MlrA, an Essential Enzyme for Microcystins and Nodularin on First Step Biodegradation in Microcystin-Degrading Bacteria.

Microcystin-degrading bacteria first degrade microcystins by microcystinase A (MlrA) to cleave the cyclic structure of microcystins at the Adda-Arg site of microcystin-LR, microcystin-RR, and microcystin-YR, but the cleavage of the other microcystins was not clear. In our study, the microcystin-degrading bacterium Sphingopyxis sp. C-1 as wild type and that of mlrA-disrupting mutant, Sphingopyxis sp. CMS01 were used for microcystins biodegradation. The results showed MlrA degraded microcystin-LA, microcystin-LW, microcystin-LY, microcystin-LF, and nodularin. MlrA could cleave the Adda-L-amino acid site.

Photosynthetic oxygen-supported algal-bacterial aerobic granular sludge can facilitate carbon, nitrogen and phosphorus removal from wastewater: Focus on light intensity selection.

Photosynthetic O2 is a promising alternative for mechanical aeration, the major energy-intensive unit in wastewater treatment plants. This study aimed to investigate the effects of light intensity varied from 190 to 1400 µmol·s-1·m-2 on photosynthetic O2-supported algal-bacterial aerobic granular sludge (AGS) system. Results indicate photosynthetic O2 can implement aerobic phosphorus (P) uptake and ammonia oxidation under the test illumination range even at dissolved oxygen concentration < 0.5 mg/L. An obvious O2 accumulation occurred after 60-90% nutrients being removed under 330-1400 µmol·s-1·m-2, and highly efficient ammonia removal, P uptake, and dissolved inorganic carbon removal were achieved under 670-1400 µmol·s-1·m-2. On the other hand, photosynthesis as O2 supplier showed little effect on major ions except for K+. This study provides a better understanding of the roles of light intensity on photosynthetic O2-supported algal-bacterial AGS system, targeting a sustainable wastewater industry.

Toxicity of ciprofloxacin and ofloxacin to Moina macrocopa and investigation of p-value adjustments for (eco)toxicological studies.

Ciprofloxacin (CFX) and ofloxacin (OFX) are commonly found as residual contaminants in aquatic environments, posing potential risks to various species. To ensure the safety of aquatic wildlife, it is essential to determine the toxicity of these antibiotics and establish appropriate concentration limits. Additionally, in (eco)toxicological studies, addressing the issue of multiple hypothesis testing through p-value adjustments is crucial for robust decision-making. In this study, we assessed the no observed adverse effect concentration (NOAEC) of CFX and OFX on Moina macrocopa across a concentration range of 0-400 µg L-1. Furthermore, we investigated multiple p-value adjustments to determine the NOAECs. Our analysis yielded consistent results across seven different p-value adjustments, indicating NOAECs of 100 µg CFX L-1 for age at first reproduction and 200 µg CFX L-1 for fertility. For OFX treatment, a NOAEC of 400 µg L-1 was observed for both biomarkers. However, further investigation is required to establish the NOAEC of OFX at higher concentrations with greater certainty. Our findings demonstrate that CFX exhibits higher toxicity compared to OFX, consistent with previous research. Moreover, this study highlights the differential performance of p-value adjustment methods in terms of maintaining statistical power while controlling the multiplicity problem, and their practical applicability. The study emphasizes the low NOAECs for these antibiotics in the zooplanktonic group, highlighting their significant risks to ecological and environmental safety. Additionally, our investigation of p-value adjustment approaches contributes to a deeper understanding of their performance characteristics, enabling (eco)toxicologists to select appropriate methods based on their specific needs and priorities.

Highly efficient carbon assimilation and nitrogen/phosphorus removal facilitated by photosynthetic O2 from algal-bacterial aerobic granular sludge under controlled DO/pH operation.

