Nitrate Reductase Research Papers - Academia.edu (original) (raw)

The factors regulating soil microbial stability (e.g. resistance and resilience) are poorly understood, even though microorganisms are essential for ecosystem functioning. In this study, we tested whether a functional microbial community subjected to different primary mild stresses was equally resistant or resilient to a subsequent severe stress. The nitrate reducers were selected as model community and analysed in terms of nitrate reduction rates and genetic structure by narG PCR-restriction fragment length polymorphism fingerprinting. Heat, copper and atrazine were used as primary stresses and mercury at a high concentration as a severe stress. None of the primary stresses had any significant impact on the nitrate reducer community. Although primary stress with heat, copper or atrazine had no effect on the resilience of the nitrate reducer activity to mercury stress, preexposure to copper, another heavy metal, resulted in increased resilience. In contrast, the resistance of both structure and activity of the nitrate reducer community to severe mercury stress was not affected by any of the primary stresses tested. Our experiment suggests that the hypothetical effect of an initial stress on the response of a microbial community to an additional stress is complex and may depend on the relatedness of the two consecutive stresses and the development of positive cotolerance.

- by
- •
- Soil, Biological Sciences, Heavy metals, Drug Resistance
- by Carlo Leifert
- •
- Microbiology, Neuroscience, Physiology, Cardiology

Besides hyperglycemia and hypertension, a recently recognized risk factor for diabetic retinopathy (DR) appears to be hyperlipidemia. While studies using earlier generation lipid lowering agents in DR were disappointing, a randomized trial using HMG-CoA Reductase Inhibitors has strong rationale, though hitherto not attempted. The aim of the present study was to compare the HMG-CoA Reductase Inhibitor, simvastatin, with placebo in patients having DR in a double-blind randomized placebo-controlled trial. Fifty patients with diabetes mellitus (Type 1 and 2) with good glycemic control and hypercholesterolemia and having DR (non-clinically significant macular edema and visual acuity 6/24 or better) in either or both eyes were randomized to simvastatin 20-mg per day or placebo, and were followed up for 180 days. On simvastatin therapy, total cholesterol and low-density lipoprotein cholesterol (LDL-C) decreased (P B0.001, respectively), and the level of high-density lipoprotein cholesterol (HDL-C) increased (P B 0.001). VA improved in four patients using simvastatin, (not statistically different from placebo group) and worsening of VA occurred in seven patients in the placebo group and none in the simvastatin group (P = 0.009). Fundus fluorescein angiography and color fundus photograph showed improvement in one patient in the simvastatin group, while seven patients showed worsening in the placebo group (P = 0.009). The observations of the current study suggest that the HMG-CoA Reductase Inhibitor simvastatin significantly retards the progression of retinopathy in diabetic patients with hypercholesterolemia. The potential of this class of drugs for the primary prevention of DR and other microvascular complications needs to be explored further.

- by Atul Kumar
- •
- Photography, Randomization, Visual acuity, Macular Degeneration

Coexpression of cytochrome P450 monooxygenases (CYPs) and reductase was found in human gastric mucosa with intestinal metaplasia. Immunohistochemistry showed reactivity to P450 reductase in metaplastic epithelial cells and in pyloric gland cells in glands showing intestinal metaplasia. These cells exhibit NADPH-diaphorase activity. Reverse transcription-PCR analysis and Western blotting showed that CYP1A1 and CYP1A2 were expressed in specimens with intestinal metaplasia. Tissue distribution of CYP1A1 coincided with that of P450 reductase. However, immunoreactivity to CYP1A2 protein was localized only in the pyloric gland cells near the intestinal metaplastic gland. Salmonella typhimurium mutagen assay definitively revealed that microsomes prepared from gastric mucosa with intestinal metaplasia, in particular in the pyloric gland, functionally activated benzo(a)pyrene and 2-amino-3-methylimidazo [4,5-f]quinoline. These results indicate that carcinogen activation by CYP enzymes expres...

- by Hideko Sone
- •
- Cancer, Gene expression, Gastric Cancer, Salmonella Typhimurium

This study assessed the efficacy of rosuvastatin for reducing plasma low-density lipoprotein (LDL) cholesterol after 18 weeks of open-label, forced titration in patients with homozygous familial hypercholesterolaemia (hoFH) and compared the efficacy of rosuvastatin 80 mg and atorvastatin 80 mg. Forty-four patients aged 8–63 years (body mass ≥32 kg) entered the study; 4 had portacaval shunts and 11 were receiving plasmapheresis. Patients sequentially received rosuvastatin 20, 40 and 80 mg/day for 6 weeks. Patients remaining in the trial after 18 weeks received double-blind, randomised crossover treatment with rosuvastatin 80 mg/day and atorvastatin 80 mg/day for 6 weeks each. After 18 weeks, mean (S.D.)% reduction from baseline in LDL cholesterol was 22 (21)% overall and by 26 (15)% in 29 patients who neither had a portacaval shunt nor were receiving plasmapheresis. Seventy-two percent of the patients had ≥15% reductions in LDL cholesterol and were considered responders and included patients who had portacaval shunts or were receiving plasmapheresis. Mean LDL reductions from baseline after crossover treatment (n = 21) with rosuvastatin 80 mg and atorvastatin 80 mg were 19 and 18%, respectively. All treatments were well tolerated. Rosuvastatin may have therapeutic value in the management of hoFH.

