Magnetically stimulated azo dye biodegradation by a newly isolated osmo-tolerant Candida tropicalis A1 and transcriptomic responses.

A recently isolated osmo-tolerant yeast Candida tropicalis A1, which could decolorize various azo dyes under high-salinity conditions, was systematically characterized in the present study. Stimulating dye-decolorization effectiveness and osmo-tolerance of the yeast by static magnetic field (SMF) was investigated and transcriptomic responses of the yeast to SMF was analyzed to propose possible mechanisms. The results demonstrated that the yeast A1 effectively decolorized (≥ 97.50% within 12 h) and detoxified (from high toxicity to low toxicity within 24 h) 70 mg/L Acid Red B (ARB) under the optimized conditions through a series of steps including naphthalene-amidine bond cleavage, reductive or oxidative deamination/desulfurization, open-loop of hydroxy-substituted naphthalene or benzene and TCA cycle. Moreover, dye decolorization performance and osmo-tolerance of the yeast A1 were further improved by 24.6 mT SMF. Genes encoding high-affinity hexose/glucose transporter proteins and NADH-ubiquinone oxidoreductase were up-regulated by 24.6 mT SMF, which might be responsible for the increase of dye decolorization. Significant up-regulation of glycerol-3-phosphate dehydrogenase and cell wall protein RHD3 suggested that osmo-tolerance was enhanced by 24.6 mT SMF through promoting production and intracellular accumulation of glycerol as compatible solute, as well as regulation of cell wall component. In conclusion, 24.6 mT SMF led to the up-regulation of related genes resulting in enhanced dye biodegradation efficiency and osmo-tolerance of the yeast A1.

Cloning, Expression and Inhibitory Effects on Lewis Lung Carcinoma Cells of rAj-Tspin from Sea Cucumber (Apostichopus japonicus).

In recent years, sea cucumber has become a favorite healthcare food due to its characteristic prevention of cardiovascular diseases, suppression of tumors, as well as enhancement of immunity. In order to screen the anti-tumoral proteins or peptides from sea cucumber (Apostichopus japonicus), its cDNA library was analyzed, and a disintegrin-like and metalloproteinase with thrombospondin type 1 motif, member 13 (ADAMTS13)-like was found. ADAMTS13-like contains 10 thrombospondin 1 (TSP1) domains. Based on analysis of bioinformatics, the third TSP1 domain of this protein, which is further named Aj-Tspin, contains an arginine-glycine-aspartate (RGD) motif. Since our previous studies showed that the recombinant RGD-containing peptide from lampreys showed anti-tumoral activity, the third TSP1 domain of ADAMTS13-like was chosen to evaluate it's effect on tumor proliferation and metastasis, despite the fact it shares almost no homologue with disintegrins from other species. After artificial synthesis, its cDNA sequence, Aj-Tspin, which is composed of 56 amino acids, was subcloned into a pET23b vector and expressed as a recombinant Aj-Tspin (rAj-Tspin) in a soluble form with a molecular weight of 6.976 kDa. Through affinity chromatography, rAj-Tspin was purified as a single protein. Both anti-proliferation and immunofluorescence assays showed that rAj-Tspin suppressed the proliferation of Lewis lung carcinoma (LLC) cells through apoptosis. Adhesion assay also displayed that rAj-Tspin inhibited the adhesion of LLC cells to ECM proteins, including fibronectin, laminin, vitronectin and collagen. Lastly, rAj-Tspin also suppressed the migration and invasion of LLC cells across the filter in transwells. Thus, the above indicates that rAj-Tspin might act as a potential anti-tumoral drug in the future and could also provide information on the nutritional value of sea cucumber.

Cometabolic Degradation of Dibenzofuran and Dibenzothiophene by a Naphthalene-Degrading Comamonas sp. JB.

