MFS formed diffusion barriers to prevent organic component from entering into the sludge flocs to be consumed.