Electrochemical-coupled sulfur-driven autotrophic denitrification for nitrogen removal from raw landfill leachate: Evaluation of performance and mechanisms.

The cost-effective and environment-friendly sulfur-driven autotrophic denitrification (SdAD) process has drawn significant attention for advanced nitrogen removal from low carbon-to-nitrogen (C/N) ratio wastewater in recent years. However, achieving efficient nitrogen removal and maintaining system stability of SdAD process in treating low C/N landfill leachate treatment have been a major challenge. In this study, a novel electrochemical-coupled sulfur-driven autotrophic denitrification (ESdAD) system was developed and compared with SdAD system through a long-term continuous study. Superior nitrogen removal performance (removal efficiency of 89.1 ± 2.5 %) was achieved in ESdAD system compared to SdAD process when treating raw landfill leachate (influent total nitrogen (TN) concentration of 241.7 ± 36.3 mg-N/L), and the effluent TN concentration of ESdAD bioreactor was as low as 24.8 ± 5.1 mg-N/L, which meets the discharge standard of China (< 40 mg N/L). Moreover, less sulfate production rate (1.3 ± 0.2 mg SO42--S/mgNOx--N vs 1.7 ± 0.2 mg SO42--S/mgNOx--N) and excellent pH modulation (pH of 6.9 ± 0.2 vs 5.8 ± 0.4) were also achieved in the ESdAD system compared to SdAD system. The improvement of ESdAD system performance was contributed to coexistence and interaction of heterotrophic bacteria (e.g., Rhodanobacter, Thermomonas, etc.), sulfur autotrophic bacteria (e.g., Thiobacillus, Sulfurimonas, Ignavibacterium etc.) and hydrogen autotrophic bacteria (e.g., Thauera, Comamonas, etc.) under current stimulation. In addition, microbial nitrogen metabolic activity, including functional enzyme (e.g., Nar and Nir) activities and electron transfer capacity of extracellular polymeric substances (EPS) and cytochrome c (Cyt-C), were also enhanced during current stimulation, which facilitated the nitrogen removal and maintained system stability. These findings suggested that ESdAD is an effective and eco-friendly process for advanced nitrogen removal for low C/N wastewater.

Bioponic systems with biochar: Insights into nutrient recovery, heavy metal reduction, and microbial interactions in digestate-based bioponics.

Bioponics is a nutrient-recovery technology that transforms nutrient-rich organic waste into plant biomass/bioproducts. Integrating biochar with digestate from anaerobic wastewater treatment process can improve resource recovery while mitigating heavy metal contamination. The overarching goal of this study was to investigate the application of biochar in digestate-based bioponics, focusing on its efficacy in nutrient recovery and heavy metal removal, while also exploring the microbial community dynamics. In this study, biochar was applied at 50 % w/w with 500 g dry weight of digestate during two 28-day crop cycles (uncontrolled pH and pH 5.5) using white stem pak choi (Brassica rapa var. chinensis) as a model crop. The results showed that the digestate provided sufficient phosphorus and nitrogen, supporting plant growth. Biochar amendment improved plant yield and phosphate solubilization and reduced nitrogen loss, especially at the pH 5.5. Furthermore, biochar reduced the heavy metal accumulation in plants, while concentrating these metals in the residual sludge. However, owing to potential non-carcinogenic and carcinogenic health risks, it is still not recommended to directly consume plants cultivated in digestate-based bioponic systems. Additionally, biochar amendment exhibited pronounced impact on the microbial community, promoting microbes responsible for nutrient solubilization and cycling (e.g., Tetrasphaera, Herpetosiphon, Hyphomicrobium, and Pseudorhodoplanes) and heavy metal stabilization (e.g., Leptolinea, Fonticella, Romboutsia, and Desulfurispora) in both the residual sludge and plants. Overall, the addition of biochar enhanced the microbial community and facilitated the metal stabilization and the cycling of nutrients within both residual sludge and root systems, thereby improving the overall efficiency of the bioponics.

Nanobubbles in water and wastewater treatment systems: Small bubbles making big difference.

