Nitrite In Medium Research Articles

Isolation of a marine-derived yeast with potential applications in industrial nitrite utilizing.

The nitrite efficient utilization microorganism Wickerhamomyces anomalus RZWP01 was identified. Using nitrite and ammonium as the sole nitrogen source, the nitrogen removal rate of W. anomalus RZWP01 was 97.4% and 87.1%, respectively. W. anomalus RZWP01 grew well in the nitrite medium with glucose or xylose as the only carbon source. However, the W. anomalus RZWP01 cannot live on the nitrite medium with lactose, citric acid, and methanol as the only carbon source. The maximal cell concentration occurred in the nitrite medium with glucose as the only carbon source at a C/N ratio of 20 for 48h, reaching 8.92 × 108 cell mL-1. W. anomalus RZWP01 was the first reported yeast that can efficiently utilize nitrite. The isolation and identification of W. anomalus RZWP01 enriched the microbial resources of nitrite-degrading microorganisms and provided functional microorganisms for the water treatment of sustainable aquaculture.

Manipulating denitrifying sulfide removal of Pseudomonas sp. C27 with nitrite as sole nitrogen source: Shotgun proteomics analysis

Pseudomonas sp. C27 can effectively conduct denitrifying sulfide removal (DSR) reactions via autotrophic denitrification, heterotrophic denitrification and coupled-cycle pathway. This study is the first to cultivate strain C27 using nitrite as the sole nitrogen source, and to conduct shotgun proteomics analysis and investigate the characteristics of DSR growth of strain C27 with nitrate or nitrite as sole nitrogen source. Shotgun proteomics analysis identified a total of 42 specially expressed proteins of C27 in the nitrite medium, based on which, together with chemical analysis data, a supplementary pathway of sulfur metabolism for C27 from sulfate to thiosulfate via intermediate adenosine-5′-phosphosulfate and 3′-phosphoadenosine-5′-phosphosulfate was proposed. Based on the newly revised scheme, the use of nitrite as sole nitrogen source expands the assessible regime of DSR reactions by C27 and provides the potential to recover renewable chemicals such as pyruvate and succinate from the coupled-cycle pathway.

Actions of Prostaglandins on Human Nucleus Pulposus Metabolism Inferred by Cyclooxygenase 2 Inhibition of Cytokine Activated Cells.

ObjectiveLow back pain is frequently treated with nonsteroidal anti-inflammatory drugs (NSAIDs), but little is known about intervertebral disc metabolism of the prostaglandins that are diminished by these drugs. Hence, this study aimed at delineating prostaglandin actions in cytokine activated disc cells by comparing the response of nucleus pulposus (NP) cells to the pro-inflammatory cytokine interleukin (IL)-1β with and without cyclooxygenase 2 (COX-2) inhibition. MethodsNP cells cultured in alginate beads were activated with IL-1β ± the COX-2 inhibitor Sc-58125. Media harvested from cultured cells were analyzed for prostaglandin E2 (PGE2), prostaglandin F2 alpha (PGF2α), IL-6, and matrix metalloproteinase (MMP)-3 by enzyme-linked immunosorbent assay and nitric oxide by Griess Reaction. Gene expression along with proteoglycan, collagen, and total protein synthesis were also measured. ResultsIL-1β increased culture media PGE2 and PGF2α, but decreased proteoglycan and collagen syntheses as well as mRNA expression of the matrix genes aggrecan, versican, collagen I, and collagen II. COX-2 inhibition partially rescued proteoglycan and collagen syntheses and collagen I mRNA, but decreased collagen II mRNA IL-1β activated NP cells. COX-2 inhibition initially enhanced and subsequently reduced IL-1β induced inducible nitric oxide synthase, without altering medium nitrite. IL-1β induction of MMP-3 mRNA was increased by COX-2 inhibition at 24 and 48 hours. ConclusionCOX-2 inhibition alters the response of NP cells to IL-1β, suggesting IL-1β action on disc cells is mediated at least in part through COX-2 and its prostaglandins. COX-2 inhibition produces minimal effects on several key catabolic mediators, with the exception of MMP-3. Blocking COX-2 might be beneficial for maintaining disc matrix since it provides an overall rescue of IL-1 induced loss of matrix protein synthesis.