- by Wim Wilpshaar
- •
- Atherosclerosis, Treatment Outcome, Adolescent, Comparative Study

The Escherichia coli nar promoter is maximally induced under anaerobic conditions in the presence of nitrate ion or under anaerobic only conditions, depending on the genotype of the E. coli nar promoter. Previously, we found that the E. coli nar promoter has some desirable characteristics as an inducible promoter in the E. coli host strains. In this study, the E. coli nar promoter with lacZ gene at the downstream was cloned onto a broad-host-range Gram-negative vector, pBBR122. It was then induced in some other Gram-negative host strains, such as Agrobacterium, Pseudomonas, and Rhizobium, to determine whether the E. coli nar promoter could be used as an inducible promoter in these strains. From shake-flask experiments it was found that the wild-type E. coli nar promoter cloned onto pBBR122, pNW61, was suppressed under aerobic conditions in an Agrobacterium host strain, was partially induced under microaerobic only conditions, and was maximally induced under microaerobic conditions in the presence of nitrate ion. Whereas the mutant-type E. coli nar promoter cloned onto pBBR122, pNW618, was suppressed under aerobic conditions and was maximally induced under microaerobic conditions, regardless of the presence of nitrate ion. This kind of induction pattern observed for the E. coli nar promoters in the Agrobacterium host strain was similar to that observed for the E. coli nar promoters in the E. coli host strain. On the other hand, it was found that both of the E. coli nar promoters, pNW61 and pNW618, in a Pseudomonas host strain were partially induced under aerobic conditions and were maximally induced under microaerobic conditions, regardless of the presence of nitrate. Finally, it was found that both of the E. coli nar promoters in a Rhizobium host strain were minimally induced, regardless of the presence of oxygen or nitrate ion. Similar induction patterns for the three strains were also observed from fermentor experiments in which the dissolved oxygen (DO) level was tightly controlled. From an evolutionary point of view, the results from the three Gram-negative host strains indicate that the E. coli nar promoter system, including the promoter and regulatory proteins, was best conserved in the Agrobacterium host strain and the least conserved in the Rhizobium host strain. From an industrial point of view, the results indicate that the E. coli nar promoter system can be used as an oxygen-dependent inducible promoter in both Agrobacterium and Pseudomonas host strains.

- by Kil Lee
- •
- Biotechnology, Metabolic Engineering, Gene expression, Multidisciplinary
- by Manoj Prasad
- •
- Kinetics, Plant Biology, Genetic Diversity, India

Xylitol is a well-known sugar substitute with low-calorie and anti-cariogenic characteristics.

- by Jin-Ho Choi
- •
- Production, Environmental Biotechnology, Saccharomyces cerevisiae, Nitrate Reductase

Small size at birth continues to be a problem worldwide and many factors, including reduced folate intake and Pb exposure, are associated with it. However, single factors rarely explain the variability in birth weight, suggesting a need for more complex explanatory models. We investigated environment-gene interactions to understand whether folate intake and maternal Pb exposure were associated with smaller newborn size in 474 women with uncomplicated pregnancies delivering term infants in Mexico City. We examined if folate intake modified the negative effects of maternal Pb burden on birth size. We also asked if maternal and infant methylenetetrahydrofolate reductase (MTHFR) genotypes (C677T, A1298C and G1793A) modified the effects of folate intake or Pb exposure on birth size. Women were aged 24·6 (SD 5·1) years; 43·5 % were primiparous. Maternal blood Pb at delivery was 86 (SD 42) μg/l, with 26·7 % having levels ≥100 μg/l. Tibia Pb level was 9·9 (SD 9·8) μg/g. Of the women, 35·3 % had folate intakes <400 μg/d. Birth weight was 3170 (SD 422) g. In covariate-adjusted regressions, higher folate intake was associated with higher birth weight (β 0·04; P<0·05). Higher bone Pb was associated with lower birth weight (β −4·9; P<0·05). Folate intake did not modify the effects of Pb on birth size, nor did MTHFR modify the association between Pb or folate intake on birth size. Although modest, the relationship between maternal nutrition, Pb burden and birth size does underscore the importance of environmental exposures to child health because patterns of fetal growth may affect health outcomes well into adulthood.

- by Alexandre Pereira
- •
- Engineering, Genetics, Nutrition and Dietetics, Nutrition

Abstact -Aspergillus genus is an ubiquitous fungal group that colonizes a wide range of substrates. A total of 100 Tunisian fungal strains isolated at harvest time from fresh and dried grapes were identified within the section Nigri and tested for their ochratoxin A (OTA) producing abilities. Of the isolates, 45% were identified as Aspergillus tubingensis, 34% as Aspergillus niger, 12% as Aspergillus japonicus and 9% as Aspergillus carbonarius. The OTA production was assessed using Czapeck yeast extract agar which revealed that 25% of the isolates belonging to the Aspergillus niger aggregate were OTA producers, while 97% were identified as belonging to the A. carbonarius species. To distinguish between A. niger and A. tubingensis, PCR was used to amplify the ITS-5.8S rDNA for these two strains. The sequence analysis of the PCR products revealed a full similarity between the two species. Thus, a new molecular strategy has been developed based on the targeting of the gene from nitrate reductase (niiA) and nitrite reductase gene (niaD), as well as the intergenic region of the gene (niiA-niaD). The subsequent screening revealed that two new specific primer pairs can be used to identify specifically A. niger and A. tubingensis.

- by Angelique Gautier
- •
- Microbiology, Medical Microbiology, Sequence Analysis, Nitrate Reductase

The narB gene from the cyanobacterium Synechococcus sp. PCC 7942 was cloned downstream from the LacI-regulated promoter Ptrc in the Escherichia coli vector pTrc99A, rendering plasmid pCSLM1. Addition of isopropyl-β-D-thiogalactoside to E. coli (pCSLM1) resulted in the parallel expression of a 76 kDa polypeptide and a nitrate reductase activity with properties identical to those known for nitrate reductase isolated from Synechococcus cells. As is the case for nitrate reductase from Synechococcus cells, either reduced methyl viologen or reduced ferredoxin could be used as an electron donor for the reduction of nitrate catalyzed by E. coli (pCSLM1) extracts. This data shows that narB is a cyanobacterial structural gene for nitrate reductase.

- by Enrique flores
- •
- Genetics, Plant Biology, Cyanobacteria, Plant Molecular Biology

The aim of this study was to determine the technological properties of 2 strains of Staphylococcus simulans (Ssm12, Ssm21) and 4 strains of S. carnosus (SC28, SC31, SC54 and SC55) for the selection of a potential starter cultures to employ in the processing of dry fermented sausages. The strains were studied to evaluate nitrate reductase, proteolytic, lipolytic, decarboxylase and antioxidant activities as well as growth ability at different temperatures, pH and NaCl concentrations. Nitrate reductase activity was determined at 15, 20 and 30°C. By spectrophotometric method all the strains were able to reduce nitrate to nitrite at the different temperatures but these results were not confirmed by the agar plate method. Antioxidant and lipolytic activities were evaluated by spectrophotometric assay. All the strains showed antioxidative enzymes superoxide dismutase (SOD) and catalase whereas all appeared unable to hydrolyse pork fat. Proteolytic activity was determined by agar plate method, spectrophotometric assay (OPA) and sodium dodecyl sulphate gel-electrophoresis (SDS-PAGE) and all strains appeared to be able to hydrolyse sarcoplasmic proteins but not myofibrillar proteins. Finally, all the strains grew at 15 and 20°C, in presence of 10%, 15% and 20% of NaCl and at pH 5.0 and 5.5 and were unable to produce histamine, cadaverine and putrescine. The results showed that all strains studied possess useful technological activities that would make them eligible as a good starter cultures for fermented sausages.