Since the discovery of nanobubbles (NBs) in 1994, NBs have been attracting growing attention for their fascinating properties and have been studied for application in various environmental fields, including water and wastewater treatment. However, despite the intensive research efforts on NBs' fundamental properties, especially in the past five years, controversies and disagreements in the published literature have hindered their practical implementation. So far, reviews of NB research have mainly focused on NBs' role in specific treatment processes or general applications, highlighting proof-of-concept and success stories primarily at the laboratory scale. As such, there lacks a rigorous review that authenticates NBs' potential beyond the bench scale. This review aims to provide a comprehensive and up-to-date analysis of the recent progress in NB research in the field of water and wastewater treatment at different scales, along with identifying and discussing the challenges and prospects of the technology. Herein, we systematically analyze (1) the fundamental properties of NBs and their relevancy to water treatment processes, (2) recent advances in NB applications for various treatment processes beyond the lab scale, including over 20 pilot and full-scale case studies, (3) a preliminary economic consideration of NB-integrated treatment processes (the case of NB-flotation), and (4) existing controversies in NBs research and the outlook for future research. This review is organized with the aim to provide readers with a step-by-step understanding of the subject matter while highlighting key insights as well as knowledge gaps requiring research to advance the use of NBs in the wastewater treatment industry.

Evaluation of Nanaerobic Digestion as a Mechanism to Explain Surplus Methane Production in Animal Rumina and Engineered Digesters.

Nanaerobes are a newly described class of microorganisms that use a unique cytochrome bd oxidase to achieve nanaerobic respiration at <2 µM dissolved oxygen (∼1% of atmospheric oxygen) but are not viable above this value due to the lack of other terminal oxidases. Although sharing an overlapping ecological niche with methanogenic archaea, the role of nanaerobes in methanogenic systems has not been studied so far. To explore their occurrence and significance, we re-analyzed published meta-omic datasets from animal rumina and waste-to-energy digesters, including conventional anaerobic digesters and anaerobic digesters with ultra-low oxygenation. Results show that animal rumina share broad similarities in the microbial community and system performance with oxygenated digesters, rather than with conventional anaerobic digesters, implying that trace levels of oxygen drive the efficient digestion in ruminants. The rumen system serves as an ideal model for the newly named nanaerobic digestion, as it relies on the synergistic co-occurrence of nanaerobes and methanogens for methane yield enhancement. The most abundant ruminal bacterial family Prevotellaceae contains many nanaerobes, which perform not only anaerobic fermentation but also nanaerobic respiration using cytochrome bd oxidase. These nanaerobes generally accompany hydrogenotrophic methanogens to constitute a thermodynamically and physiologically consistent framework for efficient methane generation. Our findings provide new insights into ruminal methane emissions and strategies to enhance methane generation from biomass.

Effect of zero-valent iron nanoparticles on taxonomic composition and hydrogen production from kitchen waste.

This study investigated the effects of varying zero-valent iron (ZVI) (0 to 5,000 mg/L) on fermentative hydrogen (H2) production, metabolic pattern, and taxonomic profile by using kitchen waste as substrate. The study demonstrated that the supplementation of 500 mg ZVI/L resulted in the highest H2 yield (219.68 ± 11.19 mL H2/g-volatile solids (VS)added), which was 19% higher than the control. The metabolic pattern analysis showed that acetic and butyric acid production primarily drove the H2 production. The taxonomic analysis further revealed that Firmicutes (relative abundance (RA): 80-96%) and Clostridium sensu stricto 1 (RA: 68-88%) were the dominant phyla and genera, respectively, during the exponential gas production phase, supporting the observation of accumulation of acetic and butyric acids. These findings suggest that supplementation of ZVI can enhance H2 production from organic waste and significantly influence the metabolic pattern and taxonomic profile, including the metalloenzymes.

Flocculation of oleaginous green algae with Mortierella alpina fungi.

Microalgae are promising sources of valuable bioproducts such as biofuels, food, and nutraceuticals. However, harvesting microalgae is challenging due to their small size and low biomass concentrations. To address this challenge, bio-flocculation of starchless mutants of Chlamydomonas reinhardtii (sta6/sta7) was investigated with Mortierella alpina, an oleaginous fungus with high concentrations of arachidonic acid (ARA). Triacylglycerides (TAG) reached 85 % of total lipids in sta6 and sta7 through a nitrogen regime. Scanning electron microscopy determined cell-wall attachment and extra polymeric substances (EPS) to be responsible for flocculation. An algal-fungal biomass ratio around 1:1 (three membranes) was optimal for bio-flocculation (80-85 % flocculation efficiency in 24 h). Nitrogen-deprived sta6/sta7 were flocculated with strains of M. alpina (NVP17b, NVP47, and NVP153) with aggregates exhibiting fatty acid profiles similar to C. reinhardtii, with ARA (3-10 % of total fatty acids). This study showcases M. alpina as a strong bio-flocculation candidate for microalgae and advances a mechanistic understanding of algal-fungal interaction.