Intracanal Metformin Promotes Healing of Apical Periodontitis via Suppressing Inducible Nitric Oxide Synthase Expression and Monocyte Recruitment

IntroductionWe have previously shown that intracanal metformin ameliorates apical periodontitis, partially by modulation of osteoblast apoptosis. The action of metformin on other cell types pertinent to the development of apical periodontitis needs to be examined. In the present study, we aimed to analyze whether its effects on the expression of inducible nitric oxide synthase (iNOS) and monocyte recruitment contribute to the therapeutic effect on apical periodontitis. MethodsLipopolysaccharide (LPS)-induced expression of iNOS in a human monocytic cell line, Mono-Mac-6, was assessed by Western blot. The amount of nitrite in culture medium was assessed to quantify nitric oxide (NO) production. C-C motif chemokine ligand-2 (CCL-2) synthesis was measured by enzyme-linked immunosorbent assay. Experimental apical periodontitis in rats was treated with root canal debridement with or without intracanal metformin medication. Lesion progression was assessed by conventional radiography and micro–computed tomographic imaging. Cellular expression of iNOS and the number of monocytes/macrophages were assessed by immunohistochemistry. ResultsMetformin suppressed LPS-induced iNOS and NO production by monocytes. More importantly, metformin inhibited LPS-enhanced CCL-2 synthesis through modulation of the iNOS/NO pathway. Intracanal metformin reduced bone resorption associated with apical periodontitis and suppressed iNOS expression and monocyte recruitment. ConclusionsOur results confirmed the therapeutic efficacy of intracanal metformin for apical periodontitis. Suppression of monocyte recruitment through modulation of iNOS expression and NO production is an important mechanism underlying the beneficial effect of metformin.

Transformation of bisphenol A by electrochemical oxidation in the presence of nitrite and formation of nitrated aromatic by-products

In this contribution, the electrocatalytic abatement of bisphenol A (BPA) with boron-doped diamond (BDD) anode had been conducted in NaNO2 electrolytes. Central composite design was used as statistical multivariate method to optimize the operating parameters adopted (applied current density, flow rate, concentration of NaNO2 and initial pH). The results from response surface analysis indicated that pH was the most influential factor for TOC decay, and a maximum TOC decay of 63.7% was achieved under the optimized operating conditions (9.04 mA cm−2 of applied current density, 400 mL min−1 of flow rate, 10 mM of NaNO2, 4.0 of initial pH and 60 min of electrolysis time). Besides, LC/MS technique was applied to identify the main reaction intermediates, and plenty of nitrated oligomers were detected at the end of the degradation. These by-products were generated via the coaction of coupling reaction of nitrated phenol and electrophilic substitution mediated by nitrogen dioxide radicals. Moreover, our results showed that the degree of nitration depended heavily on the employed initial nitrite concentration. This was one of the very few investigations dealing with nitrophenolic by-products in nitrite medium, and thus the findings exhibited important implications for electrochemical degradation of BPA and its related phenolic pollutants.

Interrelation between endogenous intoxication indicators and nitric oxide metabolites in the infectious process in chronic viral hepatitis C

Aim. To study dynamics and interrelation between final nitric oxide metabolites (nitrites) and endogenous intoxication index (medium mass molecules) in patients with chronic hepatitis C with the effect of antiviral therapy on them.Methods. The study included 99 patients with chronic hepatitis C. The combination therapy for 48 weeks in patients with genotype 1 chronic hepatitis C with pegylated interferon (peginterferon alfa-2a and alfa-2b) in combination with ribavirin and for 24 weeks with daily standard interferon alfa-2b administration in patients with virus genotypes 2 and 3 was conducted. The nitric oxide level was estimated by the content of the nearest metabolite (nitrite) in blood serum using the P.P. Golikov method. Medium mass molecules in the blood serum were determined by the N.I. Gabrielyan method.Results.In patients with chronic hepatitis C high values of nitrogen oxide metabolites and the medium mass molecules in the blood serum were revealed, whereas the level of nitroxidemia and endogenous intoxication by the values of medium mass molecules was associated with the viral replicative capacity value and alanine aminotransferase serum levels. Amid the combination antiviral therapy the nature of nitroxidergy changed with the normalization of nitrogen oxide indicators by 24 weeks and an increase to 48 weeks of treatment. There were significantly positive correlative relations between pronouncement of levels of nitric oxide metabolites and the medium mass molecules before the start of antiviral therapy (R=0.292629, pConclusion. Chronic hepatitis C promotes increase in nitrite ion content and medium mass molecules in the blood serum, their level is associated with virus replication activity and cytolysis severity; antiviral therapy has an effect on the level of endogenous intoxication with the normalization of content of the nitrite ions and medium mass molecules in blood serum by 24th week of treatment.

