Ammonium sensing in nitrogen fixing bacteria: Functions of theglnB andglnD gene products

A plentiful supply of fixed nitrogen as ammonium (or other compounds such as nitrate or amino acids) inhibits nitrogen fixation in free-living bacteria by preventing nitrogenase synthesis and/or activity. Ammonium and nitrate have variable effects on the ability ofRhizobiaceae (Rhizobium, Bradyrhizobium andAzorhizobium) species to nodulate legume hosts and on nitrogen fixation capacity in bacteroid cells contained in nodules or in plant-free bacterial cultures. In addition to effects on nitrogen fixation, excess ammonium can inhibit activity or expression of other pathways for utilization of nitrogenous compounds such as nitrate (through nitrate and nitrite reductase), or glutamine synthetase (GS) for assimilation of ammonium. This paper describes the roles of two key genesglnB andglnD, whose gene products sense levels of fixed nitrogen and initiate a cascade of reactions in response to nitrogen status. While work onEscherichia coli and other enteric bacteria provides the model system,glnB and, to a lesser extent,glnD have been studied in several nitrogen fixing bacteria. Such reports will be reviewed here. Recent results on the identity and function of theglnB andglnD gene products inAzotobacter vinelandii (a free-living soil diazotroph) and inRhizobium leguminosarum biovarviciae, hereinafter designatedR.l. viciae will be presented. New data suggests thatAzotobacter vinelandii probably contains aglnB-like gene and this organism may have twoglnD-like genes (one of which was recently identified and namednfrX). In addition, evidence for uridylylation of theglnB gene product (the PII protein) ofR. l. viciae in response to fixed nitrogen deficiency is presented. Also, aglnB mutant ofR. l. viciae has been isolated; its characteristics with respect to expression of nitrogen regulated genes is described.     

Polysaccharide production by Aureobasidium pullulans: factors affecting polysaccharide formation

Aureobasidium pullulans NRRL 6220 synthesized polysaccharide most actively in media containing sucrose, fructose or maltose with (NH₄)₂SO₄ (0.6 g/l) or ammonium acetate giving greatest yields of the polysaccharide. With (NH₄)₂SO₄ at 1.2 g/l, production of polysaccharide was decreased considerably. Polysaccharide production was highest with an initial pH of 6.5 while biomass formation was better below an initial pH of 5.5. Optimum phosphate concentration for polysaccharide production was 0.03 m.S.M. Badr-Eldin, H.G. El-Masry and O.A. Abd El-Rahman are with the Microbial Chemistry Department, National Research Center, Dokki, Cairo, Egypt; F.H.A. Mohamad is with the Chemical Engineering and Pilot Plant Department, National Research Center, Dokki, Cairo, Egypt. O.M. El-Tayeb is with the Microbiology Department, Faculty of Pharmacy, Cairo University, Egypt.     

Novel Miscanthus hybrids: Modelling productivity on marginal land in Europe using dynamics of canopy development determined by light interception

New biomass crop hybrids for bioeconomic expansion require yield projections to determine their potential for strategic land use planning in the face of global challenges. Our biomass growth simulation incorporates radiation interception and conversion efficiency. Models often use leaf area to predict interception which is demanding to determine accurately, so instead we use low-cost rapid light interception measurements using a simple laboratory-made line ceptometer and relate the dynamics of canopy closure to thermal time, and to measurements of biomass. We apply the model to project the European biomass potentials of new market-ready hybrids for 2020–2030. Field measurements are easier to collect, the calibration is seasonally dynamic and reduces influence of weather variation between field sites. The model obtained is conservative, being calibrated by crops of varying establishment and varying maturity on less productive (marginal) land. This results in conservative projections of miscanthus hybrids for 2020–2030 based on 10% land use conversion of the least (productive) grassland and arable for farm diversification, which show a European potential of 80.7–89.7 Mt year⁻¹ biomass, with potential for 1.2–1.3 EJ year⁻¹ energy and 36.3–40.3 Mt year⁻¹ carbon capture, with seeded Miscanthus sacchariflorus × sinensis displaying highest yield potential. Simulated biomass projections must be viewed in light of the field measurements on less productive land with high soil water deficits. We are attempting to model the results from an ambitious and novel project combining new hybrids across Europe with agronomy which has not been perfected on less productive sites. Nevertheless, at the time of energy sourcing issues, seed-propagated miscanthus hybrids for the upscaled provision of bioenergy offer an alternative source of renewable energy. If European countries provide incentives for growers to invest, seeded hybrids can improve product availability and biomass yields over the current commercial miscanthus variety.     

