Naematelia aurantialba is a promising basidiomycete chassis for bioproduction, yet its development is hindered by a lack of efficient genetic tools. This study established a robust genetic platform for N. aurantialba NX-20. First, protoplast regeneration was optimized to 46.16% using 20 mg/mL Lywallzyme and 0.6 M maltose. Genome-wide analysis revealed a strong G/C-ending codon preference (GC3s of 67.15%) driven primarily by natural selection, providing a basis for codon optimization. To address the common issues of genetic chimerism and fluorescence heterogeneity in transformants, a 475-bp minimal point centromere (CEN) core element was identified within a genomic GC3 valley. Integration of this CEN element into an expression vector enabled stable, autonomous episomal replication. The utility of this episomal system was validated through the heterologous expression of Vitreoscilla hemoglobin (VHb). Biochemical analysis confirmed VHb activity, which significantly enhanced cellular respiration and energy supply. In stirred-tank bioreactor fermentation, the VHb-expressing engineered strain achieved a Naematelia aurantialba polysaccharide (NAPS) yield of 25.01 g/L, representing a 44.8% increase over the wild-type strain. This versatile and stable genetic platform not only fills a critical gap in N. aurantialba research but also offers a scalable paradigm for the metabolic engineering of non-model basidiomycetes to produce high-value bioproducts.

Sulfamethoxazole (SMX) is frequently found in aquatic environments, causing ecological toxicity and accelerating the spread of antibiotic resistance genes (ARGs). The conventional constructed wetlands (CWs) face challenges in removing antibiotics and recovering energy. Constructed wetland-microbial fuel cells (CW-MFCs) are a combination of constructed wetlands and bio-electrochemical technology, enhancing pollutant removal and bioelectricity production. In this study, an iron-carbon particle-enhanced CW-MFC (FCCW) was constructed for SMX removal. The decontamination performance, electrochemical properties, metagenomic profiles, ARG distribution, and transformation products of SMX were analyzed. The results showed that the FCCW displayed superiority in power generation performance with an average voltage of 286.16 mV, a peak power density of 5.40 mW·m-2, a peak current density of 32.48 mW·m2, and a low internal resistance of 382.10 Ω. The FCCW achieved the highest removal rates of TN (51.66±1.63%), NH4+-N (65.49±1.96%), TP (96.69±2.46%), COD (80.90±2.98%), and SMX (96.49±2.77%). Metagenomic analysis revealed that Proteobacteria and Actinobacteria dominated in the three systems and the iron-carbon particles increased the relative abundance of genes associated with energy metabolism and pollutant transformation. Additionally, the FCCW showed a more diffuse distribution of ARGs and no localized accumulation. The analysis of transformation intermediates showed that the FCCW may rely on a glutathione (GSH)-related conjugation pathway associated with lower accumulation of certain toxic intermediates. Overall, the enhanced performance of the FCCW was attributed to improved redox conditions, more efficient electron transfer, and changes in microbial functional composition. Therefore, the FCCW system offers a promising approach for in-situ electricity generation and stable pollutant treatment performance.

Rice straw is rich in organic matter and essential nutrients, an important resource for improving soil fertility and enzyme activity. However, its high carbon-nitrogen ratio leads to slow decomposition, affecting nutrient transformation efficiency. The small molecular carbohydrates released during microbial enzymolysis not only serve as an energy source but may also act as signaling molecules, participating in the regulation of soil microbial community structure and plant root development. This review discusses the effects of straw return to the field on soil organic carbon (SOC) stability, microbial activity, and plant-soil feedback mechanisms, with particular emphasis on its role in carbon sequestration and CO2 emission mitigation. By enhancing SOC storage and aggregate stability, rice straw returning transforms agricultural waste into a long-term carbon sink, potentially offsetting a portion of anthropogenic CO2 emissions despite short-term increases in soil respiration and greenhouse gas (GHG) fluxes during decomposition. Optimized integration with water-nitrogen management and microbial inoculants can maximize net carbon benefits while sustaining soil fertility. This review highlights the key role of microbial-mediated straw decomposition in agricultural ecosystems and waste valorization. Future research needs to focus on the receptor recognition pathways of specific carbohydrate signals (fiber oligosaccharides, lignin oligosaccharides), the spatiotemporal dynamics of interspecies signaling between microorganisms and plants, the regulatory effects of environmental factors (soil pH, moisture) on signaling pathways, and quantitative modeling of long-term CO2 reduction potentials at regional and global scales.

