Containment of biogenic sulfide production in continuous up-flow packed-bed bioreactors with nitrate or nitrite

Produced water from the Coleville oil field in Saskatchewan, Canada was used to inoculate continuous up-flow packed-bed bioreactors. When 7.8 mM sulfate and 25 mM lactate were present in the in-flowing medium, H(2)S production (souring) by sulfate-reducing bacteria (SRB) was prevented by addition of 17.5 mM nitrate or 20 mM nitrite. Changing the sulfate or lactate concentration of the in-flowing medium indicated that the concentrations of nitrate or nitrite required for containment of souring decreased proportionally with a lowered concentration of the electron donor lactate, while the sulfate concentration of the medium had no effect. Microbial communities were dominated by SRB. Nitrate addition did not give rise to changes in community composition, indicating that lactate oxidation and H(2)S removal were caused by the combined action of SRB and nitrate-reducing, sulfide-oxidizing bacteria (NR-SOB). Apparently the nitrite concentrations formed by these NR-SOB did not inhibit the SRB sufficiently to cause community shifts. In contrast, significant community shifts were observed upon direct addition of high concentrations (20 mM) of nitrite. Strains NO3A and NO2B, two newly isolated, nitrate-reducing bacteria (NRB) emerged as major community members. These were found to belong to the epsilon-division of the Proteobacteria, to be most closely related to Campylobacter lari, and to oxidize lactate with nitrate or nitrite as the electron acceptor. Thus the mechanism of microbial H(2)S removal in up-flow packed-bed bioreactors depended on whether nitrate (SRB/NR-SOB) or nitrite (SRB/NR-SOB as well as NRB) was used. However, the amount of nitrate or nitrite needed to completely remove H(2)S was dictated by the electron donor (lactate) concentration, irrespective of mechanism.     

An ultra-high throughput screening approach for an adenine transferase using fluorescence polarization

We have developed a novel assay for measuring the activity of an enzyme that transfers multiple adenine-containing groups to an acceptor protein. The assay is based on fluorescence polarization (FP) technology in a 1536-well plate format. In the assay, a long wavelength fluorescence tracer, Texas Red (Rhodamine), was covalently conjugated to adenine of the donor substrate through a C(6) spacer arm. As a result of the transfer of the adenine-containing moieties to the acceptor protein substrate, the rotational correlation time of the Texas Red conjugate increased, hence increasing the degree of fluorescence polarization. The pharmacological profile and kinetics of the enzyme measured according to the FP method were consistent with those determined previously by conventional analysis. We have successfully executed a 250,000-compound high throughput screening program based on the FP assay method. The quality and validity of the assay were verified by a variety of statistical analyses.     

580. FRITILLARIA KARELINII

Summary.  Fritillaria karelinii Fisch. ex D.Don ( Liliaceae ) is illustrated. Its relationships with other species and the position of subgenus Rhinopetalum are discussed. Instructions for its cultivation are given.     

Quantitative proteomic analysis of cabernet sauvignon grape cells exposed to thermal stresses reveals alterations in sugar and phenylpropanoid metabolism

Grapes (Vitis vinifera) are a valuable fruit crop and wine production is a major industry. Global warming and expanded range of cultivation will expose grapes to more temperature stresses in future. Our study investigated protein level responses to abiotic stresses, with particular reference to proteomic changes induced by the impact of four different temperature stress regimes, including both hot and cold temperatures, on cultured grape cells. Cabernet Sauvignon cell suspension cultures grown at 26°C were subjected to 14 h of exposure to 34 and 42°C for heat stress, and 18 and 10°C for cold stress. Cells from the five temperatures were harvested in biological triplicates and label‐free quantitative shotgun proteomic analysis was performed. A total of 2042 non‐redundant proteins were identified from the five temperature points. Fifty‐five proteins were only detected in extreme heat stress conditions (42°C) and 53 proteins were only detected at extreme cold stress conditions (10°C). Gene Ontology (GO) annotations of differentially expressed proteins provided insights into the metabolic pathways that are involved in temperature stress in grape cells. Sugar metabolism displayed switching between alternative and classical pathways during temperature stresses. Additionally, nine proteins involved in the phenylpropanoid pathway were greatly increased in abundance at extreme cold stress, and were thus found to be cold‐responsive proteins. All MS data have been deposited in the ProteomeXchange with identifier PXD000977 ( http://proteomecentral.proteomexchange.org/dataset/PXD000977 ).     