Comamonas sp. JB was used to investigate the cometabolic degradation of dibenzofuran (DBF) and dibenzothiophene (DBT) with naphthalene as the primary substrate. Dehydrogenase and ATPase activity of the growing system with the presence of DBF and DBT were decreased when compared to only naphthalene in the growing system, indicating that the presence of DBF and DBT inhibited the metabolic activity of strain JB. The pathways and enzymes involved in the cometabolic degradation were tested. Examination of metabolites elucidated that strain JB cometabolically degraded DBF to 1,2-dihydroxydibenzofuran, subsequently to 2-hydroxy-4-(3'-oxo-3'H-benzofuran-2'-yliden)but-2-enoic acid, and finally to catechol. Meanwhile, strain JB cometabolically degraded DBT to 1,2-dihydroxydibenzothiophene and subsequently to the ring cleavage product. A series of naphthalene-degrading enzymes including naphthalene dioxygenase, 1,2-dihydroxynaphthalene dioxygenase, salicylaldehyde dehydrogenase, salicylate hydroxylase, and catechol 2,3-oxygenase have been detected, confirming that naphthalene was the real inducer of expression the degradation enzymes and metabolic pathways were controlled by naphthalene-degrading enzymes.

Performance of the biological aerated filter bioaugmented by a yeast Magnusiomyces ingens LH-F1 for treatment of Acid Red B and microbial community dynamics.

Biological aerated filters (BAFs) were constructed and operated for assessing the effectiveness of bacterial community bioaugmented by a yeast Magnusiomyces ingens LH-F1 for treatment of azo dye Acid Red B (ARB). Dynamics of both bacterial and fungal communities were analyzed through MiSeq sequencing method. The results showed that the bioaugmented BAF displayed obviously better performance for decolorization, COD removal and detoxification of ARB wastewater than the other two which were inoculated with activated sludge (AS) and single M. ingens LH-F1, respectively. Moreover, the bioaugmented BAF also exhibited higher tolerance and stability to shock loading. MiSeq sequencing results demonstrated that both of bacterial and fungal communities remarkably shifted with operation conditions, and the increasing fungal diversity in the bioaugmented BAF was probably related to the relatively high biodegradation and detoxification efficiency. Furthermore, M. ingens LH-F1 survived in the bioaugmented BAF and became one of the dominant fungal species. Therefore, bioaugmentation with yeast M. ingens LH-F1 was successful for improving traditional biological processes aiming at treatment of azo compounds. This method was also potentially useful and meaningful for treating other recalcitrant organic pollutants in practical applications.

Aerobic decolorization of Acid Brilliant Scarlet GR by microbial community and the community dynamics during sequencing batch processes.

In this research, aerobic decolorization of Acid Brilliant Scarlet GR by microbial community was studied. Effects of conditions and dye concentraion on decolorization processes were investigated. Additionally, continuous decolorization was evaluated through sequencing batch tests and the microbial dynamics during this process was analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis. The results showed that 100 mg l⁻¹ of the dye was completely decolorized within 12 h, which was mainly caused by biodegradation. The optimal decolorization conditions were as follows: inoculation size 2.07 g l⁻¹ (wet cell pellet), rotation speed 150 r min⁻¹, pH 5.0-7.0 and 30 °C. The processes were well described by zero-order kinetics, and more than 700 mg l⁻¹ of the dye would inhibit the activity of the consortium. Furthermore, the microbial community exhibited high efficiency in sequencing batch processes for continuous decolorization. Microbial community structure shifted obviously when exposed to higher concentration of the dye (500 mg l⁻¹), and all the dominant microorganisms were affiliated with four different phyla of Actinobacteria, Bacteroidetes, Proteobacteria and Firmicutes.

Influence of dye type and salinity on aerobic decolorization of azo dyes by microbial consortium and the community dynamics.

In this research, aerobic decolorization of different azo dyes by a microbial community was studied. The results showed that more than 80% of four azo dyes (100 mg/L) could be aerobically decolorized by the microbial consortium, however, the time needed was obviously different. Kinetic data indicated that the processes were well described by zero-order kinetics, and the chemical structures of dyes had obvious influence on decolorization rates. On the other hand, effects of salinity on decolorization were also investigated. There was still 40% dye removal for Acid Brilliant Red GR when the salinity increased to 250 g/L. And the microbial community structures with different salinity were detected by PCR-DGGE. It was shown that the same two bacteria were dominant in all decolorization systems, and some typical halophilic microorganisms were found under higher-salt conditions.

Simultaneously enhanced treatment efficiency of simulated hypersaline azo dye wastewater and membrane antifouling by a novel static magnetic field membrane bioreactor (SMFMBR).