- by Gianluigi Mauriello
- •
- Chemical Engineering, Meat Science, Animal Production, Catalase

Plant extracts have been suggested as a rich as yet unexplored source of potentially useful anti-diabetic drugs. Recent scientific investigations have confirmed the efficacy of many of these preparations, as effective antioxidants; able to regenerate functional pancreatic beta cells and has a hypoglycemic potential. In this study, the exotic rambutan (Nephelium lappaceum L.) fruit's rind has been shown to be effective in inhibiting the carbohydrate hydrolyzing enzymes, alpha glucosidase (EC 50 = 2.7 μg/mL) and alpha amylase (EC 50 = 70.8 μg/mL), at a much more significant level than the drug acarbose (EC 50 = 3500 μg/mL; alpha glucosidase; EC 50 =12 μg/mL; alpha amylase). In addition, the geraniin-enriched ethanolic extracts were able to inhibit the key enzyme in the polyol pathway, aldol reductase (EC 50 = 0.04 μg/mL) and prevent the formation of advanced glycation end-products (AGE) by 43%. These findings indicate that the geraniinstandardized N. lappaceum rind may be a potential source of an anti-hyperglycemic agent.

- by Theanmalar Masilamani and +1
- •
- Management, Chemical Engineering, Food, Glycation

Measurement of nitrite and nitrate, the stable oxidation products of nitric oxide (NO), provides a useful tool to study NO synthesis in vivo and in cell cultures. A simple and rapid fluorometric HPLC method was developed for determination of nitrite through its derivatization with 2,3-diaminonaphthalene (DAN). Nitrite, in standard solution, cell culture medium, or biological samples, readily reacted with DAN under acidic conditions to yield the highly fluorescent 2,3-naphthotriazole (NAT). For analysis of nitrate, it was converted to nitrite by nitrate reductase, followed by the derivatization of nitrite with DAN to form NAT. NAT was separated on a 5-mm reversed-phase C column (15034.6 mm, I.D.) guarded by a 40-mm 8 reversed-phase C column (5034.6 mm, I.D.), and eluted with 15 mM sodium phosphate buffer (pH 7.5) containing 50% 18 methanol (flow-rate, 1.3 ml / min). Fluorescence was monitored with excitation at 375 nm and emission at 415 nm. Mean retention time for NAT was 4.4 min. The fluorescence intensity of NAT was linear with nitrite or nitrate concentrations ranging from 12.5 to 2000 nM in water, cell culture media, plasma and urine. The detection limit for nitrite and nitrate was 10 pmol / ml. Because NAT is well separated from DAN and other fluorescent components present in biological samples, our HPLC method offers the advantages of high sensitivity and specificity as well as easy automation for quantifying picomole levels of nitrite and nitrate in cell culture medium and biological samples.

- by Well Dan
- •
- Cell Culture, Nitric oxide, Nitrate Reductase, High Performance Liquid Chromatography

Gas chromatography-mass spectrometry (GC-MS) and liquid chromatography with fluorescence detection (LC-FL) methods have been proposed for the determination of low level nitrite and nitrate in biological, food and environmental samples. The methods include derivatization of aqueous nitrite with 2,3-diaminonaphthalene (DAN), enzymatic reduction of nitrate to nitrite, extraction with toluene and chromatographic analyses of highly fluorescent 2,3-naphthotriazole (NAT) derivative of nitrite by using GC-MS in selected-ion-monitoring (SIM) mode and LC-FL. Nitrite and nitrate ions in solid samples were extracted with 0.5 M aqueous NaOH by sonication. The recoveries of nitrite and nitrate ions based on GC-MS and LC-FL results were 98.40% and 98.10% and the precision of these methods, as indicated by the relative standard deviations (RSDs) were 1.00% for nitrite and 1.20% for nitrate, respectively. The limits of detection of the GC-MS in SIM mode and LC-FL methods based on S/N = 3 were 0.02 and 0.29 pg/ml for nitrite and 0.03 and 0.30 pg/ml for nitrate, respectively.

- by mehmet akyuz
- •
- Analytical Chemistry, Water, Mass Spectrometry, Secondary Ion Mass Spectrometry

Expression and activity of nitrate reductase (NR; EC 1.6.6.1) and glutamine synthetase (GS; EC 6.3.1.2) were analysed in relation to the rate of CO2 assimilation in cucumber (Cucumis sativus L.) leaves. Intact plants were exposed to different atmospheric CO2 concentrations (100, 400 and 1200 μL L−1) for 14 days. A correlation between the in vivo rates of net CO2 assimilation and the atmospheric CO2 concentrations was observed. Transpiration rate and stomatal conductance remained unaffected by CO2 levels. The exposure of the cucumber plants to rising CO2 concentrations led to a concomitant increase in the contents of starch and soluble sugars, and a decrease in the nitrate content in leaves. At very low CO2, NR and GS expression decreased, in spite of high nitrate contents, whereas at normal and elevated CO2 expression and activity were high although the nitrate content was very low. Thus, in cucumber, NR and GS expression appear to be dominated by sugar levels, rather than by nitrate contents.

