Microbial phylogenetic and functional responses within acidified wastewater communities exhibiting enhanced phosphate uptake

Acid stimulated accumulation of insoluble phosphorus within microbial cells is highly beneficial to wastewater treatment but remains largely unexplored. Using single cell analyses and next generation sequencing, the response of active polyphosphate accumulating microbial communities under conditions of enhanced phosphorus uptake under both acidic and aerobic conditions was characterised. Phosphorus accumulation activities were highest under acidic conditions (pH 5.5 > 8.5), where a significant positive effect on bioaccumulation was observed at pH 5.5 when compared to pH 8.5. In contrast to the Betaproteobacteria and Actinobacteria dominated enhanced biological phosphorus removal process, the functionally active polyP accumulators at pH 5.5 belonged to the Gammaproteobacteria, with key accumulators identified as members of the families Aeromonadaceae and Enterobacteriaceae. This study demonstrated a significant enrichment of key polyphosphate kinase and exopolyphosphatase genes within the community metagenome after acidification, concomitant with an increase in P accumulation kinetics.

SOL-GEL Lanthanum Phosphate, Composite and coating for functional Applications.

Lanthanum phosphate is one among the lanthanide family of “Rare Earths” following the periodic table of elements. Known under the generic name, Monazite, the rare earth phosphates have melting points above 1900 °C, high thermal phase stability, low thermal conductivity and thermal expansion coefficient similar to some of the high temperature oxides like alumina and zirconia.

Phosphate Deficiency Induces the Jasmonate Pathway and Enhances Resistance to Insect Herbivory.

During their life cycle, plants are typically confronted by simultaneous biotic and abiotic stresses. Low inorganic phosphate (Pi) is one of the most common nutrient deficiencies limiting plant growth in natural and agricultural ecosystems, while insect herbivory accounts for major losses in plant productivity and impacts ecological and evolutionary changes in plant populations. Here, we report that plants experiencing Pi deficiency induce the jasmonic acid (JA) pathway and enhance their defense against insect herbivory. Pi-deficient Arabidopsis (Arabidopsis thaliana) showed enhanced synthesis of JA and the bioactive conjugate JA-isoleucine, as well as activation of the JA signaling pathway, in both shoots and roots of wild-type plants and in shoots of the Pi-deficient mutant pho1 The kinetics of the induction of the JA signaling pathway by Pi deficiency was influenced by PHOSPHATE STARVATION RESPONSE1, the main transcription factor regulating the expression of Pi starvation-induced genes. Phenotypes of the pho1 mutant typically associated with Pi deficiency, such as high shoot anthocyanin levels and poor shoot growth, were significantly attenuated by blocking the JA biosynthesis or signaling pathway. Wounded pho1 leaves hyperaccumulated JA/JA-isoleucine in comparison with the wild type. The pho1 mutant also showed an increased resistance against the generalist herbivore Spodoptera littoralis that was attenuated in JA biosynthesis and signaling mutants. Pi deficiency also triggered increased resistance to S. littoralis in wild-type Arabidopsis as well as tomato (Solanum lycopersicum) and Nicotiana benthamiana, revealing that the link between Pi deficiency and enhanced herbivory resistance is conserved in a diversity of plants, including crops.

Functional analysis of the 5′ flanking region of the human G6PC3 gene: regulation of promoter activity by glucose, pyruvate, AMP kinase and the pentose phosphate pathway

G6PC3 is a widely expressed isoform of glucose-6-phosphatase, found in many foetal and adult tissues. Mutations in this gene cause developmental abnormalities and severe neutropenia due to abolition of glucose recycling between the cytoplasm and endoplasmic reticulum. Low G6PC3 expression as a result of promoter polymorphisms or dysregulation could produce similar outcomes. Here we investigated the regulation of human G6PC3 promoter activity. HeLa and H4IIE cells were transiently transfected with G6PC3 promoter coupled to the firefly luciferase gene, and promoter activity was measured by dual luciferase assay. Activity was highest in a 453 bp segment of the G6PC3 promoter, from − 455 to − 3 relative to the transcriptional start site. This promoter was unresponsive to glucostatic hormones. Its activity increased significantly between 1 and 5.5 mM glucose, and was not elevated further by glucose concentrations up to 25 mM. Pyruvate increased its activity, but β-hydroxybutyrate and sodium acetate did not. Promoter activity was reduced by inhibitors of hexokinase, glyceraldehyde phosphate dehydrogenase and the oxidative branch of the pentose phosphate pathway, but not by a transketolase inhibitor. Deletion of two adjacent Enhancer-boxes (− 274 to − 279 and − 299 to − 304) reduced promoter activity and abolished the glucose effect, suggesting they could function as a glucose response element. Deletion of an additional downstream 140 bp (− 140 to − 306) restored activity, but not the glucose response, suggesting the presence of repressor elements in this region. 5-Aminoimidazole-4-carboxamide 1-β-d-ribofuranoside (AICAR) reduced promoter activity, showing dependence on AMP-kinase. Regulation of the G6PC3 promoter is thus radically different to that of the hepatic isoform, G6PC. It is sensitive to carbohydrate, but not to fatty acid metabolites, and at much lower physiological concentrations. Based on these findings, we speculate that reduced G6PC3 expression could occur during hypoglycemic episodes in vivo, which are common in utero and in the postnatal period. If such episodes lower G6PC3 expression they could place the foetus or infant at risk of impaired immune function and development, and this possibility requires further examination both in vitro and in vivo.