The addition of biostimulant particles (BPs) derived from olive-mill waste (OMW) has emerged as a promising technique to improve microalgae performance during photosynthetic biogas upgrading (PBU) and has motivated the use of pigment-producing microalgae to improve the cost-competitiveness of the process. However, a clear understanding of the interaction between the particles and microalgae is necessary to design proper materials that maximize both PBU overall performance and pigment production. Thereby, this study aims to unravel the biostimulant effects of OMW-BPs on Arthrospira platensis cultures during batch PBU tests. Three BPs with different Fe contents (0,6 and 15 % dw) were synthetized and added to A. platensis cultures at a concentration of 100 mg L-1. The results showed that the three BPs boosted A. platensis growth rate and phycocyanin (C-PC) content regardless of the Fe content. However, the higher content of humic substances in non-iron containing BPs (hydrochar) resulted in a superior growth performance and in a C-PC content up to 325 mg g-1dw. A. platensis cultures supplemented with hydrochar BPs under light-stress conditions (800 μmol m-2s-1) exhibited a similar C-PC content and a specific growth rate 15 % higher than the non-supplemented cultures. Transcriptomic and SEM-EDS analysis demonstrated that hydrochar BPs interacted extracellularly (by acting as metal chelators and improving nutrient and ions uptake) and intracellularly (by penetrating the cells and improving electron flux and PSII efficiency). Hence, this study demonstrates the potential of hydrochar BPs to improve the photosynthetic efficiency of A. platensis and ultimately the overall PBU process.

To enhance energy and material recovery from complex residual municipal wastes, the post-treated thermally hydrolyzed mechanically sorted organic fraction (PHSW) was evaluated through thermophilic acidogenic fermentation. Using a sequential approach, batch tests first assessed the influence of inoculum source and substrate-to-inoculum ratio on fermentative performance. Stabilized mixed sludge outperformed digested sewage sludge, increasing biohydrogen production by 46%, associated with a higher microbial diversity (Shannon index: 5.489) that favored complementarity between hydrolytic and acidogenic taxa. Despite thermal pretreatment, substrate conversion remained limited, with hydrolysis efficiencies ranging from 8% to 26%. Subsequently, semi-continuous stirred tank reactors were operated at 55 °C under different hydraulic retention times (HRT: 3, 5, and 8 days) to evaluate process performance. The highest specific hydrogen production rate was obtained at HRT 3 days, reaching 19.6 mL H2·g-1 volatile solids·d-1 and a volatile fatty acids productivity of 3.31 g chemical oxygen demand ·L-1·d-1, with butyrate as the predominant metabolite. Microbial analysis revealed a community shift from hydrogen-producers acidogens toward a syntrophic consortium, which was associated with the emergence of hydrogenotrophic methanogenic activity under mildly acidic conditions (pH 5.5-6.5). These findings highlight the potential of PHSW valorization through thermophilic acidogenic fermentation, supporting the transition toward a circular biorefinery model.

Lignocellulosic biomass is an abundant renewable organic carbon resource, but its compact cell-wall structure and lignin shielding limit bioconversion efficiency. Advanced oxidation processes (AOPs) have emerged as promising pretreatment approaches; however, the links between reactive species (RS) chemistry, structural attack patterns, byproduct formation, and downstream bioconversion remain fragmented. This review integrates current evidence within an RS-attack sites-byproducts-downstream conversion framework. The formation conditions and reaction characteristics of inorganic radicals, organic radicals, and non-radical pathways are compared, with emphasis on their preferential reactions with lignin linkages, aromatic moieties, and carbohydrate structures. Comparative analysis indicates that RS selectivity interacts with feedstock properties and operating conditions to shape the formation and distribution of major byproduct profiles, including phenolic compounds, furan derivatives, and small organic acids. The review further evaluates how these byproducts influence enzymatic saccharification, methanogenic anaerobic digestion, and chain elongation fermentation in lignocellulosic bioconversion processes. The dominant mechanisms involve interfacial adsorption, enzyme deactivation, redox burden, and microbial stress, which collectively reshape process kinetics and stability. Practical implementation remains constrained by oxidant cost, catalyst stability and recyclability, and residual oxidant or inhibitor carryover. Therefore, engineering scalability of AOP-based pretreatment should integrate RS regulation with catalyst recovery, inhibitor mitigation, and downstream tolerance rather than focusing only on delignification or sugar release. Overall, AOP-based pretreatment should shift from maximizing oxidative intensity or delignification toward controlling RS pathways and byproduct carryover according to downstream tolerance.