Aqueous Fraction of Beta vulgaris Ameliorates Hyperglycemia in Diabetic Mice due to Enhanced Glucose Stimulated Insulin Secretion,Mediated by Acetylcholine and GLP-1, and Elevated Glucose Uptake via Increased Membrane Bound GLUT4 Transporters

BackgroundThe study was designed to investigate the probable mechanisms of anti-hyperglycemic activity of B. Vulgaris.ConclusionFindings of the present study clearly prove the role of Acetylcholine and GLP-1 in the Insulin secreting activity of B. Vulgaris. Increased glucose uptake in the skeletal muscles and subsequent glycogen synthesis may also play a part in the anti-hyperglycemic activity of B. Vulgaris.     

Comparative Genomic and Phylogenetic Analyses of Gammaproteobacterial glg Genes Traced the Origin of the Escherichia coli Glycogen glgBXCAP Operon to the Last Common Ancestor of the Sister Orders Enterobacteriales and Pasteurellales

Production of branched α-glucan, glycogen-like polymers is widely spread in the Bacteria domain. The glycogen pathway of synthesis and degradation has been fairly well characterized in the model enterobacterial species Escherichia coli (order Enterobacteriales, class Gammaproteobacteria), in which the cognate genes (branching enzyme glgB, debranching enzyme glgX, ADP-glucose pyrophosphorylase glgC, glycogen synthase glgA, and glycogen phosphorylase glgP) are clustered in a glgBXCAP operon arrangement. However, the evolutionary origin of this particular arrangement and of its constituent genes is unknown. Here, by using 265 complete gammaproteobacterial genomes we have carried out a comparative analysis of the presence, copy number and arrangement of glg genes in all lineages of the Gammaproteobacteria. These analyses revealed large variations in glg gene presence, copy number and arrangements among different gammaproteobacterial lineages. However, the glgBXCAP arrangement was remarkably conserved in all glg-possessing species of the orders Enterobacteriales and Pasteurellales (the E/P group). Subsequent phylogenetic analyses of glg genes present in the Gammaproteobacteria and in other main bacterial groups indicated that glg genes have undergone a complex evolutionary history in which horizontal gene transfer may have played an important role. These analyses also revealed that the E/P glgBXCAP genes (a) share a common evolutionary origin, (b) were vertically transmitted within the E/P group, and (c) are closely related to glg genes of some phylogenetically distant betaproteobacterial species. The overall data allowed tracing the origin of the E. coli glgBXCAP operon to the last common ancestor of the E/P group, and also to uncover a likely glgBXCAP transfer event from the E/P group to particular lineages of the Betaproteobacteria.     

Sperm status and DNA dose play key roles in sperm/ICSI‐mediated gene transfer in caprine

In relation to the growing recent interest in the establishment of sperm‐mediated gene transfer (SMGT) technology as a convenient and effective method for the simple production of transgenic animals, in this study the possibility of using SMGT to produce transgenic caprine embryos was investigated for the first time. Buck sperm were directly incubated with different concentrations (0–500 ng) of pcDNA/his/Lac‐Z plasmid and used for IVF or ICSI. Sperm used for ICSI were categorized into motile or live‐immotile group before being injected into oocytes. In a separate experiment, dead sperm prepared by repeated freezing/thawing were used for DNA‐incubation before ICSI. Sham injection was carried out by intracytoplasmic injection of approximately the same volume of media containing different doses of DNA using an ICSI needle. Transgene expression and transmission were detected by X‐Gal staining and PCR analysis of developed embryos, respectively. A reasonable blastocyst rate was observed in all the groups. Only embryos in the sham group were negative for transgene transmission. Transgene expression was completely dependent on the delivery technique and status of sperm, and was only observed in the live‐immotile and dead ICSI groups. The results of this study showed that the technique (IVF vs. ICSI vs. sham injection), sperm status (motile vs. live‐immotile vs. dead) and to some extent DNA concentration affect embryo development, transgene transmission and expression. Mol. Reprod. Dev. 77:868–875, 2010. © 2010 Wiley‐Liss, Inc.