The Phosphate Source Influences Gene Expression and Quality of Mineralization during In Vitro Osteogenic Differentiation of Human Mesenchymal Stem Cells

For in vitro differentiation of bone marrow-derived mesenchymal stem cells/mesenchymal stromal cells into osteoblasts by 2-dimensional cell culture a variety of protocols have been used and evaluated in the past. Especially the external phosphate source used to induce mineralization varies considerably both in respect to chemical composition and concentration. In light of the recent findings that inorganic phosphate directs gene expression of genes crucial for bone development, the need for a standardized phosphate source in in vitro differentiation becomes apparent. We show that chemical composition (inorganic versus organic phosphate origin) and concentration of phosphate supplementation exert a severe impact on the results of gene expression for the genes commonly used as markers for osteoblast formation as well as on the composition of the mineral formed. Specifically, the intensity of gene expression does not necessarily correlate with a high quality mineralized matrix. Our study demonstrates advantages of using inorganic phosphate instead of beta-glycerophosphate and propose colorimetric quantification methods for calcium and phosphate ions as cost-and time-effective alternatives to X-ray diffraction and Fourier-transform infrared spectroscopy for determination of the calcium phosphate ratio and concentration of mineral matrix formed under in vitro-conditions. We critically discuss the different assays used to assess in vitro bone formation in respect to specificity and provide a detailed in vitro protocol that could help to avoid contradictory results due to variances in experimental design.

Improving the stability of an oxime based electrochemical microsensor for organo-phosphate vapor detection

A micro gas sensor has been developed by our group for the detection of organo-phosphate vapors using an aqueous oxime solution. The analyte diffuses from the high flow rate gas stream through a porous membrane to the low flow rate aqueous phase. It reacts with the oxime PBO (1-Phenyl-1,2,3,-butanetrione 2-oxime) to produce cyanide ions, which are then detected electrochemically from the change in solution potential. Previous work on this oxime based electrochemistry indicated that the optimal buffer pH for the aqueous solution was approximately 10. A basic environment is needed for the oxime anion to form and the detection reaction to take place. At this specific pH, the potential response of the sensor to an analyte (such as acetic anhydride) is maximized. However, sensor response slowly decreases as the aqueous oxime solution ages, by as much as 80% in first 24 hours. The decrease in sensor response is due to cyanide which is produced during the oxime degradation process, as evidenced by the cyanide selective electrode. Solid phase micro-extraction carried out on the oxime solution found several other possible degradation products, including acetic acid, N-hydroxy benzamide, benzoic acid, benzoyl cyanide, 1-Phenyl 1,3-butadione, 2-isonitrosoacetophenone and an imine derived from the oxime. It was concluded that degradation occurred through nucleophilic attack by a hydroxide or oxime anion to produce cyanide, as well as a nitrogen atom rearrangement similar to Beckmann rearrangement. The stability of the oxime in organic solvents is most likely due to the lack of water, and specifically hydroxide ions. The reaction between oxime and organo-phosphate to produce cyanide ions requires hydroxide ions, and therefore pure organic solvents are not compatible with the current micro-sensor electrochemistry. By combining a concentrated organic oxime solution with the basic aqueous buffer just prior to being used in the detection process, oxime degradation can be avoided while preserving the original electrochemical detection scheme. Based on beaker cell experiments with selective cyanide sensitive electrodes, ethanol was chosen as the best organic solvent due to its stabilizing effect on the oxime, minimal interference with the aqueous electrochemistry, and compatibility with the current microsensor material (PMMA). Further studies showed that ethanol had a small effect on micro-sensor performance by reducing the rate of cyanide production and decreasing the overall response time. To avoid incomplete mixing of the aqueous and organic solutions, they were pre-mixed externally at a 10:1 ratio, respectively. To adapt the microsensor design to allow for mixing to take place within the device, a small serpentine channel component was fabricated with the same dimensions and material as the original sensor. This allowed for seamless integration of the microsensor with the serpentine mixing channel. Mixing in the serpentine microchannel takes place via diffusion. Both detector potential response and diffusional mixing improve with increased liquid residence time, and thus decreased liquid flowrate. Micromixer performance was studies at a 10:1 aqueous buffer to organic solution flow rate ratio, for a total rate of 5.5 μL/min. It was found that the sensor response utilizing the integrated micromixer was nearly identical to the response when the solutions were premixed and fed at the same rate.