A dual system formed by the ARC and NR molybdoenzymes mediates nitrite-dependent NO production in Chlamydomonas.

Nitric oxide (NO) is a relevant signal molecule involved in many plant processes. However, the mechanisms and proteins responsible for its synthesis are scarcely known. In most photosynthetic organisms NO synthases have not been identified, and Nitrate Reductase (NR) has been proposed as the main enzymatic NO source, a process that in vitro is also catalysed by other molybdoenzymes. By studying transcriptional regulation, enzyme approaches, activity assays with in vitro purified proteins and in vivo and in vitro NO determinations, we have addressed the role of NR and Amidoxime Reducing Component (ARC) in the NO synthesis process. N\R and ARC were intimately related both at transcriptional and activity level. Thus, arc mutants showed high NIA1 (NR gene) expression and NR activity. Conversely, mutants without active NR displayed an increased ARC expression in nitrite medium. Our results with nia1 and arc mutants and with purified enzymes support that ARC catalyses the NO production from nitrite taking electrons from NR and not from Cytb5-1/Cytb5-Reductase, the component partners previously described for ARC (proposed as NOFNiR, Nitric Oxide-Forming Nitrite Reductase). This NR-ARC dual system would be able to produce NO in the presence of nitrate, condition under which NR is unable to do it.

Mechanisms of toxicity by proinflammatory cytokines in a novel human pancreatic beta cell line, 1.1B4

BackgroundMolecular mechanisms of toxicity and cell damage were investigated in the novel human beta cell line, 1.1B4, after exposure to proinflammatory cytokines — IL-1β, IFN-γ, TNF-α. MethodsMTT assay, insulin radioimmunoassay, glucokinase assay, real time reverse transcription PCR, western blotting, nitrite assay, caspase assay and comet assay were used to investigate mechanisms of cytokine toxicity. ResultsViability of 1.1B4 cells decreased after 18h cytokine exposure. Cytokines significantly reduced cellular insulin content and impaired insulin secretion induced by glucose, alanine, KCl, elevated Ca2+, GLP-1 or forskolin. Glucokinase enzyme activity, regulation of intracellular Ca2+ and PDX1 protein expression were significantly reduced by cytokines. mRNA expression of genes involved in secretory function — INS, GCK, PCSK2 and GJA1 was downregulated in cytokine treated 1.1B4 cells. Upregulation of transcription of genes involved in antioxidant defence — SOD2 and GPX1 was observed, suggesting involvement of oxidative stress. Cytokines also upregulated transcriptions of NFKB1 and STAT1, which was accompanied by a significant increase in NOS2 transcription and accumulation of nitrite in culture medium, implicating nitrosative stress. Oxidative and nitrosative stresses induced apoptosis was evident from increased % tail DNA, DNA fragmentation, caspase 3/7 activity, apoptotic cells and lower BCL2 protein expression. ConclusionsThis study delineates molecular mechanisms of cytokine toxicity in 1.1B4 cells, which agree with earlier observations using human islets and rodent beta cells. General significanceThis study emphasizes the potential usefulness of this cell line as a human beta cell model for research investigating autoimmune destruction of pancreatic beta cells.

Abstract 91: Activation of The Antioxidant Response Element (ARE) in Human Renal Glomerular Endothelial Cells (HRGECs) Reduces Asymmetric Dimethylarginine (ADMA) And Increases Nitric Oxide by Transcriptional Activation of Dimethylarginine Dimethylaminohydrolase (DDAH) and Endothelial Nitric Oxide Synthase (eNOS)

