Whole Mitochondrial and Plastid Genome SNP Analysis of Nine Date Palm Cultivars Reveals Plastid Heteroplasmy and Close Phylogenetic Relationships among Cultivars

Date palm is a very important crop in western Asia and northern Africa, and it is the oldest domesticated fruit tree with archaeological records dating back 5000 years. The huge economic value of this crop has generated considerable interest in breeding programs to enhance production of dates. One of the major limitations of these efforts is the uncertainty regarding the number of date palm cultivars, which are currently based on fruit shape, size, color, and taste. Whole mitochondrial and plastid genome sequences were utilized to examine single nucleotide polymorphisms (SNPs) of date palms to evaluate the efficacy of this approach for molecular characterization of cultivars. Mitochondrial and plastid genomes of nine Saudi Arabian cultivars were sequenced. For each species about 60 million 100 bp paired-end reads were generated from total genomic DNA using the Illumina HiSeq 2000 platform. For each cultivar, sequences were aligned separately to the published date palm plastid and mitochondrial reference genomes, and SNPs were identified. The results identified cultivar-specific SNPs for eight of the nine cultivars. Two previous SNP analyses of mitochondrial and plastid genomes identified substantial intra-cultivar ( = intra-varietal) polymorphisms in organellar genomes but these studies did not properly take into account the fact that nearly half of the plastid genome has been integrated into the mitochondrial genome. Filtering all sequencing reads that mapped to both organellar genomes nearly eliminated mitochondrial heteroplasmy but all plastid SNPs remained heteroplasmic. This investigation provides valuable insights into how to deal with interorganellar DNA transfer in performing SNP analyses from total genomic DNA. The results confirm recent suggestions that plastid heteroplasmy is much more common than previously thought. Finally, low levels of sequence variation in plastid and mitochondrial genomes argue for using nuclear SNPs for molecular characterization of date palm cultivars.     

Characterization of S-Triazine Herbicide Metabolism by a Nocardioides sp. Isolated from Agricultural Soils

Atrazine, a herbicide widely used in corn production, is a frequently detected groundwater contaminant. Nine gram-positive bacterial strains able to use this herbicide as a sole source of nitrogen were isolated from four farms in central Canada. The strains were divided into two groups based on repetitive extragenic palindromic (rep)-PCR genomic fingerprinting with ERIC and BOXA1R primers. Based on 16S ribosomal DNA sequence analysis, both groups were identified as Nocardioides sp. strains. None of the isolates mineralized [ring-U-¹⁴C]atrazine. There was no hybridization to genomic DNA from these strains using atzABC cloned from Pseudomonas sp. strain ADP or trzA cloned from Rhodococcus corallinus. S-Triazine degradation was studied in detail in Nocardioides sp. strain C190. Oxygen was not required for atrazine degradation by whole cells or cell extracts. Based on high-pressure liquid chromatography and mass spectrometric analyses of products formed from atrazine in incubations of whole cells with H₂¹⁸O, sequential hydrolytic reactions converted atrazine to hydroxyatrazine and then to the end product N-ethylammelide. Isopropylamine, the putative product of the second hydrolytic reaction, supported growth as the sole carbon and nitrogen source. The triazine hydrolase from strain C190 was isolated and purified and found to have a K_m for atrazine of 25 μM and a V_max of 31 μmol/min/mg of protein. The subunit molecular mass of the protein was 52 kDa. Atrazine hydrolysis was not inhibited by 500 μM EDTA but was inhibited by 100 μM Mg, Cu, Co, or Zn. Whole cells and purified triazine hydrolase converted a range of chlorine or methylthio-substituted herbicides to the corresponding hydroxy derivatives. In summary, an atrazine-metabolizing Nocardioides sp. widely distributed in agricultural soils degrades a range of s-triazine herbicides by means of a novel s-triazine hydrolase.     

Kinetic Modeling and Error Analysis for Decontamination of Different Petroleum Hydrocarbon Components in Biostimulation of Oily Soil Microcosm

A microcosm study was constructed to investigate the effect of complex co-substrate (corn steep liquor, CSL) addition on indigenous bacterial community, rate and extent of petroleum hydrocarbons (PH) degradation in an oily soil with total petroleum hydrocarbons (TPH) content of 63353 mg kg^?1. TPH degradation was found to be characterized by a rapid phase of degradation during the first three weeks where 76% removal of TPH occurred, followed by a slower degradation phase, where further 7% of the initial TPH was removed by the end of incubation period, 35 d. Branched alkanes are more resistant to microbial degradation than n-alkanes. Furthermore, the unresolved complex mixtures (UCM) of hydrocarbons are less degradable than n- and iso-alkanes. Pristane (Pr) was the most recalcitrant aliphatic compound studied in this work. These results in addition to the extensive bacterial growth observed (from 10⁷ to 10¹⁰ CFU g^?1 soil) give strong support that the addition of CSL resulted in increased degradation rates. The indigenous bacteria grew exponentially during the incubation period of 35 d with a growth rate of 0.26 d^?1. Kinetic modeling was performed to estimate the rates of biodegradation of each hydrocarbon type component in the studied system. Five different error functions were used in this study to evaluate the fitness of the model equation to the obtained experimental data. This showed that the degradation of ∑nC₂₀-nC_24, ∑nC₃₅-nC₄₂ and nC₁₈ can be better represented by a second order model, whereas the TPH, total resolvable peaks (TRP), nC₁₇, UCM, ∑nC₁₀-nC₁₄, ∑nC₁₅-nC_19, ∑nC₂₅-nC₂₉, ∑nC₃₀-nC_34, ∑nC_n, and ∑isoC_n and isoprenoids Pr and phytane (Ph) were similarly following the first order model.     