Plasma Protein Profile of Lactating Women from Two Primary Health Centers in Jakarta,Indonesia

Background:Plasma protein profile test is a potential laboratory method to assess nutritional status especially albumin and globulins levels which reflects protein adequacy. The purpose of this study is to evaluate plasma protein profile of lactating women from two primary health centers in Jakarta.Methods:A cross-sectional study was conducted involving lactating women attending routine maternal examinations in two public primary health centers in Jakarta, Indonesia. The mother’s plasma total protein, albumin, globulin, and immunoglobulin levels were measured.Results:Sixty lactating women were recruited, mostly were 28 years old, slightly overweight, bearing two children, and their recent children were 2 months old. The mean total protein level was 8.13 g/dl, albumin 5.00 g/dl, globulin 3.18 g/dl, albumin: globulin ratio 1.558, mean total IgG level of 1255.98 and mean total IgM level of 135.819. All the measured plasma protein parameters were shown to be not correlated with maternal age, maternal BMI, or maternal parity.DiscussionThe plasma total protein, albumin, globulin, as well as total IgG and IgM level of lactating women in Jakarta were within normal range. These biochemical parameters were shown to be not correlated with anthropometrical data such as maternal age and BMI. The small and relatively homogenous samples were supposed to be the cause of such findings.Conclusion:The plasma protein profile of lactating women in Jakarta was adequate. Further studies are required to evaluate the eligibility of plasma protein profile as biochemical parameter of nutritional status in lactating women.Key Words: Blood protein, lactation, protein, albumin/globulin, immunoglobulin M/G     

Isolated Diaphorase From Bovine Erythrocyte Cannot Reduce Oxidized Cytoglobin (Metcygb)

Background:Cytoglobin (Cygb) is a relatively newly identified globin protein that acts as an oxygen transporter in tissues like hemoglobin (Hb) in erythrocytes and myoglobin (Mb) in muscles. The natural oxidation of the Fe²⁺ ion in its heme group into metglobin (globin-Fe³⁺) made the loses of oxygen binding functions. It is known metHb and metMb can be reduced enzymatically using diaphorase or cyb5r3. However, metCygb reductase had not been previously identified. This study aims to analyze the reducing activity of bovine diaphorase on metCygb.Methods:Diaphorase was isolated from bovine erythrocyte and purified using gel filtration and cationic-exchanger chromatography. Its purity was verified by SDS-PAGE and western blot (WB). The metCygb was obtained from Cygb oxidation with potassium ferrocyanide and its reducing activity was determined by spectroscopy.Results:The diaphorase (MW=30.09 kDa) was purified 10.77-fold from crude enzyme with specific activity against metHb 8.479 U/mg. The purity was confirmed by WB using primary antibody anti-cyb5r3. The purified enzyme reduced metCygb at 0.785 µgmin^-1, which was 13.7 times less than the Vmax of metHb.DiscussionIn conclusion, the purified diaphorase from bovine erythrocytes did not significantly reduce metCygb rather than metHb, a natural substrate in cells.Key Words: Bovine Erythrocyte, Cytochrome B5 Reductase, Diaphorase, Metcytoglobin, Reduction     

Efficacy of intra-lesional injections of meglumine antimoniate once a week vs. twice a week in the treatment of cutaneous leishmaniasis caused by L. tropica in Iran: A randomized controlled clinical trial