Effects of vermicompost leachate on nitrogen, phosphorus, and microbiome in a food waste bioponic system.

Food waste is rich in nutrients, such as nitrogen and phosphorus, and can be integrated with bioponics, a closed-loop agricultural system that combines hydroponics with biological nutrient recovery. Vermicompost leachate (VCL) supplementation has been shown to improve the co-composting of organic waste (i.e., compost quality) and the biodegradation of organic compounds. Thus, VCL has high potential for enhancing nutrient availability in bioponics from food waste. However, the understanding of nitrogen and phosphorus availability in food waste-based bioponics is limited, both with and without VCL. In this study, food waste derived from cafeteria vegetable waste was used as the substrate (500 g dry wt./system) in bioponics to grow lettuce (Lactuca sativa L.) for two consecutive cycles (35 days/cycle) without substrate replacement. VCL was applied weekly (1-5% v/v) and compared to the control without VCL. The results showed that the food waste in bioponics provided nitrogen and phosphorus for plant growth (15.5-65.8 g/lettuce head). Organic-degrading and nutrient-transforming bacteria (Hydrogenispora, Clostridium_sensu_stricto_1, Ruminiclostridium_1, Cellvibrio, Thauera, Hydrogenophaga, and Bacillus) were predominantly found in plant roots and residual food waste. VCL addition significantly increased nitrate, phosphate, and chemical oxygen demand levels in bioponics, owing to the nutrients in VCL and the enhancement of keystone microorganisms responsible for organic degradation and nutrient cycling (e.g., Ellin6067, Actinomyces, and Pirellula). These findings suggest that nitrogen, phosphorus, and organic carbon concentrations in an ecosystem of nutrient-transforming and organic-degrading microbes are key in managing nutrient recovery from food waste in bioponics.

Artificial intelligence and machine learning for smart bioprocesses.

In recent years, the digital transformation of bioprocesses, which focuses on interconnectivity, online monitoring, process automation, artificial intelligence (AI) and machine learning (ML), and real-time data acquisition, has gained considerable attention. AI can systematically analyze and forecast high-dimensional data obtained from the operating dynamics of bioprocess, allowing for precise control and synchronization of the process to improve performance and efficiency. Data-driven bioprocessing is a promising technology for tackling emerging challenges in bioprocesses, such as resource availability, parameter dimensionality, nonlinearity, risk mitigation, and complex metabolisms. This special issue entitled "Machine Learning for Smart Bioprocesses (MLSB-2022)" was conceptualized to incorporate some of the recent advances in applications of emerging tools such as ML and AI in bioprocesses. This VSI: MLSB-2022 contains 23 manuscripts, and summarizes the major findings that can serve as a valuable resource for researchers to learn major advances in applications of ML and AI in bioprocesses.

Use of nanobubble water bioaugmented anaerobically digested sludge for high-efficacy energy production from high-solids anaerobic digestion of corn straw.

An increasing attention has been paid to the secure and sustainable management of agricultural wastes, especially lignocellulosic biomass. Nanobubble water (NBW) contains 106-108 bubbles/mL with diameter <1000 nm. Although previous studies have examined the enhancement effects of NBW on methane production from organic solid wastes, the NBW-based anaerobic digestion (AD) system is still restrained from practical application due to the large increase in AD reactor volume, generation of wastewater, and increase in energy consumption as well. In this study, NBW bioaugmentation of anaerobically digested sludge for the first time was performed for high-solids AD of corn straw. Results show that cellulase, xylanases and lignin peroxidase activities were increased by 2-55% during the NBW bioaugmentation process. Significant enrichment of hydrolytic/acidogenic bacteria and methanogenic archaea were noticed in the NBW bioaugmented sludge. This study clearly demonstrated 47% increase in methane production from high-solids AD of corn straw when O2-NBW bioaugmented sludge was applied, achieving a net energy gain of 5138 MJ/t-volatile solids of corn straw with an energy recovery of 34%. The NBW-based high-solids AD system can provide a novel and sustainable management solution for renewable energy production from agricultural wastes, targeting the reduction of environmental pollution and energy crisis.