The E2-related factor 2 (Nrf2) binds to AREs and transcriptionally activates several genes that protect against oxidative stress. DDAH phosphorylates and activates eNOS and degrades ADMA which is an endogenous inhibitor of eNOS. We detected 2-3 putative AREs in the promoter regions of the human genes for DDAH-1 and -2, peroxisome proliferator-activated receptor-γ (PPAR-γ) and eNOS. PPAR-γ can increase eNOS expression and eNOS phosphorylation. Therefore, we hypothesized that AREs regulate the DDAH/eNOS/PPAR-γ/ADMA/ NO pathway. Tert-butylhydroquinone (tBHQ) activates Nrf2 by translocation to the nucleus. Incubation of HRGECs for 24 hours with tBHQ enhanced cell proliferation dose-dependently (5-20μM). tBHQ (20μM; n=4) increased medium nitrite by 69 ± 3.7% (P<0.01), intracellular NO (from DAF-FM fluorescence ) by 62 ± 17.1% (P<0.05), and NOS and DDAH activities( from conversion of [ 3 H]-arginine or [ 14 C]-ADMA to [ 3 H]- or [ 14 C] -citrulline) by 58 ± 3.6% and 47 ± 8.5% (P<0.05) and decreased ADMA (by capillary zone electrophoresis) from 1.2±0.03 vs. 2.3±0.16μM/L, (P<0.001, n=6). tBHQ increased the mRNAs and proteins for eNOS by 79 ± 9% and 43 ± 17%, for DDAH-1 by 129 ± 10% and 62 ± 21%, and for DDAH-2 by 119 ± 25% and 48 ± 18% (all P<0.05) and increased the protein expression of PPAR-γ and eNOS phosphorylation and increased tBHQ nuclear translocation markedly. Knockdown of the cell Nrf2 by transfection with a specific siRNA abolished all these effects and attenuated cell proliferation. Chromatin immunoprecipitation (CHIP)-based PCR assays demonstrated that tBHQ enhanced the binding of Nrf2 to one ARE region in the promoters for DDAH-1 and -2 but there was no binding to the eNOS promoter. In conclusion, activation of a specific ARE in the promoter region of the DDAH-1 and -2 genes causes their transcriptional activation. The ensuing enhancement of DDAH activity enhances ADMA metabolism. Nrf2 also increases eNOS expression, NOS activity and NO generation without binding to the ARE for eNOS, likely by upregulation of PPAR-γ. Thus, Nrf2 improves endothelial cell function by coordinating a diverse set of pathways that enhance NO generation.

Characteristics of denitrifying granular sludge grown on nitrite medium in an upflow sludge blanket (USB) reactor

While inoculating pre-acclimatized floccular sludge, nitrite-denitrifying granular sludge was obtained after approximately 40 days of cultivation in a 10 L upflow sludge blanket (USB) reactor. The nitrite removal efficiency was approximately 95% when the nitrite concentration was 50 mg L(-1)at an influent flow rate of 20 L h(-1). The nitrite granular sludge had several notable features including good settleability (110 m h(-1)), high ash content (79%), and high density (1.248 g cm(-3)). The mixed liquor suspended solids (MLSS) of the sludge bed remained at 130.04 g L(-1), at a hydraulic upflow velocity of 2 m h(-1). These interesting characteristics were attributed to a high effluent pH (9.7) caused by the release of alkalinity during the nitrite denitrification process. The surfaces of the granules were dominated by cocci bacteria with a diameter of approximately 3 μm, which could be classified as Nitrosomonas-like species based on our analysis of 16 S rDNA sequences.

Activation of Nrf‐2 Enhances the Function of Human Renal Glomerular Endothelial Cells

Diabetes causes nitric oxide (NO) deficiency dysfunction and nephropathy. Recently, bardoxolone methyl, an activator of nuclear E2‐related factor 2 (Nrf2)–antioxidant response element (ARE) pathway was shown to increase the GFR in diabetic nephropathy, but the mechanism was not established. We detected 1 to 3 potential AREs in the human dimethylarginine dimethylaminohydrolase (DDAH‐1 and ‐2) gene promoter regions. Since DDAH metabolizes the endogenous NOS inhibitor ADMA and upregulates eNOS, we tested the hypothesis that activation of Nrf‐2 enhances endothelial function. Incubation of human renal glomerular endothelial cells (HRGECs) with the Nrf2 activator, tert‐butylhydroquinone (tBHQ; 5–20uM) dose‐dependently and significantly (P<0.05) increased medium nitrite(69±3.7%), NO (62±17.1%) and DDAH activity (47±8.5%), and increased endothelial eNOS, DDAH‐1 and‐2 protein expression by 43±17.1%, 62±20.9% and 48±17.6% (P<0.05). tBHQ markedly increased Nrf‐2 protein in the cell nuclear fraction. In contrast, siRNA knockdown of Nrf‐2, compared tocontrol siRNA, abolished all these tBHQ‐mediated effects and attenuated basal cell proliferation, nitrite and NO production, DDAH activity, and eNOS and DDAH protein expression. In conclusion, activation of Nrf‐2 enhanced endothelial function by simultaneously increasing expression of eNOS and DDAH, thereby increasing NO generation and activity that leads to endothelial cell proliferation. These actions might explain the beneficial effects of Nrf‐2 activation in human diabetic nephropathy.