Anaerobic methane oxidation coupled to extracellular electron transfer (EET) has shown promising application in bioenergy recovery, yet whether this process can simultaneously drive biological nitrogen fixation (BNF) remains unexplored. Here, we demonstrated that methane-fed microbial fuel cells (M-MFCs) under nitrogen-limited conditions achieve concurrent electrogenesis and BNF via a syntrophic division of labor mechanism. The M-MFC system generated a peak current of 1.17 ± 0.18 mA with a Coulombic efficiency of 78.1%, and the ammonium in the medium accumulated to 0.22 ± 0.01 mg/L over a 30-day operation. 15N2 stable isotope probing (SIP) confirmed direct methane-driven BNF, with the 15N abundances in biomass and aqueous ammonium reaching 13.55% and 7.15%, respectively. Raman spectroscopy revealed 15N incorporation into cytochrome c, while electrochemical analyses and inhibitor assays identified cytochrome c-mediated electron transfer as the dominant pathway for electrogenesis. DNA-SIP combined with high-throughput sequencing further revealed the distinct metabolic specialization of the consortium: Methanobacterium mediated anaerobic methane oxidation, Geobacter and Desulfovibrio specialized in EET, and Sulfurivermis, Desulfobulbus and Azoarcus acted as the core diazotrophs. This work establishes a new paradigm for multifunctional bioelectrochemical systems, which enables energy recovery from methane while simultaneously co-producing bioavailable nitrogen, thereby achieving a trinity of benefits for sustainable environmental biotechnology.

Floating treatment wetlands (FTWs) are increasingly used as nature-based water infrastructure, but their performance is often limited by oxygen transfer, hydrodynamic variability, and unstable microbiomes. This review synthesizes bioaugmentation evidence from FTWs and related wetlands for nutrients, metals, hydrocarbons, pharmaceuticals, antibiotics, microplastics, and emerging contaminants like per- and polyfluoroalkyl substances (PFAS), noting that destructive transformation is rarely demonstrated for some compounds. Separating contaminant removal from confirmed degradation, we review mechanisms, robustness, and scale. This review compares liquid consortia with carrier-immobilized or encapsulated inoculum and extract-based practical design criteria for plant-microbial partnerships, rhizosphere engineering, and biofilm development and persistence under shear, seasonality, and pulse loading. Targeted aeration, supportive media, and electrochemical augmentation, such as wetland microbial fuel cells and electrochemical oxidation (electro-oxidation) and coagulation-based processes (e.g., conventional chemical coagulation or electrocoagulation), can boost performance and stabilize new functions. Mechanistic sections map attenuation pathways to catalytic modules, including oxygenases, reductases, and hydrolases, as well as to biosorption and biomineralization, and identify dominant failure modes that hinder translation, including washout, community reversion, inhibitory intermediates, and uncertain long-term ecological effects. Building on this evidence base, we outline a testable translational roadmap toward more programmable FTWs, emphasizing near-term decision support rather than fully automated control, and specifying validation needs for multi-omics-guided strain selection, division-of-labor consortia, safety-by-design containment, and model-informed monitoring and operation under climate variability. Finally, we propose reporting and governance metrics, including effect sizes versus controls, persistence, ecological risk monitoring, and life-cycle trade-offs, to support responsible field deployment and water reuse.

Anoxic zone played essential roles in nutrient removal in biological wastewater treatment processes. This study compared the impacts of pre-anoxic (PreA) and post-anoxic (PostA) configurations on sludge characteristics, nutrient removal performance, and microbial succession. Results showed that shifting from PreA to PostA deteriorated sludge settleability, with sludge volume index (SVI30) increasing from 71 to 192 mL·g[-1]. Meanwhile, sludge structures became looser and more hydrophilic, accompanied by a decrease in the protein (PN) to polysaccharides (PS) ratio of extracellular polymeric substances (EPS) from 0.78 to 0.62. Denitrification batch tests revealed that PostA promoted soluble organic matter release, with the release rates of 1.17 and 1.54 mg COD·(g·h)[-1] under PreA and PostA, respectively. In addition, PostA denitrification relied more on the EPS-derived soluble products as electron donors, including tyrosine- and tryptophan-like substances, which contributed to sludge structural loosening. Furthermore, the sludge bulking under PostA was not attributed to filamentous bacteria proliferation, as their relative abundance remained unchanged (from 3.00 % to 3.03 %), but by the loose and hydrophilic sludge. Overall, the layout of anoxic zone regulated the sludge structure by altering the distribution of carbon sources and electron acceptors, reshaping microbial metabolic functions rather than community composition.