Operation performance and membrane fouling of a novel static magnetic field membrane bioreactor (SMFMBR) for treatment of hypersaline azo dye wastewater was investigated. The results showed that SMFMBRs possessed higher efficiency of dye decolorization, COD removal and detoxification than the control MBR without SMF. The (3#) SMFMBR equipped with 305.0 mT (the highest intensity) SMF displayed the best treatment performance among all the four reactors (named as 0#-3#, equipped with SMFs of 0 mT, 95.0 mT, 206.3 mT and 305.0 mT, respectively). Potentially effective microbes belonging to Rhodanobacter, Saccharibacteria genera incertae sedis, Defluviimonas, Cellulomonas, Cutaneotrichosporon, Candida and Pichia were enriched in three SMFMBRs, in both of suspended sludge and bio-cakes. The relative abundance of Candida and Pichia in suspended sludge of 3# SMFMBR was the highest among all the four reactors, suggesting their successful colonization and potentially persistent effect of bioaugmentation. On the other hand, SMF of higher intensity effectively mitigated membrane fouling. Less production of soluble microbial products (SMP) and extracellular polymeric substances (EPS), lower protein/polysaccharide (PN/PS) ratio in SMP and EPS, looser structure of bio-cakes on membrane surface, as well as lower relative abundance of potential fouling causing microbes (mainly bacteria) in microbial communities were determined in 3# SMFMBR than the other three groups.

Improving Azo Dye Decolorization Performance and Halotolerance of Pichia occidentalis A2 by Static Magnetic Field and Possible Mechanisms Through Comparative Transcriptome Analysis.

A halotolerant yeast, Pichia occidentalis A2, was recently isolated that can decolorize various azo dyes. The azo dye decolorization performance of this strain was characterized, including the degradation pathway and detoxification effects of this yeast. Additionally, the effect of static magnetic field (SMF) on this decolorization process was investigated. Activities of key enzymes were analyzed to estimate the change of metabolic activity. Furthermore, possible mechanisms were analyzed through detecting differentially expressed genes between yeast A2 in the absence and presence of SMF. The results indicated that yeast A2 displayed the optimal decolorization performance when the concentrations (in g/L) of glucose, (NH4)2SO4, yeast extract, and NaCl were 4.0, 1.0, 0.1, and ≤30.0, respectively. Meanwhile, the optimal rotation speed, temperature, and pH were 160 rpm, 30°C, and 5.0, respectively. Acid Red B was decolorized and detoxified by yeast A2 through successive steps, including cleavage of the naphthalene-amidine bond, reductive deamination, oxidative deamination/desulfurization, open-loop of hydroxy-substituted naphthalene, and tricarboxylic acid cycle. The dye decolorization efficiency and halotolerance of yeast A2 were enhanced by 206.3 mT SMF. The activities of manganese peroxidase, and laccase were elevated 1.37- and 1.16-fold by 206.3 mT SMF, but lignin peroxidase activity showed little change. It was suggested from the transcriptome sequence that the enhanced halotolerance might be related to the upregulated genes encoding the enzymes or functional proteins related to intracellular synthesis and accumulation of glycerol.

Enhanced azo dye biodegradation performance and halotolerance of Candida tropicalis SYF-1 by static magnetic field (SMF).

Enhancing Acid Red B (ARB) decolorization by growing cells of a halotolerant yeast Candida tropicalis SYF-1 with static magnetic field (SMF) was investigated. Activity of key enzymes and membrane phospholipid fatty acids (PLFAs) were analyzed for estimating the change of metabolic activity and membrane salt-stress response, respectively. Possible enhancement mechanisms were revealed through comparative transcriptome analysis. The results showed that 95.0 mT SMF enhanced ARB decolorization by growing cells of a yeast SYF-1, as well as cell growth and halotolerance capability. Activity of intracellular lignin peroxidase (LiP) and laccase (Lac) was 1.51- and 1.47-fold higher with 95.0 mT SMF than that without SMF, respectively. Unsaturation degree and chain length of dominant PLFAs was increased by 95.0 mT SMF treatment. Several functional protein encoding unigenes related to organics biodegradation, cell growth and halotolerance were 1.17- to 4.19-fold up-regulated in response to 95.0 mT SMF.