Water-energy-greenhouse gas nexus of a novel high-rate activated sludge-two-stage vertical up-flow constructed wetland system for low-carbon wastewater treatment.

Municipal wastewater treatment which is associated with high energy consumption and excessive greenhouse gas (GHG) emissions, has been facing severe challenges toward carbon emissions. In this study, a high-rate activated sludge-two-stage vertical up-flow constructed wetland (HRAS-TVUCW) system was developed to reduce carbon emissions during municipal wastewater treatment. Through carbon management, optimized mass and energy flows were achieved, resulting in high treatment efficiency and low operational energy consumption. The carbon emission of the HRAS-TVUCW system (i.e., 0.21 kg carbon dioxide equivalent/m3 wastewater) was 4.1-folds lower than that of the conventional anaerobic/anoxic/aerobic (A2O) process. Meanwhile, the recovered energy from the HRAS-TVUCW system increased its contribution to carbon neutrality to 40.2%, 4.6-folds higher than that of the A2O process. Results of functional microbial community analysis at the genus level revealed that the controlled dissolved oxygen allocation led to distinctive microbial communities in each unit of HRAS-TVUCW system, which facilitated denitrification efficiency increase and carbon emissions reduction. Overall, the HRAS-TVUCW system could be considered as a cost-effective and sustainable low-carbon technology for municipal wastewater treatment.

Effects of Cu and Zn contamination on chicken manure-based bioponics: Nitrogen recovery, bioaccumulation, microbial community, and health risk assessment.

In bioponics, although chicken manure is an efficient substrate for vegetable production and nitrogen recovery, it is often contaminated with high Cu and Zn levels, which could potentially cause bioaccumulation in plants and pose health risks. The objectives of this study were to assess nitrogen recovery in lettuce- and pak choi-based bioponics with Cu (50-150 mg/kg) and Zn (200-600 mg/kg) supplementation, as well as their bioaccumulation in plants, root microbial community, and health risk assessment. The supplementation of Cu and Zn did not affect nitrogen concentrations and plant growth (p > 0.05) but reduced nitrogen use efficiency. Pak choi showed higher Cu and Zn bioconcentration factors than lettuce. Bacterial genera Ruminiclostridium and WD2101_soil_group in lettuce roots and Mesorhizobium in pak choi roots from Cu and Zn supplemented conditions were significantly higher (p < 0.05) than controls, suggesting microbial biomarkers in plant roots from Cu and Zn exposure bioponics depended on plant type. Health risk assessment herein revealed that consumption of bioponic vegetables with Cu and Zn contamination does not pose long-term health risks (hazard index <1) to children or adults, according to the US EPA. This study suggested that vegetable produced from chicken manure-based bioponics has low health risk in terms of Cu and Zn bioaccumulation and could be applied in commercial-scale system for nutrient recovery from organic waste to vegetable production; however, health risk from other heavy metals and xenobiotic compounds must be addressed.

Microbiological insights into anaerobic digestion for biogas, hydrogen or volatile fatty acids (VFAs): a review.

In the past decades, considerable attention has been directed toward anaerobic digestion (AD), which is an effective biological process for converting diverse organic wastes into biogas, volatile fatty acids (VFAs), biohydrogen, etc. The microbial bioprocessing takes part during AD is of substantial significance, and one of the crucial approaches for the deep and adequate understanding and manipulating it toward different products is process microbiology. Due to highly complexity of AD microbiome, it is critically important to study the involved microorganisms in AD. In recent years, in addition to traditional methods, novel molecular techniques and meta-omics approaches have been developed which provide accurate details about microbial communities involved AD. Better understanding of process microbiomes could guide us in identifying and controlling various factors in both improving the AD process and diverting metabolic pathway toward production of selective bio-products. This review covers various platforms of AD process that results in different final products from microbiological point of view. The review also highlights distinctive interactions occurring among microbial communities. Furthermore, assessment of these communities existing in the anaerobic digesters is discussed to provide more insights into their structure, dynamics, and metabolic pathways. Moreover, the important factors affecting microbial communities in each platform of AD are highlighted. Finally, the review provides some recent applications of AD for the production of novel bio-products and deals with challenges and future perspectives of AD.

Bioconversion of waste-to-resources (BWR-2021): Valorization of industrial and agro-wastes to fuel, feed, fertilizer, and biobased products.