Reducing CO2 emission and energy consumption is crucial for the sustainable management of wastewater treatment plants (WWTPs). In this study, an algal-bacterial aerobic granular sludge (AGS) system was developed for efficient carbon (C) assimilation and nitrogen (N)/phosphorus (P) removal without the need for mechanical aeration. The photosynthetic O2 production by phototrophic organisms maintained the dissolved oxygen (DO) level at 3-4 mg/L in the bulk liquid, and an LED light control system reduced 10-30% of light energy consumption. Results showed that the biomass assimilated 52% of input dissolved total carbon (DTC), and the produced O2 simultaneously facilitated aerobic nitrification and P uptake with the coexisting phototrophs serving as a C fixer and O2 supplier. This resulted in a stably high total N removal of 81 ± 7% and an N assimilation rate of 7.55 mg/(g-MLVSS∙d) with enhanced microbial assimilation and simultaneous nitrification/denitrification. Good P removal of 92-98% was maintained during the test period at a molar ∆P/∆C ratio of 0.36 ± 0.03 and high P release and uptake rates of 10.84 ± 0.41 and 7.18 ± 0.24 mg/(g- MLVSS∙h), respectively. Photosynthetic O2 was more advantageous for N and P removal than mechanical aeration. This proposed system can contribute to a better design and sustainable operation of WWTPs using algal-bacterial AGS.

Enhanced alginate-like exopolymers recovery from algal-bacterial aerobic granular sludge: Optimal cultivation condition and contribution of bacteria and microalgae during the transport/storage period.

Extracting alginate-like exopolymers (ALE) is a promising approach for valuable resources recovery from excess algal-bacterial aerobic granular sludge (AGS) to achieve circular bioeconomy and environmental sustainability in wastewater treatment plants (WWTPs). In this study, six batch cultivation tests were conducted to investigate the optimal cultivation duration or transport/storage period, light intensity, and temperature for algal-bacterial AGS after sampling and before further processing or ALE extraction. At a light intensity of 5 klux, the highest ALE content (36.33 mg/g-VSS) was detected at a low temperature of 10 °C, which increased by 300% from its original level after 6-h cultivation. Results from levofloxacin (LVX) exposure and dark condition imply that microalgae contributed more to ALE synthesis in the algal-bacterial granules. This work not only provides a better understanding of the mechanisms involved in ALE biosynthesis but also offers useful guidance for maintaining or improving ALE recovery after algal-bacterial biomass sampling.

Exosomes in ascites from patients with human pancreatic cancer enhance remote metastasis partially through endothelial-mesenchymal transition.

BACKGROUND: Despite advances in multidisciplinary treatment, the prognosis of pancreatic cancer remains poor. Since distant metastasis defines prognosis, elucidation of the mechanism of metastasis is important for improving survival. Exosomes are extracellular secretory vesicles and are responsible for intercellular communication. In this study, we investigated whether exosomes secreted by human pancreatic cancer cells are involved in promoting distant metastasis of cancer and the mechanism that underlies the promotion of metastasis. METHODS: Exosomes were isolated from ascites of a patient with pancreatic cancer and a patient with liver cirrhosis as a control. Three days after the administration of exosomes to nude mice, GFP-labeled human pancreatic cancer cells were injected via the spleen or tail vein, and then the liver and lungs were histologically analyzed. To elucidate the mechanism, vascular permeability was estimated using FITC-dextran in place of pancreatic cancer cells in vivo and human umbilical vascular endothelial cells (HUVECs) were used to analyze vascular permeability and the induction of endothelial-mesenchymal transition (EndMT) in vitro. RESULTS: Distant metastasis and vascular permeability were significantly enhanced in mice treated with exosomes from pancreatic cancer patients in comparison to exosomes from a control patient in vivo. In addition, exosomes from pancreatic cancer patients significantly enhanced vascular permeability and the induction of EndMT in HUVECs in vitro. CONCLUSION: Exosomes derived from pancreatic cancer cells form a pre-metastatic niche and promote the extravasation and colonization of pancreatic cancer cells to remote organs, partially through endothelial-mesenchymal transition.