Biochar (BC), particularly micro/nanoscale biochar (MNBC), exhibits great potential for remediating heavy metal-contaminated soils. However, the mechanisms underlying their effects on microbial life-history strategies and plant physiological responses remain poorly understood. Here, a 50-day pot experiment was conducted to evaluate the effects of BC/MNBC applied at 1%, 2% and 4% (w/w) on soybean growth and rhizosphere bacterial life-history strategies under cadmium (Cd) stress. Compared to Cd-alone, BC decreased malondialdehyde and superoxide anion contents by 33.3%-40.7% and 11.2%-23.0%, respectively. The corresponding reductions under MNBC treatment were 41.1%-52.7% and 18.6%-31.2%, respectively, indicating a greater alleviation of Cd-induced oxidative damage. Principal component analysis further indicated a shift in soybean physiological status from Cd stress to homeostasis, coinciding with biochar application. Notably, 4% MNBC significantly decreased shoot and root Cd by 70.7% and 65.2%, respectively, outperforming BC. BC/MNBC amendments were associated with higher relative abundances of bacterial r-strategists and lower abundances of K-strategists under Cd stress. These findings highlight the potential of MNBC to enhance crop resilience and decrease Cd accumulation, providing a microbial ecological perspective on the restoration of Cd-contaminated agroecosystems.

Lipids in food waste (FW) are promising biofuel precursors, but their recovery from slurry-type FW is limited by entrapment within carbohydrate-protein-solid matrices and emulsion-like structures. This study developed a short-term pre-fermentation pretreatment strengthened by a mixed acidifying inoculum to improve intrinsic lipid recovery before thermal centrifugation. In laboratory tests, inoculated pre-fermentation increased lipid yield by 36.3% relative to the initial slurry and recovered 97.8% of the Soxhlet-determined total solvent-extractable lipid content after 24 h. Particle refinement, viscosity reduction, total organic carbon release, lactate accumulation, pH decline, and Lactobacillus dominance indicated that hydrolysis-assisted acidification disintegrated the slurry matrix and improved lipid accessibility. Recovered lipids also showed moderate compositional shifts, including increased saturation, slight C8-C16 enrichment, and minor branched-chain fatty acids, suggesting limited microbial effects on the recovered lipid profile. Full-scale validation at a 600 tonnes/day FW treatment facility achieved a 13.9% lipid-yield increase under an 8 h retention time and lower inoculum dosage, demonstrating practical applicability in an operating wet-thermal recovery line. An incremental cost-benefit estimate indicated a positive annual net benefit under conservative assumptions. Overall, inoculation-strengthened pre-fermentation provides a low-retrofit and process-compatible strategy for enhancing FW lipid recovery within existing treatment chains.

Microalgae are regarded as a promising platform for carbon capture and storage (CCS). A key target for enhancing CCS in microalgae is the carbon-concentrating mechanism (CCM), which is heavily reliant on carbonic anhydrase (CA) activity. Overexpressing CA in microalgae may offer an effective approach to capture excess CO2. A directed-evolution-generated variant of Desulfovibrio vulgarisCA (DvCA) with ultrastable properties and efficient carbon capture capability in vitro was previously reported. Here, the DvCA was heterologously expressed in the green alga Chlamydomonas reinhardtii (Chlamydomonas). The engineered Chlamydomonas strain, expressing the DvCA in the chloroplast, exhibited significantly higher inorganic carbon uptake, total carbon conversion efficiency and more biomass than parental line (PL). Under photoautotrophic conditions (ambient CO2), photosynthetic rate and photoprotection were also enhanced in DvCA expressing transformants. Moreover, the DvCA transformants showed a significant increase in carbon storage, with starch and triacylglycerol (TAG) content rising by 25 % and 53 %. In addition, the expression levels of major genes directly involved in photosynthesis, TAG and starch biosynthesis were upregulated in DvCA transformants comparted to PL. These results demonstrated that heterologous expression of highly stable CA is an effective strategy for optimizing microalgal CCMs, offering considerable promise for enhancing CCS using microalgae.