The mitigation of greenhouse gas (GHG) emission is one of the major focuses of The Glasgow Climate Pact, a global agreement that is believed to accelerate climate action. Following the energy sector, industrial and agro-wastes are the major contributors to global GHG emission. With the rapid growth in population, affluence, and urbanization, the GHG emission from waste sector is likely to be further aggravated if timely measures are not taken to address this burning issue. Thus, a significant research and development efforts are being made in shifting conventional waste treatment approach to resource recovery from waste, incorporating a circular bioeconomy concept. There have been significant advances in technologies such as anaerobic digestion, composting, pyrolysis, algae farming, and microbial fuel cell for recovering resources from organic wastes. This virtual special issue (VSI), "Bioconversion of Waste-to-Resources (BWR-2021)", contains 25 manuscripts covering various aspects of wastes and residual biomass valorization to high value products, including development of new technologies, optimization of current technologies for more efficient utilization of wastes and residues. The key findings of each manuscript are briefly summarized here, which can serve as a valuable resource for researchers in the field of resource recovery from wastes.

Dynamic anaerobic membrane bioreactor coupled with sulfate reduction (SrDMBR) for saline wastewater treatment.

This study investigated organic removal performance, characteristics of the membrane dynamics, membrane fouling and the effects of biological sulfate reduction during high-salinity (1.0%) and high-sulfate (150 mgSO42--S/L) wastewater treatment using a laboratory-scale upflow anaerobic sludge bed reactor integrated with cross-flow dynamic membrane modules. Throughout the operational period, dynamic membrane was formed rapidly (within 5-10 min) following each backwashing cycle (21-16 days), and the permeate turbidity of <5-7 NTU was achieved with relatively high specific organic conversion (70-100 gTOC/kgVSS·d) and specific sulfate reduction (50-70 gSO42--S/kgVSS·d) rates. The sulfide from sulfate reduction can be reused for downstream autotrophic denitrification. 16S rRNA gene amplicon sequencing revealed that the microbial communities enriched in the sludge were different than those accumulated on the dynamic layer. Overall, this study demonstrates that the anaerobic dynamic membrane bioreactor coupled with sulfate reduction (SrDMBR) shows promising applicability in saline wastewater treatment.

Application of machine learning in anaerobic digestion: Perspectives and challenges.

Anaerobic digestion (AD) is widely adopted for remediating diverse organic wastes with simultaneous production of renewable energy and nutrient-rich digestate. AD process, however, suffers from instability, thereby adversely affecting biogas production. There have been significant efforts in developing strategies to control the AD process to maintain process stability and predict AD performance. Among these strategies, machine learning (ML) has gained significant interest in recent years in AD process optimization, prediction of uncertain parameters, detection of perturbations, and real-time monitoring. ML uses inductive inference to generalize correlations between input and output data, subsequently used to make informed decisions in new circumstances. This review aims to critically examine ML as applied to the AD process and provides an in-depth assessment of important algorithms (ANN, ANFIS, SVM, RF, GA, and PSO) and their applications in AD modeling. The review also outlines some challenges and perspectives of ML, and highlights future research directions.

Anaerobic co-digestion of various organic wastes: Kinetic modeling and synergistic impact evaluation.

Anaerobic mono- and co-digestion of coffee pulp (CP), cattle manure (CM), food waste (FW) and dewatered sewage sludge (DSS), were assessed using biochemical methane potential tests. The effects of two different inocula, anaerobically digested cattle manure (ADCM) and anaerobically digested waste activated sludge (ADWAS), and five different co-feedstock ratios for CP:CM and FW:DSS (1:0, 4:1, 2:1, 4:3, and 0:1) on specific methane yields were also evaluated. Mono-digestions of both CP and FW yielded the highest methane yield compared to the co-digestion ratios examined. Furthermore, no synergistic or antagonistic effect was observed for any of the co-digestion ratios tested. Nine different kinetic models (five conventional mono-digestion models and four co-digestion models) were compared and evaluated for both mono- and co-digestion studies. For CP:CM, cone and modified Gompertz with second order equation models were the best-fit for mono- and co-digestion systems, respectively, while for FW:DSS, superimposed model showed the best-fit for all systems.

Chicken manure-based bioponics: Effects of acetic acid supplementation on nitrogen and phosphorus recoveries and microbial communities.