The Anti-tumor Activity and Mechanisms of rLj-RGD3 on Human Laryngeal Squamous Carcinoma Hep2 Cells.

BACKGROUND: Laryngeal Squamous Cell Carcinoma (LSCC) is a malignant epithelial tumor with poor prognosis and its incidence rate increased recently. rLj-RGD3, a recombinant protein cloned from the buccal gland of Lampetra japonica, contains three RGD motifs that could bind to integrins on the tumor cells. METHODS: MTT assay was used to detect the inhibitory rate of viability. Giemsa's staining assay was used to observe the morphological changes of cells. Hoechst 33258 and TUNEL staining assay, DNA ladder assay were used to examine the apoptotic. Western blot assay was applied to detect the change of the integrin signal pathway. Wound-healing assay, migration, and invasion assay were used to detect the mobility of Hep2 cells. H&E staining assay was used to show the arrangement of the Hep2 cells in the solid tumor tissues. RESULTS: In the present study, rLj-RGD3 was shown to inhibit the viability of LSCC Hep2 cells in vitro by inducing apoptosis with an IC50 of 1.23µM. Western blot showed that the apoptosis of Hep2 cells induced by rLj- RGD3 was dependent on the integrin-FAK-Akt pathway. Wound healing, transwells, and western blot assays in vitro showed that rLj-RGD3 suppressed the migration and invasion of Hep2 cells by integrin-FAKpaxillin/ PLC pathway which could also affect the cytoskeleton arrangement in Hep2 cells. In in vivo studies, rLj-RGD3 inhibited the growth, tumor volume, and weight, as well as disturbed the tissue structure of the solid tumors in xenograft models of BALB/c nude mice without reducing their body weights. CONCLUSION: These results suggested that rLj-RGD3 is an effective and safe suppressor on the growth and metastasis of LSCC Hep2 cells from both in vitro and in vivo experiments. rLj-RGD3 might be expected to become a novel anti-tumor drug to treat LSCC patients in the near future.

Performance of a newly isolated salt-tolerant yeast strain Pichia occidentalis G1 for degrading and detoxifying azo dyes.

A salt-tolerant yeast named G1 which could decolorize various azo dyes was recently isolated and identified as Pichia occidentalis. Systematic researches on characterization, degradation pathway, detoxification effects and enzymes analysis of this yeast were done. The results showed that the optimal metabolism and growth parameters for strain G1 were: 2.0gL-1 glucose, 0.6gL-1 ammonium sulfate, 0.08gL-1 yeast extract, 30gL-1 NaCl, 160rmin-1, 30°C and pH 5.0. More than 98% of 50mgL-1 Acid Red B (ARB) could be decolorized within 16h under the optimal conditions. Additionally, strain G1 degraded and obviously detoxified ARB through a possible pathway successively consisting of decolorization, deamination/desulfonation and TCA cycle processes. Moreover, NADH-DCIP reductase was estimated as the key reductase for decolorization and ligninases including lignin peroxidase, manganese peroxidase and laccase were important oxidoreductases for further degradation of decolorization intermediates.

A novel integration system of magnetically immobilized cells and a pair of graphite plate-stainless iron mesh electrodes for the bioremediation of coking wastewater.

Magnetically immobilized cells of Comamonas sp. JB coupling with electrode reaction was developed to enhance the treatment efficiency of coking wastewater containing phenol, carbazole (CA), dibenzofuran (DBF), and dibenzothiophene (DBT). The pair of graphite plate-stainless iron mesh electrodes was chosen as the most suitable electrodes. Magnetically immobilized cells coupling with graphite plate-stainless iron mesh electrodes (coupling system) exhibited high degradation activity for all the compounds, which were significantly higher than the sum by single magnetically immobilized cells and electrode reaction at the optimal voltage. Recycling experiments demonstrated that the degradation activity of coupling system increased gradually during eight recycles, indicating that there was a coupling effect between the biodegradation and electrode reaction. Phenol hydroxylase and qPCR assays confirmed that appropriate electrical stimulation could improve phenol hydroxylase activity and promote cells growth. Toxicity assessment suggested the treatment of the coking wastewater by coupling system led to less toxicity than untreated wastewater.

Aerobic decolorization and degradation of azo dyes by suspended growing cells and immobilized cells of a newly isolated yeast Magnusiomyces ingens LH-F1.