Enhanced fixation of dissolved inorganic carbon by algal-bacterial aerobic granular sludge during treatment of low-organic-content wastewater.

The microalgae-based wastewater treatment technologies are believed to contribute to carbon neutrality. This study investigated the inorganic carbon fixation performance in the algal-bacterial aerobic granular sludge (A-BAGS) process under cultivation at different concentrations of organic carbon (OC) and inorganic carbon (IC). The results indicated that A-BAGS in treating wastewater containing organics of 77 mg-C/L contributed little to the fixation of inorganic carbon, while the highest inorganic carbon removal efficiency of 50 % was achieved at the influent IC of 100 mg/L and OC of 7 mg/L. This high IC condition contributed to enhanced biomass growth rate and enhanced extracellular polymeric substances, while it did not affect the granular stability and nitrification efficiency. The microbial diversity was also largely enhanced. The results demonstrated the great potential of A-BAGS for simultaneous resource recovery in wastewater and inorganic carbon fixation, while operation conditions need to be further optimized.

Algal-bacterial aerobic granular sludge for real municipal wastewater treatment: Performance, microbial community change and feasibility of lipid recovery.

Despite various research works on algal-bacterial aerobic granular sludge for wastewater treatment and resource recovery processes, limited information is available on its application in real wastewater treatment in terms of performance, microbial community variation and resource recovery. This study investigated the performance of algal-bacterial aerobic granular sludge on real low-strength wastewater treatment in addition to the characterization of microbial community and fatty acid compositions for biodiesel production. The results demonstrated 71% COD, 77% NH4+-N and 31% phosphate removal efficiencies, respectively. In addition, all the water parameters successfully met the effluent standard A, imposed by the Department of Environment (DOE) Malaysia. Core microbiome analyses revealed important microbial groups (i.e., Haliangium ochraceum, Burkholderiales and Chitinophagaceae) in bacterial community. Meanwhile the photosynthetic microorganisms, such as Oxyphotobacteria and Trebouxiophyceae dominated the algal-bacterial aerobic granular sludge, suggesting their important roles in granulation and wastewater treatment. Up to 12.51 mg/gSS lipid content was recovered from the granules. In addition, fatty acids composition showed high percetages of C16:0 and C18:0, demonstrating high feasibility to be used for biodiesel production application indicated by the cetane number, iodine value and oxidation stability properties.

Revealing calcium ion behavior during anaerobic phosphorus release process in aerobic granular sludge system.

Calcium ions (Ca2+) are important for biological phosphorus (P) removal from wastewater, but its behavior has not been well documented during the anaerobic P release process. This study is aimed to explore the mechanisms of Ca2+ release in bacterial aerobic granular sludge (AGS) system. During the non-aeration (anaerobic) phase, nearly 40 % increase in Ca2+ concentration was detected at the bottom of AGS reactor where decrease in pH and increase in Mg2+ concentration occurred. The pH decrease due to anaerobic P release caused CaCO3 dissolution inside the granules, leading to Ca2+ release. In addition, the increased Mg2+ ions from hydrolysis of polyphosphates were detected to reversibly exchange with Ca2+ in granules at a molar ΔCa/ΔMg ratio of 0.51-0.65. Results from this work revealed that dissolution of CaCO3 and ions exchange between Ca2+ and Mg2+ were the two major contributors to Ca2+ release during anaerobic P release process.

Enhanced economic benefit of recycling Fe3O4 for promotion of volatile fatty acids production in anaerobic fermentation of food waste.