Bioponics has the potential to recover nutrients from organic waste streams, such as chicken manure and digestate with high volatile fatty acid (VFA) contents through crop production. Acetic acid, a dominant VFA, was supplemented weekly (0, 500, 1000, and 1500 mg/L) in a chicken manure-based bioponic system, and its effect on the performance of bioponics (e.g., plant yield and nitrogen and phosphorus availabilities) was examined. Microbial communities were analyzed using 16S rRNA gene sequencing, and the functional gene abundances were predicted using PICRUSt. Although acetic acid negatively affected plant yield, no significant difference (p > 0.05) was noted in the average nitrogen or phosphorus concentration. In terms of nutrient recovery, the bioponic systems still functioned well, although higher concentrations of acetic acid decreased plant yield and altered the bacterial communities in plant roots and chicken manure sediments. These data suggest that an acetic acid concentration of < 500 mg/L or a longer loading interval is recommended for the effective operation of chicken manure and digestate-based bioponics.

Sewage sludge digestion beyond biogas: Electrochemical pretreatment for biochemicals.

Low economic gains from biogas drive research on shifting to volatile fatty acid (VFA) production during anaerobic sludge digestion. pH control and methanogenesis inhibition are widely used strategies for VFA production via anaerobic digestion of sludge. However, these strategies require perpetual dosing of chemicals, increasing cost and operation complexity. Here, we applied electrochemical pretreatment (EPT) (12 V/30 min) for VFA production during anaerobic sludge digestion. The underlying mechanisms of the VFA production induced by EPT were explored systematically through analyses of the changes in the EPT operation parameters, the sludge characteristics, and the microbial community structure and functional enzymes involving in the subsequent sludge digestion. EPT with carbon-based electrodes selectively inhibited methanogenesis by down-regulating heterodisulfide reductase without affecting enzymatic acidogenesis and hydrolysis, resulting in accumulation of VFAs (up to 389±12 mg acetic acid equivalent/L). Propionate and acetate were, respectively enriched to 89 and 75% of the total VFAs after carbon- and graphite- EPT. Titanium-EPT produced lower levels of VFA; instead, biogas yield increased by ∼20%. We anticipate that EPT will advance VFA recovery from diverse organic wastes to meet the global challenge of resource supply and waste management.

Catalytic co-hydrothermal carbonization of food waste digestate and yard waste for energy application and nutrient recovery.

Hydrothermal carbonization (HTC) provides a promising alternative to valorize food waste digestate (FWD) and avoid disposal issues. Although hydrochar derived from FWD alone had a low calorific content (HHV of 13.9 MJ kg-1), catalytic co-HTC of FWD with wet lignocellulosic biomass (e.g., wet yard waste; YW) and 0.5 M HCl exhibited overall superior attributes in terms of energy recovery (22.7 MJ kg-1), stable and comprehensive combustion behaviour, potential nutrient recovery from process water (2-fold higher N retention and 129-fold higher P extraction), and a high C utilization efficiency (only 2.4% C loss). In contrast, co-HTC with citric acid provided âˆ¼3-fold higher autogenous pressure, resulting in a superior energy content of 25.0 MJ kg-1, but the high C loss (∼74%) compromised the overall environmental benefits. The results of this study established a foundation to fully utilize FWD and YW hydrochar for bioenergy application and resource recovery from the process water.

Biogenic sulfide for azo dye decolorization from textile dyeing wastewater.

Azo dye is the most versatile class of dyes used in the textile industry. Although the sulfidogenic process shows superiority in the removal of azo dye, the role of biogenic sulfide produced by sulfate-reducing bacteria (SRB) in the decolorization of azo dye is unclear. This study explored the mechanism of biogenic sulfide for removal of a model azo dye (Direct Red 81 (DR 81)) through biotic and abiotic batch tests with analysis of intermediates of the azo dye degradation. The results showed that biogenic sulfide produced from sulfate reduction directly cleaved two groups of azo bond (-NN-), thereby achieving decolorization. Moreover, the decolorization rate was enhanced by nearly 3-fold (up to 42 ± 1 mg/L-hr; removal efficiency > 99%) by adding an external carbon source or elevating the initial azo dye concentration. This study showed that biogenic sulfide plays an essential role in azo dye decolorization and provides a new avenue for the potential application of biogenic sulfide from the sulfidogenic system for the treatment of azo dye-laden wastewater.