Ultra high pressure homogenization (UHPH) inactivation of Bacillus amyloliquefaciens spores in phosphate buffered saline (PBS) and milk

Ultra high pressure homogenization (UHPH) opens up new areas for dynamic high pressure assisted thermal sterilization of liquids. Bacillus amyloliquefaciens spores are resistant to high isostatic pressure and temperature and were suggested as potential surrogate for high pressure thermal sterilization validation. B. amyloliquefaciens spores suspended in PBS buffer (0.01 M, pH 7.0), low fat milk (1.5%, pH 6.7), and whole milk (3.5%, pH 6.7) at initial concentration of similar to 10(6) CFU/mL were subjected to UHPH treatments at 200, 300, and 350 MPa with an inlet temperature at similar to 80 degrees C. Thermal inactivation kinetics of B. amyloliquefaciens spores in PBS and milk were assessed with thin wall glass capillaries and modeled using first-order and Weibull models. The residence time during UHPH treatments was estimated to determine the contribution of temperature to spore inactivation by UHPH. No sublethal injury was detected after UHPH treatments using sodium chloride as selective component in the nutrient agar medium. The inactivation profiles of spores in PBS buffer and milk were compared and fat provided no clear protective effect for spores against treatments. Treatment at 200 MPa with valve temperatures lower than 125 degrees C caused no reduction of spores. A reduction of 3.5 log(10)CFU/mL of B. amyloliquefaciens spores was achieved by treatment at 350 MPa with a valve temperature higher than 150 degrees C. The modeled thermal inactivation and observed inactivation during UHPH treatments suggest that temperature could be the main lethal effect driving inactivation.

Absorption Measurements of Periodically Poled Potassium Titanyl Phosphate (PPKTP) at 775 nm and 1550 nm

The efficient generation of second-harmonic light and squeezed light requires non-linear crystals that have low absorption at the fundamental and harmonic wavelengths. In this work the photo-thermal self-phase modulation technique is exploited to measure the absorption coefficient of periodically poled potassium titanyl phosphate (PPKTP) at 1,550 nm and 775 nm. The measurement results are (84 +/- 40) ppm/cm and (127 +/- 24) ppm/cm, respectively. We conclude that the performance of state-of-the-art frequency doubling and squeezed light generation in PPKTP is not limited by absorption.

High alkaline phosphatase activity in phosphate replete waters: The case of two macrotidal estuaries

The occurrence of alkaline phosphatase activity (APA) that hydrolyses organic phosphorus into phosphate (PO4) is commonly related to PO4 deficiency of oceanic, coastal and fresh waters. APA is almost never investigated in PO4-rich estuaries, since very low activities are expected to occur. As a consequence, microbial mineralization of organic phosphorus into PO4 has often been ignored in estuaries. In this study, we examined the importance of potential APA and the associated microbial dynamics in two estuaries, the Aulne and the Elorn (Northwestern France), presenting two different levels of PO4 concentrations. Unexpected high potential APA was observed in both estuaries. Values ranged from 50 to 506 nmol L−1 h−1, which range is usually found in very phosphorus-limited environments. High potential APA values were observed in the oligohaline zone (salinity 5–15) in spring and summer, corresponding to a PO4 peak and a maximum bacterial production of particle-attached bacteria. In all cases, high potential APA was associated with high suspended particulate matter and total particulate phosphorus. The low contribution of the 0.2–1 μm fraction to total APA, the strong correlation between particulate APA and bacterial biomass, and the close relationship between the production of particle-attached bacteria and APA, suggested that high potential APA is mainly due to particle-attached bacteria. These results suggest that the microbial mineralization of organic phosphorus may contribute to an estuarine PO4 production in spring and summer besides physicochemical processes.

Analysis of β-tricalcium phosphate granules prepared with different formulations by nano-computed tomography and scanning electron microscopy

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