The operational stability of continuous-flow partial denitrification-anammox (PD/A) systems is frequently constrained by insufficient nitrite supply and temperature sensitivity of anammox bacteria, particularly under low-temperature stress. In such conditions, filamentous bacteria often proliferate excessively, and their overgrowth has long been associated with reactor instability and performance deterioration. Here, we demonstrate stable nitrogen removal in a filamentous-dominated continuous-flow PD/A reactor at an average temperature of 16.7 °C through operational optimizations and microbial responses. The reactor achieved 89.8 % total nitrogen removal, with ammonium and nitrate removal efficiencies of 97.2 % and 91.5 %, respectively, with anammox contributing up to 98.3 % of nitrogen removal. Metagenomic analyses revealed that the filamentous genus Sphaerotilus dominated the microbial community (29.3-41.5 %) but sustained the genomic potential for nitrite availability to support anammox. Genome-centric reconstruction confirmed that a Sphaerotilus-affiliated MAG5 possessed adaptive features under low temperature. Additional heterotrophs, including Leptothrix, Rubrivivax, and Thauera, harbored genomic potential for auxiliary nitrate-to-nitrite conversion. Crucially, the synergy between this genomic potential for nitrite provision and engineered biomass retention (specifically mesh filtration and regular sludge return) facilitated the enrichment of Ca. Brocadia, increasing its relative abundance from 2.4 % to 5.4 %. Concurrently, Ca. Brocadia reinforced low-temperature adaptability by expanding the genetic potential of energy-generating carbon metabolic pathways and increasing its contribution to the cold shock protein gene cspA from 6.6 % to 21.0 %. Collectively, this study reveals that integrating strategic biomass retention with microbial responses provides a viable pathway to sustain stable nitrogen removal in filamentous-dominated continuous-flow PD/A systems.

Single-atom catalysts (SACs) have attracted considerable attention owing to their high catalytic activity and atom utilization. Nevertheless, their application to antibiotic removal and associated biosafety remains insufficiently explored. In this work, a single-atom Co catalyst supported on N-doped biochar (SA-Co-NC) was synthesized by impregnation-pyrolysis to activate peroxymonosulfate (PMS) for the degradation of tetracycline (TC). The porous biochar stabilized atomically dispersed Co-N3 sites. The experimental results demonstrate that the SA-Co-NC/PMS system removed 96.6% of TC within 18 min and achieved 75.7% mineralization. X-ray absorption spectroscopy and density functional theory calculations indicate that coordinatively unsaturated Co-N3 sites regulate the local electronic structure, strengthen TC and PMS adsorption, and promote PMS activation, with [1]O2 dominating the nonradical pathway and Co(IV)=O possibly contributing. Phytotoxicity assessments demonstrate that the treated solution is less phytotoxic than untreated TC solution. Animal toxicity tests reveal that a 24-h oral exposure to SA-Co-NC caused no overt histopathological damage in the mouse liver, kidney, or thyroid and produced only limited changes in the measured oxidative-stress and inflammatory markers. These results demonstrate the catalytic effectiveness and preliminary biosafety of SA-Co-NC for antibiotic-contaminated water treatment.

Black soldier fly larvae (BSFL) are rich in crude protein (CP) and crude fat (EE), offering substantial potential for animal feed and biodiesel production. However, their nutritional composition is dynamically influenced by multiple factors. Traditional chemical detection methods are inefficient to meet the practical demands of modern farming. Therefore, this study developed a machine learning-based prediction method for the major nutritional components of BSFL. By collecting larval growth data under different feeding substrates and environmental conditions, six regression models were evaluated. Results demonstrated that Light Gradient Boosting Machine (LightGBM), achieved optimal performance in predicting CP content (determination coefficient (R[2]) = 0.711, mean absolute error (MAE) = 3.515, root mean square error (RMSE) = 4.913), while K-Nearest Neighbors outperformed the other five models in predicting EE content (R[2] = 0.702, MAE = 4.508, RMSE = 5.975). Validation on five practical substrates produced mean absolute errors of 3.14 percentage points for CP and 3.42 percentage points for EE, indicating promising predictive potential under tested conditions. SHapley Additive exPlanations analysis identified larval instar stage and initial weight as the most important predictive factors for CP, and feeding substrate as the most influential factor for EE prediction. This study provided a data-driven tool to rapidly evaluate BSFL nutritional components, supporting optimization of farming strategies, although validation under broader conditions is still required.