Fe3O4 addition in anaerobic fermentation of food waste (FW) is promising for enhancing volatile fatty acids (VFAs) production. However, the large amount of Fe3O4 in the digestate fertilizer leads to the waste of resources and possible toxicity to organisms. Thus, this study investigated the feasibility of Fe3O4 recycling for VFAs enhancement in anaerobic fermentation of FW and performed the cost-benefit evaluation of this process. Results revealed that Fe3O4 could be successfully recycled twice with recovery rates of 71.5% and 65.5%, respectively. X-ray diffraction analysis revealed a slight change to the Fe2O3-like structure after 2-time recycling. The VFAs yields were enhanced by 17.2% and 17.0% in Cycles 1 and 2 owing to the enhanced activities of hydrolytic and acid-forming enzymes. The net income of the Fe3O4 recycling process was about 13-fold higher than that of the conventional treatment process, suggesting a promising and economically feasible strategy for enhancing VFAs production.

Pb(II) bioremediation using fresh algal-bacterial aerobic granular sludge and its underlying mechanisms highlighting the role of extracellular polymeric substances.

Lead (Pb) discharged from rural industries poses a significant threat to the environment and human health. Algal-bacterial aerobic granular sludge (A-B AGS) is a promising alternative for sewage treatment with high efficiency and good settleability. In this study, Pb(II) biosorption using fresh A-B AGS was investigated for the first time. The important role of extracellular polymeric substances (EPS) was revealed with the involved mechanisms being clarified. The desorbents for Pb recovery from Pb-loaded A-B AGS were also screened. Results showed that A-B AGS has an excellent maximum Pb adsorption capacity of 72.4 mg·g-1 at pH 6.0. EPS plays an important role in keeping microbial activity, Pb bonding, and providing metal ions (Ca, Na and Mg) for Pb ion exchanges. Electrostatic interaction, ion exchange, and bonding to functional groups may occur orderly in the Pb biosorption process and the formation of pyromorphite (Pb5(PO4)3Cl) contributes to Pb biosorption. About 66 % of the adsorbed Pb was accumulated in the A-B AGS microbial cells. Na2EDTA (0.05 M) can recover 60.3 % of the loaded Pb with the highest microbial activity of granules being remained. All the findings will provide the theoretical basis for the large-scale application of A-B AGS to bioremediate Pb(II)-containing wastewater.

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.

Fe3O4 enhanced efficiency of volatile fatty acids production in anaerobic fermentation of food waste at high loading.

High treatment capacity for food waste (FW) is required due to the huge amount generated worldwide. Conversion of FW to volatile fatty acids (VFAs) via anaerobic fermentation is a promising technology; however, inhibition of VFAs production could easily occur at high loadings. In this study, Fe3O4 was used to enhance VFAs production in anaerobic fermentation of FW at high loading, and the mechanisms involved were revealed at microbial levels. Results showed that Fe3O4 significantly enhanced VFAs yield and VFAs productivity of microbes by 160% at high loading (substrate to inoculum (S/I) ratio of 3). The enhancement effect of Fe3O4 was mainly due to the accelerated hydrolysis of particulate/soluble organics, the enriched hydrolytic and acidogenic bacteria, and the reduced relative abundance of Lactobacillus. This study provides a new approach for the high-efficient treatment of FW at high loadings, while the performance and economic benefit should be further studied for practical application.

Characteristics of Antibiotic Resistance and Tolerance of Environmentally Endemic Pseudomonas aeruginosa.

Antibiotic-resistant bacteria remain a serious public health threat. In order to determine the percentage of antibiotic-resistant and -tolerant Pseudomonas aeruginosa cells present and to provide a more detailed infection risk of bacteria present in the environment, an isolation method using a combination of 41 °C culture and specific primers was established to evaluate P. aeruginosa in the environment. The 50 strains were randomly selected among 110 isolated from the river. The results of antibiotic susceptibility evaluation showed that only 4% of environmental strains were classified as antibiotic-resistant, while 35.7% of clinical strains isolated in the same area were antibiotic-resistant, indicating a clear difference between environmental and clinical strains. However, the percentage of antibiotic-tolerance, an indicator of potential resistance risk for strains that have not become resistant, was 78.8% for clinical strains and 90% for environmental strains, suggesting that P. aeruginosa, a known cause of nosocomial infections, has a high rate of antibiotic-tolerance even in environmentally derived strains. It suggested that the rate of antibiotic-tolerance is not elicited by the presence or absence of antimicrobial exposure. The combination of established isolation and risk analysis methods presented in this study should provide accurate and efficient information on the risk level of P. aeruginosa in various regions and samples.