Treatment of Cutaneous leishmaniasis (CL) is based on using antimoniate derivatives; patients’ compliance for systemic injections is low due to the pain and systemic complications. In this randomized open trial, the efficacy of intra-lesional (IL) injections of meglumine antimoniate (MA) once a week vs. twice a week in the treatment of Anthrpoponothic CL caused by L. tropica was studied. Eligible volunteer patients were selected according to inclusion/exclusion criteria. The included patients were randomly allocated to receive IL-MA injections once a week or twice a week. The primary outcome was set as complete healing of the lesion(s), and defined as complete re-epithelialization and absence of induration in the lesions. A total of 180 parasitologicaly proven CL patients caused by L. tropica were recruited, 90 patients were treated with weekly IL-MA and 90 patients received IL-MA twice a week. The complete cure was 87.9% vs. 89.2% in the group received weekly and twice a week IL-MA injections, respectively (P = 0.808). Patients’ compliance was acceptable and side effects were limited to a few local allergic reactions to MA. Median time to healing was significantly shorter in patients who received IL-MA twice a week (median ± SE) 37±3.8, (CI: 29.6–44.4) days compared to whom received IL-MA once a week 60±2.3, (CI: 55.6–64.5) days (P< 0.001), however the number of injections was higher in group who received IL-MA twice a week (12 vs. 9 injections). In conclusion, the rate of cure in the group of CL patients with IL-MA twice a week was not significantly different from the group who received IL-MA once a week shorten, but the duration of healing was shorter in the group who received IL-MA twice a week while the group received more injections so is recommended to use IL-MA once a week due to the fact the compliance is acceptable with limited side effects.Clinical Trial Registration: IRCT20081130001475N13; https://en.irct.ir/.     

Database and tools for metabolic network analysis

Metabolic network analysis has attracted much attention in the area of systems biology. It has a profound role in understanding the key features of organism metabolic networks and has been successfully applied in several fields of systems biology, including in silico gene knockouts, production yield improvement using engineered microbial strains, drug target identification, and phenotype prediction. A variety of metabolic network databases and tools have been developed in order to assist research in these fields. Databases that comprise biochemical data are normally integrated with the use of metabolic network analysis tools in order to give a more comprehensive result. This paper reviews and compares eight databases as well as twenty one recent tools. The aim of this review is to study the different types of tools in terms of the features and usability, as well as the databases in terms of the scope and data provided. These tools can be categorised into three main types: standalone tools; toolbox-based tools; and web-based tools. Furthermore, comparisons of the databases as well as the tools are also provided to help software developers and users gain a clearer insight and a better understanding of metabolic network analysis. Additionally, this review also helps to provide useful information that can be used as guidance in choosing tools and databases for a particular research interest.     

Emergence of <Emphasis Type="Italic">Aureobasidium pullulans</Emphasis> as human fungal pathogen and molecular assay for future medical diagnosis

Despite the great importance of Aureobasidium pullulans in biotechnology, the fungus had emerged as an opportunistic human pathogen, especially among immunocompromised patients. Clinical detection of this rare human fungal pathogen presently relies on morphology diagnosis which may be misleading. Thus, a sensitive and accurate quantitative molecular assay for A. pullulans remains lacking. In this study, we presented the microscopy observations of A. pullulans that reveals the phenotypic plasticity of the fungus. A. pullulans-specific primers and molecular beacon probes were designed based on the fungal 18S ribosomal RNA (rRNA) gene. Comparison of two probes with varied quencher chemistry, namely BHQ-1 and Tamra, revealed high amplification efficiency of 104% and 108%, respectively. The optimized quantitative real-time PCR (qPCR) assays could detect and quantify up to 1 pg concentration of A. pullulans DNA. Both assays displayed satisfactory performance parameters at fast thermal cycling mode. The molecular assay has great potential as a molecular diagnosis tool for early detection of fungal infection caused by A. pullulans, which merits future study in clinical diagnosis.     

Cationic liposome-mediated gene delivery: biophysical study and mechanism of internalization

To identify factors affecting cationic liposome-mediated gene delivery efficiency, we studied the relationship between the biophysical characteristics of liposome/DNA complexes (lipoplexes) at different (+/-) charge ratios, their structures as monitored by atomic force microscopy (AFM), and their mechanism(s) of internalization into the cells. Significant changes were observed in the particle size and zeta potential of liposomes and their structures assessed by AFM upon addition of DNA, which depended on (+/-) charge ratios. AFM images showed that lipoplexes were formed from extensively fused and apparently homogeneous lipid particles encapsulating DNA. Lipoplexes were found to internalize the cells through the endocytosis pathway. Lipoplex-cell fusion was found to occur mainly at the plasma membrane level; however, this lipoplex-cell membrane fusion was found to be essential for the uptake of the large particles. A new perspective for the internalization of large lipoplex particles into cytoplasm is discussed.