A Computational Model of a Microfluidic Device to Measure the Dynamics of Oxygen-Dependent ATP Release from Erythrocytes

Erythrocytes are proposed to be involved in blood flow regulation through both shear- and oxygen-dependent mechanisms for the release of adenosine triphosphate (ATP), a potent vasodilator. In a recent study, the dynamics of shear-dependent ATP release from erythrocytes was measured using a microfluidic device with a constriction in the channel to increase shear stress. The brief period of increased shear stress resulted in ATP release within 25 to 75 milliseconds downstream of the constriction. The long-term goal of our research is to apply a similar approach to determine the dynamics of oxygen-dependent ATP release. In the place of the constriction, an oxygen permeable membrane would be used to decrease the hemoglobin oxygen saturation of erythrocytes flowing through the channel. This paper describes the first stage in achieving that goal, the development of a computational model of the proposed experimental system to determine the feasibility of altering oxygen saturation rapidly enough to measure ATP release dynamics. The computational model was constructed based on hemodynamics, molecular transport of oxygen and ATP, kinetics of luciferin/luciferase reaction for reporting ATP concentrations, light absorption by hemoglobin, and sensor characteristics. A linear model of oxygen saturation-dependent ATP release with variable time delay was used in this study. The computational results demonstrate that a microfluidic device with a 100 µm deep channel will cause a rapid decrease in oxygen saturation over the oxygen permeable membrane that yields a measurable light intensity profile for a change in rate of ATP release from erythrocytes on a timescale as short as 25 milliseconds. The simulation also demonstrates that the complex dynamics of ATP release from erythrocytes combined with the consumption by luciferin/luciferase in a flowing system results in light intensity values that do not simply correlate with ATP concentrations. A computational model is required for proper interpretation of experimental data.     

Testicular disorders induced by plant growth regulators: cellular protection with proanthocyanidins grape seeds extract

The present study aims to investigate the adverse effects of plant growth regulators : gibberellic acid (GA₃) and indoleacetic acid (IAA) on testicular functions in rats, and extends to investigate the possible protective role of grape seed extract, proanthocyanidin (PAC). Male rats were divided into six groups; control group, PAC, GA₃, IAA, GA₃ + PAC and IAA + PAC groups. The data showed that GA₃ and IAA caused significant increase in total lipids, total cholesterol, triglycerides, phospholipids and low-density-lipoprotein cholesterol in the serum, concomitant with a significant decrease in high-density-lipoprotein cholesterol, total protein, and testosterone levels. In addition, there was significant decrease in the activity of alkaline phosphatase, acid phosphatase, and gamma-glutamyl transferase. A significant decrease was detected also in epididymyal fructose along with a significant reduction in sperm count. Testicular lipid peroxidation product and hydrogen peroxide (H₂O₂) levels were significantly increased. Meanwhile, the total antioxidant capacity, glutathione, sulphahydryl group content, as well as superoxide dismutase, catalase, and glucose-6-phosphate dehydrogenase activity were significantly decreased. Moreover, there were a number of histopathological testicular changes including Leydig’s cell degeneration, reduction in seminiferous tubule and necrotic symptoms and sperm degeneration in both GA₃- and IAA-treated rats. However, an obvious recovery of all the above biochemical and histological testicular disorders was detected when PAC seed extract was supplemented to rats administered with GA₃ or IAA indicating its protective effect. Therefore it was concluded that supplementation with PAC had ameliorative effects on those adverse effects of the mentioned plant growth regulators through its natural antioxidant properties.     