Energy saving and rapid establishment of granular microalgae system from tiny microalgae cells: Effect of decrease in upflow air velocity under intermittent aeration condition.

The novel type of microalgae granules (MGs) derived from tiny microalgae cells has received extensive attention due to its great potential for nutrient remediation and resource recovery in wastewater treatment whereas the long start-up time with increased labor expenses remains a bottleneck. In this study, an operation strategy at reduced upflow air velocity (UAV = 0.49 cm/s in RA) under intermittent aeration mode was proposed and compared with RB at a higher UAV (0.98 cm/s) in terms of MGs formation, maintenance, and energy consumption. Although the formation of MGs in RA was delayed for 12 days compared to RB, 40.78 % increase in chlorophyll-a content was detected in MGs in RA along with more cost-effective carbon, nitrogen, and phosphorus removals due to efficient microalgae assimilation and energy reduction. Results from this study provide new insight into minimizing energy input for rapid establishment and stable operation of MG systems towards environmentally sustainable wastewater management.

Microcystin-LR incorporated into colonic cells through probenecid-sensitive transporters leads to upregulated MCP-1 expression induced by JNK activation.

Harmful algae that inhabit eutrophic lakes produce cyanotoxic microcystins. Therefore, the relationship between chronic exposure to microcystins via drinking water and organ disorders has been investigated. The present study aimed to determine whether representative microcystin-LR is involved in increased monocyte chemoattractant protein-1 (MCP-1) expression in rat colonic mucosa and enterocyte-like differentiated Caco-2 cells. The mRNA expression of MCP-1 was increased in the colons of rats administered with microcystin-LR, compared with controls. Furthermore, mRNA levels of MCP-1 expression significantly and positively correlated with those of Adhesion G Protein-Coupled Receptor E1 (ADGRE1; EMR1; F4/80), an indicator of macrophage infiltration, suggesting that increased MCP-1 expression induced by microcystin-LR promotes macrophage infiltration into the colon. Microcystin-LR increased MCP-1 expression in enterocyte-like differentiated Caco-2 cells, by activating c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase (ERK) or p38. The findings of transporter inhibitors indicated that microcystin-LR is incorporated into cells via ATP Binding Cassette (ABC) or solute carrier (SLC) transporters other than the organic anion transporting polypeptides (OATPs)1B1, 1B3, 2B1, and 1A2, which this leads to increased MCP-1 expression in the colon through activating JNK. Thus, increased MCP-1 expression induced by microcystin-LR might be a trigger for initiating tumorigenesis with inflammation in the colon because increased MCP-1 expression induces inflammation associated with macrophage infiltration into the colon, and chronic inflammation is associated with the initiation of tumorigenesis.

Insight into enhanced acetic acid production from food waste in anaerobic hydrolysis/acidification with Fe3O4 supplementation.

Fe3O4 supplementation has been reported as a high-efficient approach to enhance biogas production in anaerobic digestion (AD). Volatile fatty acids (VFAs), especially acetic acid (HAc), are considered as important products in acidification process of AD. However, the possible mechanisms involved in promotion effect of Fe3O4 on HAc production in hydrolysis and acidification processes of AD have not been comprehensively studied. This study first investigated the promotion effect of Fe3O4 on hydrolysis, acidogenesis and acetogenesis stages of AD and proposed the underlying mechanisms, using food waste (FW) as the feedstock, which is considered as the most suitable substrate for VFAs production. Results indicated that the HAc production (77.38 g-C/kg-VS) was enhanced by 79 % in AD of FW with addition of 10 g/L Fe3O4. The duration to reach the maximum HAc production was also shortened from 14 days to 10 days. The AD tests using model substrates revealed that Fe3O4 enhanced hydrolysis, acidogenesis, and degradation of propionic acid, thus resulting in enhanced HAc production. The enhanced activities of hydrolytic and acid-forming enzymes, and electron transport system (ETS) with Fe3O4 addition further demonstrated its function as an electron acceptor to stimulate electron transfer and accelerate microbial metabolisms in AD, which contributed to the enhanced HAc production from FW.