- by David Ruano
- •
- Carbon Dioxide, Photosynthesis, Plant Biology, Plant Physiology

Nitrate uptake and reduction to nitrite and ammonium are driven in cyanobacteria by photosynthetically generated assimilatory power, i.e., ATP and reduced ferredoxin. High-affinity nitrate and nitrite uptake takes place in different cyanobacteria through either an ABC-type transporter or a permease from the major facilitator superfamily (MFS). Nitrate reductase and nitrite reductase are ferredoxin-dependent metalloenzymes that carry as prosthetic groups a [4Fe–4S] center and Mo-bis-molybdopterin guanine dinucleotide (nitrate reductase) and [4Fe–4S] and siroheme centers (nitrite reductase). Nitrate assimilation genes are commonly found forming an operon with the structure: nir (nitrite reductase)-permease gene(s)-narB (nitrate reductase). When the cells perceive a high C to N ratio, this operon is transcribed from a complex promoter that includes binding sites for NtcA, a global nitrogen-control regulator that belongs to the CAP family of bacterial transcription factors, and NtcB, a pathway-specific regulator that belongs to the LysR family of bacterial transcription factors. Transcription is also affected by other factors such as CnaT, a putative glycosyl transferase, and the signal transduction protein PII. The latter is also a key factor for regulation of the activity of the ABC-type nitrate/nitrite transporter, which is inhibited when the cells are incubated in the presence of ammonium or in the absence of CO2. Notwithstanding significant advance in understanding the regulation of nitrate assimilation in cyanobacteria, further post-transcriptional regulatory mechanisms are likely to be discovered.

- by Enrique Flores
- •
- Photosynthesis, Plant Biology, Cyanobacteria, Signal Transduction

Low-density lipoprotein (LDL) cholesterol is an established risk factor for coronary heart disease (CHD). In the presence of oxidative stress LDL particles can become oxidized to form a lipoprotein species that is particularly atherogenic. Indeed, oxidized LDL (oxLDL) is pro-inflammatory, it can cause endothelial dysfunction and it readily accumulates within the arterial wall. Several factors may influence the susceptibility of LDL to oxidation, including its size and composition, and the presence of endogenous antioxidant compounds, such as ␣-tocopherol. Individuals with type 2 diabetes or the metabolic syndrome have high levels of oxidative stress and consequently are at an increased risk for cardiovascular events. Reducing oxidative stress has been proposed as a potential approach to prevent CHD and antioxidant vitamins have been employed with encouraging results in experimental models of atherosclerosis. However, clinical trials have not demonstrated consistent beneficial effects of antioxidants on cardiovascular outcomes. Statins (3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors) are the first-line choice for lowering total and LDL cholesterol levels and they have been proven to reduce the risk of CHD. Recent data suggest that these compounds, in addition to their lipid-lowering ability, can also reduce the production of reactive oxygen species and increase the resistance of LDL to oxidation. It may be that the ability of statins to limit the oxidation of LDL contributes to their effectiveness at preventing atherosclerotic disease. (R.S. Rosenson). may also be at increased risk of developing CHD. Indeed, although levels of LDL may not be elevated, its atherogenic potential may be increased by oxidative modification.

- by Robert Rosenson
- •
- Clinical Trial, Atherosclerosis, Treatment Outcome, Oxidative Stress

In order to improve wastewater treatment processes, a need exists for tools that rapidly give detailed insight into the community structure of activated sludge, supplementary to chemical and physical data. In this study, the advantages of microarrays and quantitative polymerase chin reaction (PCR) methods were combined into a real-time PCR assay that allows the simultaneous quantification of phylogenetic and functional genes involved in nitrification and denitrification processes. Simultaneous quantification was possible along a 5-log dynamic range and with high linear correlation (R 2 > 0.98). The specificity of the assay was confirmed by cloning and sequencing analyses of PCR amplicons obtained from activated sludge. The real-time assay was validated on mixed liquid samples of different treatment plants, which varied in nitrogen removal rate. The abundance of ammonia oxidizers was in the order of magnitude of 106 down to 104 ml−1, whereas nitrite oxidizers were less abundant (103–101 order of magnitude). The results were in correspondence with the nitrite oxidation rate in the sludge types. As for the nirS, nirK, and nosZ gene copy numbers, their abundance was generally in the order of magnitude of 108–105. When sludge samples were subjected to lab-scale perturbations, a decrease in nitrification rate was reflected within 18 h in the copy numbers of nitrifier genes (decrease with 1 to 5 log units), whereas denitrification genes remained rather unaffected. These results demonstrate that the method is a fast and accurate tool for the analysis of the (de)nitrifying community structure and size in both natural and engineered environmental samples.

- by kim heylen
- •
- NIRS, Activated Sludge, Applied microbiology, Multidisciplinary

Asymmetric nuclear hybrids have been obtained by fusion of cells from a nitrate-reductase deficient mutant of Nicotiana plumbaginifolia (cnx20) and gamma irradiated protoplasts of Atropa belladonna (irradiation doses tested were 10, 30, 50 and 100 krad). The hybrid formation frequency following selection for genotypic complementation in the NR function was in the range of 0.7%-3.7%. Cytogenetic studies demonstrated that all hybrid plants tested possessed multiple (generally tetra-or hexaploid) sets of N. plumbaginifolia (n = 10) chromosomes along with 6-29 Atropa chromosomes (n = 36), some of which were greatly deleted. Besides the cnxA gene (the selection marker), additional material of the irradiated partner was expressed in some of the lines, as shown by analyses of multiple molecular forms of enzymes. Surprisingly, rDNA genes of both parental species were present and amplified in the majority of the hybrids. Whenever studied, the chloroplast DNA in the hybrids was derived from the Nicotiana parent. Regenerants from some lines flowered and were partially fertile. It is concluded that irradiation of cells of the donor parent before fusion can be used to produce highly asymmetric nuclear hybrid plants, although within the dose range tested, the treatment determined the direction of the elimination but not the degree of elimination of the irradiated genome.

- by Alex Parokonny
- •
- Technology, Biological Sciences, Nitrate Reductase, Enzyme
- by محمود الشاهر
- •
- Nutrition, Plant Biology, Models, Copper

The grapevine (Vitis vinifera L. cv. Riesling) plants subjected to water deficit were studied for changes in relative water content (RWC), leaf dry mass, contents of chlorophyll (Chl), total leaf proteins, free amino acids, and proline, and activities of ribulose-1,5-bisphosphate carboxylase (RuBPC), nitrate reductase (NR), and protease. In water-stressed plants RWC, leaf dry matter, Chl content, net photosynthetic rate (P N), and RuBPC and NR activities were significantly decreased. The total leaf protein content also declined with increase in the accumulation of free amino acids. Concurrently, the protease activity in the tissues was also increased. A significant two-fold increase in proline content was recorded.