Study on expression of NO and iNOS in J774A. 1 cells induced by rVvhA

Objective To investigate the effect of recombinant Vibrio vulnificus hemolysin (rVvhA) on the expression of nitric oxide(NO) and induced nitric oxide synthase(iNOS) in J774A. 1 cells.Methods The inhibitory effect of rVvhA on J774A. 1 proliferation was measured by MTF colorimetry technique. The content of nitrite in culture medium was determined by Griess reagent. The expression of iNOS protein and mRNA were measured by immunofluorescence and RT-PCR, respectively. Results The viability of J774A. 1 cells was apparently inhibited when exposed to 0.8 HU/ml rVvhA and up. With the help of IFN-γ, the expression of NO and the activity of iNOS in J774A. 1 cells were remarkably increased when exposed to 0.4 HU/ml rVvhA. Conclusion rVvhA can increase the expression of NO and iNOS, it may play an important role in the pathogenic mechanism of Vibrio vulnificus. Key words: Vibrio vulnificus; Cytolysin; J774A. 1; Nitric oxide; Induced nitric oxide synthase

Screening and denitrification characteristics of a slight halophilic heterotrophic nitrobacteria

A slight halophilic heterotrophic nitrobacteria named gs1 was separated from the matured activated sludge. According to the morphological observation, physiological biochemical tests and sequence analysis of the 16S rDNA, strain gs1 was identified to be as Pseudomonas sp. Sodium acetate and ammonium chloride were used as carbon and nitrogen sources, respectively, to investigate the characteristics of the bacterium. When cultured for 24 h under aerobic conditions, with the removal rates of the NH4+-N and COD being 82.2% and 74.73%, respectively, strain gs1 will have a nitrification function of producing NO2−-N. When cultured for 24 h under aerobic conditions in nitrite medium, the removal rate of the NO2−-N became 100%, and when cultured for 24 h under aerobic conditions in nitrate medium, the removal rate of the NO3−-N became 97%. The result shows that this strain functions for either nitrification or denitrification, i.e., it can complete the full process of biological deoxidation.

C-peptide stimulates nitrites generation via the calcium-JAK2/STAT1 pathway in murine macrophage Raw264.7 cells

Aims C-peptide is a product of pro-insulin cleavage. Numerous studies have demonstrated that C-peptide, although not influencing blood glucose control, may play a role in preventing and potentially reversing some of the chronic complications of type 1 diabetes. The aim of this paper was to present a novel function of C-peptide, focusing on its role in nitric oxide (NO) generation. Main methods Murine macrophage Raw264.7 cells and primary peritoneal macrophages were incubated under control conditions, or with C-peptide. Expression level of iNOS and phosphorylation status of JAK2/STAT1 were analyzed by Western blot. Fluorometric NO assay kit was used to assess the concentration of nitrite in culture medium. Intracellular calcium concentration was measured with calcium indicator dyes, such as Fura-2 and Fluo-3 AM. Key findings C-peptide increased the level of nitrites in murine macrophage Raw264.7 cells. The nitrites production induced by lipopolysaccharide (LPS) was further enhanced by co-treatment of C-peptide. This up-regulation of nitrites generation also correlated with the induction of inducible nitric oxide synthase (iNOS), a prominent marker of macrophage activation. In addition, C-peptide increased the intracellular concentration of calcium levels. Moreover, C-peptide-induced nitrites generation and increase in calcium was observed in freshly isolated primary peritoneal macrophages. In addition, C-peptide specifically affected the Janus activated kinase (JAK)/signal transducer and activated transcription (STAT) pathway. Finally, C-peptide-mediated nitrites generation and JAK2/STAT1 phosphorylation were not detected in the presence of the intracellular calcium chelator, BAPTA-AM. Significance These results suggest that C-peptide may elicit immune modulatory function via modulation of the calcium/JAK–STAT pathway.