A comparative study on simultaneous recovery of phosphorus and alginate-like exopolymers from bacterial and algal-bacterial aerobic granular sludges: Effects of organic loading rate.

The effects of organic loading rate (OLR) on simultaneous phosphorus (P) and alginate-like exopolymers (ALE) recovery from bacterial aerobic granular sludge (AGS) and algal-bacterial AGS were examined and compared during 70 days' operation. With the increase of OLR (0.6-1.2 g COD/(L·day)), both AGS showed good settleability and granular strength with P bioavailability > 92% (Stage III). The moderate increase in OLR had a positive influence on simultaneous recovery of P and ALE. On day 60, the contents of ALE and guluronic acid/guluronic acid (GG) blocks reached the highest in algal-bacterial AGS, about 13.37 and 2.13 mg/g-volatile suspended solids (VSS), respectively. Meanwhile, about daily 0.55 kg of P is estimated to be recovered from the wastewater treatment plant with a treatment capacity of 10,000 m3/day. P mass balance analysis during ALE extraction from both AGS was conducive to further evaluation of P removal pathway and its application potentials.

Insight into aerobic phosphorus removal from wastewater in algal-bacterial aerobic granular sludge system.

This study aimed to figure out the main contributors to aerobic phosphorus (P) removal in the algal-bacterial aerobic granular sludge (AGS)-based wastewater treatment system. Kinetics study showed that aerobic P removal was controlled by macropore (contributing to 64-75% P removal) and micropore diffusion, and the different light intensity (0, 4.0, 12.3, and 24.4 klux) didn't exert significant (p > 0.05) influence on P removal. On the other hand, the increasing light intensity did promote microalgae metabolism, leading to the elevated wastewater pH (8.0-9.8). The resultant pH increase had a strongly negative relationship (R2 = 0.9723) with P uptake by polyphosphate-accumulating organisms, while promoted chemical Ca-P precipitation at a molar Ca/P ratio of 1.05. Results from this work could provide an in-depth understanding of microalgae-bacteria symbiotic interaction, which is helpful to better design and operate the algal-bacterial AGS systems.

Pancreatic cancer cell-derived exosomes induce epithelial-mesenchymal transition in human pancreatic cancer cells themselves partially via transforming growth factor ß1.

Distant metastasis is a dismal prognostic factor of pancreatic cancer. Metastasis is established in several steps, but the mechanism underlying the very early stages remains unclear. Epithelial-mesenchymal transition (EMT) is involved in these stages. Although signaling molecules have been reported to induce EMT, the mechanism underlying their origin is unclear. In this study, we hypothesized that pancreatic cancer cell-derived exosomes induce EMT in cancer cells themselves, a notion we entertained because we found EMT in in vitro three-dimensional colonies of cancer cells, with vimentin-positive cells observed in some of the budding pancreatic cancer cells and in single cells outside the colony as well. First, we clarified that pancreatic cancer cell-derived exosomes induce EMT in cancer cells themselves. Next, we examined the involvement of transforming growth factor-ß1 (TGF-ß1), and TGF-ß1 knock-down in pancreatic cancer cells with TGF-ß1 siRNA significantly suppressed TGF-ß1 gene expression in cancer cells, and exosomal TGF-ß1 was significantly reduced in the secretory exosomes. Exosomes from TGF-ß1 knock-down cells suppressed EMT induction in cancer cells themselves and TGF-ß1 protein expression in target cells. Taken together, these findings suggest that TGF-ß1 is involved in EMT induction via exosomes, results that may support the production of effective metastasis inhibitors.