- by Massimo Bertamini
- •
- Plant Biology, Water Stress, Nitrate Reductase, Physiological Response

Mycorrhizal symbioses are a rule in nature and may have been crucial in plant and fungal evolution. Ectomycorrhizas are mutualistic interactions between tree roots and soil fungi typical of temperate and boreal forests. The functional analysis of genes involved in developmental and metabolic processes, such as N nutrition, is important to understand the ontogeny of this mutualistic symbiosis. RNA silencing was accomplished in the model mycorrhizal fungus Laccaria bicolor by Agrobacterium-mediated gene transfer. Promoter-directed expression of double-stranded RNA with a partial coding sequence of the Laccaria nitrate reductase gene resulted in fungal transgenic strains strongly affected in growth with nitrate as N source in a medium with high concentration of an utilizable C source. The phenotype correlated with a clear reduction of the target gene mRNA level and this effect was not caused by homologous recombination of the T-DNA in the nitrate reductase locus. Transformation with the hairpin sequence resulted in specific CpG methylation of both the silenced transgene and the nitrate reductase encoding gene. The methylation in the target gene was restricted to the silencing trigger sequence and did not represent the entire genomic DNA in the dikaryon suggesting that the epigenetic changes accompanying RNA silencing affected only the transformed nucleus. Mycorrhization experiments of Populus with strongly silenced fungal strains revealed a systematic inhibition of symbiosis under mycorrhization conditions (C starvation) and nitrate as N source compared with the wild type. This inhibition of mycorrhization was reversed by an organic N source only utilizable by the fungus. These observations would indicate that the plant may be capable of monitoring and detecting the nutritional status of a potential symbiont avoiding the establishment of an unsatisfactory interaction. A probable control mechanism conducted by the plant would inhibit symbiosis when the metabolic profile of the fungal partner is not proper and mutual benefit from the symbiotic structure cannot be assured. Our results are the first report showing that the alteration of expression of a fungal gene impairs mycorrhization. Moreover, this work is the first demonstration of RNA silencing in mycorrhizal fungi and clearly shows that gene knock-down is a powerful tool for further functional genomic studies in mycorrhizal research.

- by Sébastien Duplessis
- •
- Microbiology, Environmental microbiology, Nutrition, RNA

In the present study, species like Mangifera indica, Linn., Cassia fistula, Linn., and Eucalyptus hybrid were exposed to different air pollution load for short duration (active biomonitoring). Variation in biochemical parameters like chlorophyll, protein, soluble sugar, free amino acid, ascorbic acid, nitrate reductase, superoxide dismutase and peroxidase in the leaves were found to be pollution load dependent. These variations can be used as indicators of air pollution for early diagnosis of stress or as a marker for physiological damage to trees prior to the onset of visible injury symptoms. Just by analyzing these biochemical indicators air quality can also be assessed.

- by Mukesh K Gautam
- •
- Bioindicators, Peroxidase, Nitrate Reductase, Superoxide Dismutase

This review summarizes the historic, correlative field evidence and experimental research that implicate cultural eutrophication as a major cause of seagrass disappearance. We summarize the underlying physiological responses of seagrass species, the potential utility of various parameters as indicators of nutrient enrichment in seagrasses, the relatively sparse available information about environmental conditions that exacerbate eutrophication effects, and the better known array of indirect stressors imposed by nutrient over-enrichment that influence seagrass growth and survival. Seagrass recovery following nutrient reductions is examined, as well as the status of modeling efforts to predict seagrass response to changing nutrient regimes.

- by David Tomasko
- •
- Ecological Engineering, Botany, Environmental Science, Oceanography

There is a growing awareness that lignocellulose will be a major raw material for production of both fuel and chemicals in the coming decades—most likely through various fermentation routes. Considerable attention has been given to the problem of finding efficient means of separating the major constituents in lignocellulose (i.e., lignin, hemicellulose, and cellulose) and to efficiently hydrolyze the carbohydrate parts into sugars. In these processes, by-products will inevitably form to some extent, and these will have to be dealt with in the ensuing microbial processes. One group of compounds in this category is the furaldehydes. 2-Furaldehyde (furfural) and substituted 2-furaldehydes—most importantly 5-hydroxymethyl-2-furaldehyde—are the dominant inhibitory compounds found in lignocellulosic hydrolyzates. The furaldehydes are known to have biological effects and act as inhibitors in fermentation processes. The effects of these compounds will therefore have to be considered in the design of biotechnological processes using lignocellulose. In this short review, we take a look at known metabolic effects, as well as strategies to overcome problems in biotechnological applications caused by furaldehydes.

- by João R. M. Almeida
- •
- Metabolism, Biotechnology, Fungi, Applied microbiology

Investigations were carried out to envisage the effect of the cypress canker infection on some features of the thylakoids from field grown cypress (Cupressus sempervirens) needles. Changes in photosynthetic pigments, soluble proteins, soluble starch, starch, ribulose-1,5-bisphosphate carboxylase, nitrate reductase, photosynthetic activities and thylakoid membrane proteins were investigated. The level of total chlorophyll and carotenoids were markedly reduced in cypress canker-infected needles. Similar results were also observed for soluble proteins, nitrate reductase and ribulose-1,5-bisphosphate carboxylase activity. In contrast, the content of soluble starch and sugar were increased in infected needles. In isolated thylakoids, cypress canker infection caused marked inhibition of whole chain and photosystem II activity while the inhibition of photosystem I activity was only marginal. The artificial exogenous electron donors, diphenyl carbazide and hydroxylamine significantly restored the loss of photosystem II activity in infected needles. The same results were obtained when Fv/Fm was evaluated by chlorophyll fluorescence measurements. The marked loss of photosystem II activity in infected needles could be due to the loss of 47, 43, and 15 kDa polypeptides. It is concluded that cypress canker infection inactivates the donor side of photosystem II. This conclusion was confirmed by immunological studies showing that the content of the 33 kDa protein of the water-splitting complex was diminished significantly in infected needles. q

- by Massimo Bertamini
- •
- Microbiology, Plant Biology, Chlorophyll Fluorescence, Nitrate Reductase

Nitric oxide (NO) has now gained significant place in plant science, mainly due to its properties (free radical, small size, no charge, short-lived, and highly diffusible across biological membranes) and multifunctional roles in plant growth, development, and regulation of remarkable spectrum of plant cellular mechanisms. In the last few years, the role of NO in tolerance of plants to abiotic stress has established much consideration. As it is evident from the present review, recent progress on NO potentiality in tolerance of plants to environmental stresses has been impressive. These investigations suggest that NO, itself, possesses antioxidant properties and might act as a signal in activating ROSscavenging enzyme activities under abiotic stress. NO plays an important role in resistance to salt, drought, temperature (high and low), UV-B, and heavy metal stress. Rapidly increasing evidences indicate that NO is essentially involve in several physiological processes; however, there has been much disagreement regarding the mechanism(s) by which NO reduces abiotic stress.