Aerobic decolorization and degradation of azo dyes by both of suspended growing cells and immobilized cells of a newly isolated yeast strain LH-F1 were investigated in this study. A yeast strain LH-F1 capable of aerobically decolorizing various azo dyes (20mg/L) was identified as Magnusiomyces ingens basing on 26S rDNA analysis. Meanwhile, effects of different parameters on decolorization of Acid Red B by both of suspended growing cells and immobilized cells of strain LH-F1 were investigated. Furthermore, possible degradation pathway of the dye was proposed through analyzing metabolic intermediates using UV-Vis and HPLC-MS methods. As far as it is known, it is the first systematic research on a M. ingens strain which is capable of efficiently decolorizing azo dyes under aerobic condition. Additionally, this work would also provide a potentially useful microbial strain LH-F1 for treatment of industrial wastewaters containing azo dyes.

Aerobic decolorization and degradation of Acid Orange G (AOG) by suspended growing cells and immobilized cells of a yeast strain Candida tropicalis TL-F1.

In this study, aerobic decolorization and degradation of azo dye Acid Orange G (AOG) by both suspended growing cells and immobilized cells of a yeast strain Candida tropicalis TL-F1 were studied. The effects of different parameters on decolorization of AOG by both growing suspended and immobilized strain TL-F1 were investigated. Furthermore, a possible decolorization mechanism of AOG was proposed through analyzing metabolic intermediates using UV-vis and high-performance liquid chromatography-mass spectrometry (HPLC-MS) methods. Strain TL-F1 could decolorize AOG in both liquid and solid mediums through degradation. The optimal conditions for decolorization with suspended growing cells of strain TL-F1 were as follows: 6-10 g/L sucrose, 5-7 g/L urea, ≥6 % (v/v) inoculation size, ≥160 rpm, 35-40 °C, and pH 5.0-6.0; and those for immobilized cells, the conditions were as follows: 4-6 g/L glucose, 0.2-0.4 g/L urea, 6-10 g/L (wet cell pellets) inoculation size, ≥160 rpm, 35-40 °C, and pH 5.0-7.0. Results of UV-vis scanning spectra suggested that AOG was decolorized through biodegradation, and the possible pathway was proposed through the results of HPLC-MS analysis and related literature. This is a systematic research on aerobic decolorization and degradation of AOG by both suspended and immobilized cells of a C. tropicalis strain.

Aerobic decolorization and degradation of azo dyes by growing cells of a newly isolated yeast Candida tropicalis TL-F1.

The aim of this work was to investigate the decolorization and degradation of azo dyes by growing cells of a new yeast strain TL-F1 which was isolated from the sea mud. Strain TL-F1 was identified as Candida tropicalis on the basis of 28S rDNA analysis. Various azo dyes (20mg/L) were efficiently decolorized through aerobic degradation. Meantime, the effects of different parameters on both decolorization of Acid Brilliant Scarlet GR and growth of strain TL-F1 were investigated. Furthermore, possible degradation pathway of the dye GR was proposed through analysis of metabolic products using UV-Vis spectroscopy and HPLC-MS methods. As far as it is known, it is the first systematic research on a C. tropicalis strain which is capable of efficiently decolorizing various azo dyes under aerobic condition. This work provides a potentially useful microbial strain TL-F1 for treatment of azo dye contaminated wastewater.

Preliminary study on responses of marine nematode community to crude oil contamination in intertidal zone of Bathing Beach, Dalian.

This study investigated the responses of marine nematodes to crude oil contamination in polluted and relatively uncontaminated sites in Dalian Xingang, China, 40 days after an oil spill. Samples were taken at different tide levels on the beach and at different positions along the beach. We present the results of a comparison of nematode assemblages from undisturbed sediment from the Xiajiahezi Bathing Beach with those from sediment from the Xinghai Bathing Beach contaminated with crude oil. A total of 1666 nematodes from 26 genera were found in this study. Results showed significant differences in nematode assemblages between samples from undisturbed controls and those from the polluted area. Nematode abundance, number of species, diversity and species richness decreased significantly with increasing levels of crude oil contamination. Fifteen genera were eliminated and seemed to be composed of species intolerant to crude oil contamination; only the abundance of Marylynnia sp. increased slightly.