Temperature influences the population structure of nitrite‐oxidizing bacteria in activated sludge

Activated sludge from the municipal waste water treatment plant in Hamburg was seeded with mineral nitrite medium and incubated at 10°C, 17°C and 28°C. Dominant lithoautotrophic nitrite-oxidizing bacteria have been identified by electron microscopy, denaturing and temperature gradient gel electrophoresis and PCR with genus-specific primer pairs. The results have revealed the existence of three different genera of nitrite-oxidizing bacteria, namely Nitrospira, Nitrobacter and a novel cold-adapted nitrite oxidizer. As shown by electron microscopy members of the novel genus coexisted in activated sludge together with Nitrospira. A temperature-dependent shift in the population structure was demonstrated by cultivation-based approaches. The novel nitrite oxidizer was enriched at temperatures of 10°C and 17°C. Representatives of Nitrospira were able to grow in a broad temperature range between 10°C and 28°C and members of Nitrobacter were enriched during incubations at 17°C and 28°C. By subsequent 16S rDNA sequencing, the cold-adapted nitrite oxidizer was shown to be closely related to the betaproteobacterium 'Candidatus Nitrotoga arctica'. These findings demonstrated that the population structure of nitrite-oxidizing bacteria in activated sludge is more complex than previously thought and responds strongly to long-term temperature changes.

Suppression of bank vole pancreatic islet function by proinflammatory cytokines

Bank voles kept in captivity may develop diabetes. We recently characterized β-cell function of pancreatic islets from normal and glucose intolerant/diabetic bank voles. These animals had features of both human type 1 and type 2 diabetes. Cytokines may impair β-cell function in both types of diabetes. Presently, we studied how pancreatic islets isolated from normal, i.e. glucose tolerant bank voles are affected by proinflammatory cytokines in vitro. Islets were exposed to hIL-1β (25 U/ml) alone or in combination with hTNF-α (1000 U/ml) + mIFN-γ (1000 U/ml) for 48 h, whereupon islet functions were assessed. Cytokines markedly reduced insulin gene expression and the (pro)insulin biosynthesis rate, which was accompanied by a profound depletion of the islet insulin content. The cytokines did not affect the culture medium insulin accumulation and the glucose oxidation rate, but caused a modest increase in medium nitrite, an indicator of nitric oxide (NO) generation. Cytokine-induced decrease in islet insulin content was not prevented by the preferential inducible NO synthase inhibitor aminoguanidine. These findings suggest that the reduction in islet insulin content is not attributed to enhanced exocytosis or related to altered glucose metabolism, but is rather due to a decline in insulin production. The suppressive effects of islet functions elicited by cytokines seem to be mediated by an NO-independent mechanism. In relation to previous studies on cytokine effects on islets from various species, the bank vole islets show a pattern which more resembles human islets than rat or murine islets.

Determination of trace nitrite by direct and indirect methods using differential pulse polarography and application

Trace nitrite determination plays an important role since it is being used in food-stuff as protecting agent. For this purpose, direct and indirect methods are developed using differential pulse polarography, DPP. It was found that in acidic medium nitrite was escaping as NO. In direct method, the peak of nitrite at about −1.43 V in pH 5 acetate buffer has been used. The calibration graph was linear in the concentration range of nitrite from 6 to 110 μM. No interference was observed from Pb, Zn, Cu, and Cd ions. However, Fe(III) had a peak very near to nitrite peak, this overlap could be eliminated by the addition of EDTA. The reaction between nitrite and sulfite has been used for indirect determination since sulfite peak at −0.7 V is sharp and very reproducible. It was found that sulfite concentration had to be 2.5–3 times larger than nitrite in order to have the reaction quantitative. The limit of quantification for nitrite in this medium was 2 × 10 −6 M. The nitrite in sausage could be determined using both of these methods. While with direct method the result was 0.21 mg g −1, it was 0.24 mg g −1 with indirect method.