- by Mohamed Al-Whaibi
- •
- Plant Biology, Abiotic Stress, Signal Transduction, Free Radical

A fast screening procedure has been applied for the isolation of staphylococcal strains with possible value as starter cultures in fermented meat products. Colonies isolated in MSA (mannitol-salt phenol red-agar) from a naturally... more

- by jose Flores
- •
- Microbiology, Industrial Biotechnology, Food Microbiology, Catalase

The objective of the present work was to determine the effect of NH4NO3 toxicity on nitrogen metabolism in Phaseolus vulgaris L. cv. Strike. The NH4NO3 was applied to the nutrient solution at 6 (N1), 12 (N2), 18 (N3), and 24 mM (N4). The availability of NO3− and NH4+ in the roots and leaves of green bean proved to be influenced by the different NH4NO3 treatments, being the NO3− and NH4+ translocated towards the aerial part of the plant and subsequently assimilated in the leaves. The most important enzymatic activities within nitrogen metabolism, such as nitrate reductase, nitrite reductase, glutamine synthetase and glutamate synthase were affected negatively by the highest NH4NO3 rate (N4). Finally, in our experiment, possibly the high accumulation of NO3−, and principally of NH4+, in both organs due to the application of high NH4NO3 rates could be the direct causes of the reduced biomass production in plants and of the reduction of N assimilation process in roots and leaves, indicating that green bean plants are highly sensitive to high N levels.

- by esteban sanchez
- •
- Glutamate, Nitrate Reductase, Nitrogen metabolism, Biomass production

Four chickpea genotypes (Tyson, ICC 4958, JG 315 and DCP 92-3) were treated with 1.0 mM and 1.5 mM salicylic acid (SA) and subjected to pre-and post flowering drought stress to analyse its influence on nitrate reductase (NR) activity, relative water content (RWC), proline and antioxidant enzymes activity (superoxide dismutase and peroxidase). Leaf RWC significantly reduced during stress at both the growth stages and ranged between 71.67-74.43% (unstressed) and 67.96-71.67% (stressed), whereas in 1.5 mM SA treated plants leaf RWC increased comparable to the control (unstressed plant). NR activity significantly reduced under stress at the post anthesis stage of growth but was maintained higher in 1.5 mM SA treated plants in all the four genotypes studied. On the other hand, activities of antioxidant enzymes superoxide dismutase (SOD) and peroxidase (POX) were upregulated by drought stress and interestingly further enhanced by 1.5 mM SA treatment. The response of SA (1.5 mM) was relatively more in ICC 4958 and Tyson cultivars of chickpea. Hence, results signify the role of SA in protecting metabolic activity along with regulating the drought response of plants.

- by A Hemantaranjan
- •
- Biochemistry, Agronomy, Agriculture, Nitrate Reductase

Strawberry plants cv. Selva was grown with four NO3 - : NH4 + ratios, (7:0, 6.5:0.5, 6:1, 5.5:1.5 mM) at constant nitrogen concentration of 7 mM/L and constant pH of 5.8. Other essential elements were prepared for the plants adequately.... more

- by Mohammad Ali
- •
- Agriculture, Nitrogen, Nitrate Reductase, Nitrogen metabolism

nutrient conditions and lowest when phosphate was limiting. In general, we found that enzymatic responses were strongly influenced by nutrient availability, indicating its important role in dictating the local responses of the calcifying... more

- by Mirta Teichberg
- •
- Carbon Dioxide, Carbon, Plant Biology, Coral Reefs

This research evaluated the effects of arbuscular mycorrhizal fungi (AMF) on growth, nutritional status, total antioxidant activity (AOX), total soluble phenolics content (TPC), and total nitrate reductase activity (NRA) of leaves and roots of Melilotus albus Medik planted in diesel-contaminated sand (7500 mg kg À1 ). Seedlings of Melilotus either Non inoculated (Non-AMF) or pre-inoculated plants (AMF) with the AMFinoculum Glomus Zac-19 were transplanted to non-contaminated or contaminated sand. After 60 days, diesel significantly reduced plant growth. AMF-plants had no significant greater (64% and 89%, respectively) shoot and leaf dry weight than Non-AMF plants, but AMF plants had lower specific leaf area. AMF-plants had significantly greater content of microelements than non-AMF plants. Regardless diesel contamination, the total AOX and TPC were significantly higher in leaves when compared to roots; in contrast, NRA was higher in roots than leaves. Diesel increased total AOX of leaves, but AMF-plants had significantly lower AOX than non-AMF plants. In contrast, roots of AMF-plants had significantly higher AOX but lower NRA than non-AMF plants. AMF-colonization in roots detected via the fungal alkaline phosphatase activity was significantly reduced by the presence of diesel. AMF-inoculation alleviated diesel toxicity on M. albus by enhancing plant biomass, nutrient content, and AOX activity. In addition, AMF-plants significantly contributed in higher degradation of total petroleum hydrocarbons when compared to non-AMF-plants.

- by Ronald Ferrera-cerrato
- •
- Biomass, Environmental Management, Research evaluation, Multidisciplinary

The development of an efficient and homologous transformation system for Aspergillus oryzae is described. This is based on nitrate reductase (niaD) of the nitrate assimilation pathway. The niaD system offers a number of inherent advantages over many other systems and may be of general use for nitrate-utilising filamentous fungi. Transformation frequencies of up to 800 transformants per microgram DNA are observed with A. oryzae. The preponderance of integration events take place at the resident niaD locus either by gene conversion (41%), single integration (23%) or multiple tandem integration (36%). Heterologous expression of the A. oryzae niaD gene in the filamentous fungi A. nidulans, A. niger and Penicillium chrysogenum is observed. That heterologous putative niaD hybridisation signals are seen with other fungal DNAs affords the opportunity to isolate the corresponding niaD from various fungi in order to develop homolgous transformation. Co-transformation with the introduction of the non-selected markers pyrG, tub-2, and uidA has been achieved.