Acclimation of the Marine Diatom Pseudo-nitzschia australis to Different Salinity Conditions: Effects on Growth,Photosynthetic Activity,and Domoic Acid Content1

Toxic Pseudo-nitzschia australis strains isolated from French coastal waters were studied to investigate their capacity to adapt to different salinities. Their acclimation to different salinity conditions (10, 20, 30, 35, and 40) was studied on growth, photosynthetic capacity, cell biovolume, and domoic acid (DA) content. The strains showed an ability to acclimate to a salinity range from 20 to 40, with optimal growth rates between salinities 30 and 40. The highest cell biovolume was observed at the lowest salinity 20 and was associated with the lowest growth rate. Salinity did not affect the photosynthetic activity; F_v/F_m values and the pigment contents remained high with no significant difference among salinities. An enhanced production of zeaxanthin was, however, observed in the late stationary and decline phases in all cultures except for those acclimated to salinity 20. In terms of cellular toxin content, DA concentrations were 2 to 3-fold higher at the lowest salinity (20) than at the other salinities and were combined with a low amount of dissolved DA. The fact that P. australis accumulate more DA per cell in less saline waters, illustrates that climate-related changes in salinity may affect Pseudo-nitzschia physiology through direct effects on growth, physiology, and toxin content.     

Chemical Variability of the Essential Oil of Pituranthos scoparius from Algeria

The chemical composition of 93 oil samples from the aerial parts of Pituranthos scoparius, harvested in three regions of Algeria, was investigated by GC‐FID, GC/MS and ¹³C‐NMR. Monoterpene hydrocarbons dominated in association with phenylpropanoids and a chemical variability was found highlighting three clusters. The composition of group I (36 samples) exhibited an atypical composition characterized by a very high contents of 6‐methoxyelemicine (13.0 – 59.6%), followed by sabinene (1.1 – 43.0%) and limonene (6.6 – 39.0%), while the samples of group II (12 samples) contained a high content of limonene (9.2 – 44.0%), followed by myristicine (0.0 – 29.4%) and a lower amount of sabinene (0.8 – 2.3%). Group III (45 samples) could be divided in two subgroups. Subgroup SGIIIA was characterized by a very high content of sabinene (28.0 – 55.6%), followed by elemicine (0.0 – 29.1%), while the samples belonging to SGIIIB were characterized by the lower content of sabinene (6.2 – 35.5%) and a significant content of myristicine (1.5 – 32.4%), α‐pinene (4.2 – 31.0%) and dill apiole (0.1 – 31.4%). Each harvested region was characterized by a different chemical composition.     

Cell culture models of fatty acid overload: Problems and solutions

High plasma levels of fatty acids occur in a variety of metabolic diseases. Cellular effects of fatty acid overload resulting in negative cellular responses (lipotoxicity) are often studied in vitro, in an attempt to understand mechanisms involved in these diseases. Fatty acids are poorly soluble, and thus usually studied when complexed to albumins such as bovine serum albumin (BSA). The conjugation of fatty acids to albumin requires care pertaining to preparation of the solutions, effective free fatty acid concentrations, use of different fatty acid species, types of BSA, appropriate controls and ensuring cellular fatty acid uptake. This review discusses lipotoxicity models, the potential problems encountered when using these cellular models, as well as practical solutions for difficulties encountered.     

Isolation, characterization, and structural organization of 10-formyltetrahydrofolate synthetase from spinach leaves

One-carbon metabolism mediated by folate coenzymes plays an essential role in several major cellular processes. In the prokaryotes studied, three folate-dependent enzymes, 10-formyltetrahydrofolate synthetase (EC 6.3.4.3), 5,10-methenyltetrahydrofolate cyclohydrolase (EC 3.5.4.9), and 5,10-methylenetetrahydrofolate dehydrogenase (EC 1.5.1.5) generally exist as monofunctional or bifunctional proteins, whereas in eukaryotes the three activities are present on one polypeptide. The structural organization of these enzymes in plants had not previously been examined. We have purified the 10-formyltetrahydrofolate synthetase activity from spinach leaves to homogeneity and raised antibodies to it. The protein was a dimer with a subunit molecular weight of Mr = 67,000. The Km values for the three substrates, (6R)-tetrahydrofolate, ATP, and formate were 0.94, 0.043, and 21.9 mM, respectively. The enzyme required both monovalent and divalent cations for maximum activity. The 5,10-methylenetetrahydrofolate dehydrogenase and 5,10-methenyltetrahydrofolate cyclohydrolase activities of spinach coeluted separately from the 10-formyltetrahydrofolate synthetase activity on a Matrex Green-A column. On the same column, the activities of the yeast trifunctional C1-tetrahydrofolate synthase coeluted. In addition, antibodies raised to the purified spinach protein immunoinactivated and immunoprecipitated only the 10-formyltetrahydrofolate synthetase activity in a crude extract of spinach leaves. These results suggest that unlike the trifunctional form of C1-tetrahydrofolate synthase in the other eukaryotes examined, 10-formyltetrahydrofolate synthetase in spinach leaves is monofunctional and 5,10-methyl-enetetrahydrofolate dehydrogenase and 5,10-methenyltetrahydrofolate cyclohydrolase appear to be bifunctional. Although structurally dissimilar to the other eukaryotic trifunctional enzymes, the 35 amino-terminal residues of spinach 10-formyltetrahydrofolate synthetase showed 35% identity with six other tetrahydrofolate synthetases.