Application of anaerobic ammonium-oxidizing consortium to achieve completely autotrophic ammonium and sulfate removal

The simultaneous ammonium and sulfate removal was detected in an anammox reactor, consisted of ammonium oxidization with sulfate deoxidization, and subsequently traditional anammox process, in via of middle medium nitrite with solid sulfur and N2 as the terminal products. The pure anammox bacteria offered a great biotechnological potential for the completely autotrophic reaction indicated by batch tests. Denaturing gradient gel electrophoresis (DGGE) analysis further revealed that a new organism belonging to Planctomycetales was strongly enriched in the defined niche: the redox of ammonium and sulfate. The new species “Anammoxoglobussulfate” was so considered as holding a critical role in the ammonium oxidization with sulfate deoxidization to nitrite. Afterwards, the Planctomyces existing in the bacteria community performed the anammox process together to achieve the complete nitrogen and sulfate removal. The potential use of sulfate as electron acceptor for ammonium oxidizing widens the usage of anammox bacteria.

Nitrite utilization by Chaetoceros muelleri under elevated CO2 concentration

Chaetoceros muelleri (Lemn.) was cultured with nitrite (NO2-) or nitrate (NO3-) as the sole nitrogen source and aerated with air or with CO2-enriched air. Cells of C. muelleri excreted into the medium nitrite produced by reduction of nitrate when grown with 100 mu M NaNO3 as nitrogen source. Accordingly, NO2- concentration reached 10.4 mu M after 95 h at the low CO2 condition (aerated with air); while the maximum NO2- concentration was only around 2.0 mu M at the high CO2 condition (aerated with 5% CO2 in air), furthermore, after 30 h it decreased to no more than 1.0 mu M. NO2- was almost assimilated in 80 h when C. muelleri was cultured at the high CO2 condition with 100 mu M NaNO2 as sole nitrogen source. At the high CO2 condition, after 3 h the activity of nitrite reductase was as much as 50% higher than that at the low CO2 condition. It was indicated that enriched CO2 concentration could inhibit nitrite excretion and enhance nitrite assimilation by cells. Therefore, aeration with enriched CO2 might be an effective way to control nitrite content in aquaculture systems.

Alkaloids from Prosopis juliflora leaves induce glial activation, cytotoxicity and stimulate NO production.

Prosopis juliflora is used for feeding cattle and humans. Intoxication with the plant has been reported, and is characterized by neuromuscular alterations and gliosis. Total alkaloidal extract (TAE) was obtained using acid/basic-modified extraction and was fractionated. TAE and seven alkaloidal fractions, at concentrations ranging 0.03–30 μg/ml, were tested for 24 h on astrocyte primary cultures derived from the cortex of newborn Wistar rats. The MTT test and the measure of LDH activity on the culture medium, revealed that TAE and fractions F29/30, F31/33, F32 and F34/35 were cytotoxic to astrocytes. The EC 50 values for the most toxic compounds, TAE, F31/33 and F32 were 2.87 2.82 and 3.01 μg/ml, respectively. Morphological changes and glial cells activation were investigated through Rosenfeld's staining, by immunocytochemistry for the protein OX-42, specific of activated microglia, by immunocytochemistry and western immunoblot for GFAP, the marker of reactive and mature astrocytes, and by the production of nitric oxide (NO). We observed that astrocytes exposed to 3 μg/ml TAE, F29/30 or F31/33 developed compact cell body with many processes overexpressing GFAP. Treatment with 30 μg/ml TAE and fractions, induced cytotoxicity characterized by a strong cell body contraction, very thin and long processes and condensed chromatin. We also observed that when compared with the control (±1.34%), the proportion of OX-42 positive cells was increased in cultures treated with 30 μg/ml TAE or F29/30, F31/33, F32 and F34/35, with values raging from 7.27% to 28.74%. Moreover, incubation with 3 μg/ml F32, 30 μg/ml TAE, F29/30, F31/33 or F34/35 induced accumulation of nitrite in culture medium indicating induction of NO production. Taken together these results show that TAE and fractionated alkaloids from P. juliflora act directly on glial cells, inducing activation and/or cytotoxicity, stimulating NO production, and may have an impact on neuronal damages observed on intoxicated animals.