- by Roland Contreras
- •
- Genetics, Nitrate Reductase, Gene Conversion, Filamentous fungi

Excised roots of wild-type and nitrate-reductase deficient mutant Arabidopsis thaliana (L.) HEYNrI. Can be propagated as sustained root cultures in liquid medium. Culture initiation from a single seedling required a two-day indoleacetic acid treatment at 0.05 mg/1 concentration. Indoleacetic acid facilitated subculture but was not essential for sustained growth. This procedure has allowed the clonal propagation of roots derived from individual wildtype and mutant seedlings for more than 21 months. The cultured roots retained their shoot regeneration ability; however, a controlled desiccation treatment was required to restore it to the level of freshly excised roots. The chromosome number remained diploid and no evidence for the accumulation of recessive mutations was observed. The cultured roots are competent for Agrobacterium-mediated transformation. The sustained root culture technology allowed the maintenance of transgenic tissues in which expression of a dominant, seed-lethal gene (seed-specific pea vicilin promoter fused to diphtheria toxin A chain gene) precluded generative propagation.

- by Laszlo Marton and +1
- •
- Plant Biology, Arabidopsis thaliana, Nitrate Reductase, Clonal propagation

Nitrate is one of the most important stimuli in nitrate reductase (NR) induction, while ammonium is usually an inhibitor. We evaluated the influence of nitrate, ammonium or urea as nitrogen sources on NR activity of the agarophyte Gracilaria chilensis. The addition of nitrate rapidly (2 min) induced NR activity, suggesting a fast post-translational regulation. In contrast, nitrate addition to starved algae

- by Fungyi Chow
- •
- Plant Biology, Environmental Biotechnology, Protein synthesis, Negative Affect

Improved establishment of mycorrhizal tissue culture derived plantlets during acclimatization (Stage IV) is commonly attributed to the enhanced vegetative growth as a resultant of different morphological and in vivo changes. These changes are early and better cuticle development, high biomass accumulation, enhanced physiological changes, improved mineral nutrition, especially phosphorus and micronutrients, etc. However, improvement in establishment of micropropagated plantlets during acclimatization may not only be limited to these mechanisms. In the present investigation, biochemical status of micropropagated grape plantlets in response to six single and a mixed strains of arbuscular-mycorrhizal fungi (AMF) during hardening were studied under glasshouse conditions. The histochemical studies revealed that the mycorrhizal inoculation resulted in accumulation of different biochemicals in the plant system such as chlorophyll, carotenoids, proline, phenol and enzymes like polyphenol oxidase (PPO) and nitrate reductase (NR). The mycorrhizal plantlets showed enhanced survival and improved tolerance against stresses experienced during weaning phase. The mycorrhizal plants also exhibited improved physiological and nutritional status and had higher relative water content (RWC) and photosynthetic rate. These plantlets also accumulated higher N, P, Mg and Fe concentrations, which may primarily be as a result of biochemical changes brought about by mycorrhizal association. Mycorrhizal plantlets also showed www.elsevier.com/locate/scihorti Scientia Horticulturae 106 better hardening under glasshouse conditions. The result suggests that the biochemical changes brought about by mycorrhization were helpful in mitigating different stresses experienced by the tissue culture plants during hardening, which determine their performance later in field. #

- by minakshi grover
- •
- Phosphorus, Tissue culture, Arbuscular mycorrhizal fungi, Nitrate Reductase

The inhibition of seedling growth and nitrate reductase activity in 5 d old Vigna radiata (L.) Wilczek cv. Pusa Baisakhi in the presence of 1.0 mM lead acetate increased drastically, if NaCl (6 and 12 EC) was also present in the nutrient media along with the metal salt. Correspondingly higher endogenous Na þ levels were accumulated in the roots and leaves of seedlings in presence of the two stresses. On the other hand, the levels of endogenous lead get reduced in presence of NaCl in both the roots and leaves. Roots accumulated more Pb 2þ and Na þ than the leaves. The two stresses affect more drastically in the additive or even synergistic manner during the early growth phase of the seedlings.

- by dharmendra gupta
- •
- Biomass, Multidisciplinary, Lead, Nitrogen

An eco-friendly process for the silver nanoparticles (Ag-NPs) biosynthesis was investigated using the fungus Monascus purpureus as a safe and commercially used microorganism. M. purpureus growth filtrate was used for the reduction of the aqueous silver nitrate into Ag-NPs with almost 100% size range of 1-7 nm, which was considered as one of the smallest microbial biosynthesized Ag-NPs. The biosynthesized Ag-NPs were structurally characterized using UV, FTIR, DLS, TEM, and XRD. The biosynthesized Ag-NPs were stable after 3 months with no alteration in shape or size. M. purpureus showed no nitrate reductase activity, whereas its pigments reducing power was decreased after nanoparticles formation indicating its role in the Ag-NPs biosynthesis. The synthesized Ag-NPs exhibited strong antimicrobial activity against different bacteria and yeasts species. The anti-Candida activity of M. purpureus culture filtrate was enhanced in the presence of Ag-NPs; the maximum increase in microbial inhibition was observed against Candida albicans with 1.73 increased folds of inhibition zones, followed by their activity against C. tropicalis and C. glabrata with 0.919-and 0.694-folds of increase, respectively. The obtained results suggest that the biosynthesized Ag-NPs offers a promising cost-effective, eco-friendly, and an alternative way to the conventional method of synthesis that could have wide applications in medicine.

- by Ahmed Tayel
- •
- Microbiology, Medical Microbiology, Metal Nanoparticles, Silver

To investigate the effect of exogenously applied 28-homobrassinolide (HBL) on drought-stressed plants, photosynthesis and antioxidant systems were examined in Indian mustard (Brassica juncea L.). Seedlings of Indian mustard were subjected to drought stress for 7 days at the 8–14 (DS1)/15–21 (DS2) days’ stage of growth and then returned to normal conditions of growth. These seedlings were sprayed with HBL (0.01 μM) at the 30-day stage and were sampled at 60 days to assess the changes in growth, photosynthesis and antioxidant enzymes. Plants exposed to stress at either of the stages of growth exhibited a significant decrease in growth and photosynthesis. The exposure of plants to stress at an earlier stage (DS1) was more inhibitory than that at a later stage (DS2). However, the follow-up treatment with HBL significantly improved the values of these parameters and also overcame the inhibitory effect of water stress. The activity of antioxidant enzymes [catalase (E.C. 1.11.1.6), peroxidase (E.C. 1.11.1.7) and superoxide dismutase (E.C. 1.15.1.1)] and proline content in leaves exhibited an increase in response to both the treatment factors, where their interaction had an additive effect. It was, therefore, concluded that the elevated antioxidant system, at least in part, was responsible for amelioration of the drought stress.

- by Aiman Ahmad
- •
- Plant Biology, Drought Stress, Peroxidase, Water Stress

The effect of ultraviolet-B (UV-B) radiation on plant growth, photosynthetic competence, nitrate content, and extractable activity of nitrate reductase (NR; EC 1.6.6.1) was evaluated in maize (Zea mays L.) seedlings during a 13 days growth period. The results showed that under our experimental conditions UV-B radiation decreased the O 2 evolution rates and the sugar contents of maize leaves without affecting the chlorophyll content, the chloroplast ultrastructure or the plant weight.

- by Silvia Quaggiotti
- •
- Plant Biology, Transcription Regulation, Plant Science, Gene expression

Nitric oxide (NO) and hydrogen peroxide (H 2 O 2 ) are key signalling molecules produced in response to various stimuli and involved in a diverse range of plant signal transduction processes. Nitric oxide and H 2 O 2 have been identified as essential components of the complex signalling network inducing stomatal closure in response to the phytohormone abscisic acid (ABA). A close inter-relationship exists between ABA and the spatial and temporal production and action of both NO and H 2 O 2 in guard cells. This study shows that, in Arabidopsis thaliana guard cells, ABA-mediated NO generation is in fact dependent on ABA-induced H 2 O 2 production. Stomatal closure induced by H 2 O 2 is inhibited by the removal of NO with NO scavenger, and both ABA and H 2 O 2 stimulate guard cell NO synthesis. Conversely, NO-induced stomatal closure does not require H 2 O 2 synthesis nor does NO treatment induce H 2 O 2 production in guard cells. Tungstate inhibition of the NOgenerating enzyme nitrate reductase (NR) attenuates NO production in response to nitrite in vitro and in response to H 2 O 2 and ABA in vivo. Genetic data demonstrate that NR is the major source of NO in guard cells in response to ABA-mediated H 2 O 2 synthesis. In the NR double mutant nia1, nia2 both ABA and H 2 O 2 fail to induce NO production or stomatal closure, but in the nitric oxide synthase deficient Atnos1 mutant, responses to H 2 O 2 are not impaired. Importantly, we show that in the NADPH oxidase deficient double mutant atrbohD/F, NO synthesis and stomatal closure to ABA are severely reduced, indicating that endogenous H 2 O 2 production induced by ABA is required for NO synthesis. In summary, our physiological and genetic data demonstrate a strong inter-relationship between ABA, endogenous H 2 O 2 and NO-induced stomatal closure.

- by Iain Weir
- •
- Plant Biology, Signal Transduction, NO, Nitric oxide

Several studies report about intra-specific trait variation of nitrogen-metabolism related traits, such as N(itrogen)-use efficiency, protein content, N-storage and remobilization in barley and related grass species. The goal of this study was to assess the intra-specific genetic diversity present in primary N-metabolism genes of barley and to investigate the associations of the detected haplotype diversity with malting and kernel quality related traits. Results: Partial sequences of five genes related to N-metabolism in barley (Hordeum vulgare L.) were obtained, i.e. nitrate reductase 1, glutamine synthetase 2, ferredoxin-dependent glutamate synthase, aspartate aminotransferase and asparaginase. Two to five haplotypes in each gene were discovered in a set of 190 various varieties. The development of 33 SNP markers allowed the genotyping of all these barley varieties consisting of spring and winter types. Furthermore, these markers could be mapped in several doubled haploid populations. Cluster analysis based on haplotypes revealed a more uniform pattern of the spring barleys as compared to the winter barleys. Based on linear model approaches associations to several malting and kernel quality traits including soluble N and protein were identified.

- by Inge Matthies
- •
- Genetics, Haplotypes, Cluster Analysis, Nitrogen

Denitrification beds are containers filled with wood by-products that serve as a carbon and energy source to denitrifiers, which reduce nitrate (NO 3 À ) from point source discharges into non-reactive dinitrogen (N 2 ) gas. This study investigates a range of alternative carbon sources and determines rates, mechanisms and factors controlling NO 3 À removal, denitrifying bacterial community, and the adverse effects of these substrates. Experimental barrels (0.2 m 3 ) filled with either maize cobs, wheat straw, green waste, sawdust, pine woodchips or eucalyptus woodchips were incubated at 16.8 C or 27.1 C (outlet temperature), and received NO 3 À enriched water (14.38 mg N L À1 and 17.15 mg N L À1 ). After 2.5 years of incubation measurements were made of NO 3 À eN removal rates, in vitro denitrification rates (DR), factors limiting denitrification (carbon and nitrate availability, dissolved oxygen, temperature, pH, and concentrations of NO 3 À , nitrite and ammonia), copy number of nitrite reductase (nirS and nirK ) and nitrous oxide * Corresponding author. Tel.: þ64 7 858 3700; fax: þ64 7 858 4964. E-mail address: warni1@yahoo.com (S. Warneke). A v a i l a b l e a t w w w . s c i e n c e d i r e c t . c o m j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / w a t r e s

- by Ian McDonald and +1
- •
- Water, Carbon, Water Purification, Multidisciplinary

Artemisia annua L. is an aromatic-antibacterial herb that destroys malarial parasites, lowers fevers and checks bleeding, and of which the secondary compound of interest is artemisinin. Enhanced production of the artemisinin content in... more

- by Masroor A Khan
- •
- Photosynthesis, Plant Biology, Essential Oil, Plant