ENHANCED PRACTICAL PHOTOSYNTHETIC CO2 MITIGATION

This quarterly report documents significant achievements in the Enhanced Practical Photosynthetic CO{sub 2} Mitigation project during the period from 4/03/2001 through 7/02/2001. Most of the achievements are milestones in our efforts to complete the tasks and subtasks that constitute the project objectives. Note that this version of the quarterly technical report is a revision to add the reports from subcontractors Montana State and Oak Ridge National Laboratories The significant accomplishments for this quarter include: Development of an experimental plan and initiation of experiments to create a calibration curve that correlates algal chlorophyll levels with carbon levels (to simplify future experimental procedures); Completion of debugging of the slug flow reactor system, and development of a plan for testing the pressure drop of the slug flow reactor; Design and development of a new bioreactor screen design which integrates the nutrient delivery drip system and the harvesting system; Development of an experimental setup for testing the new integrated drip system/harvesting system; Completion of model-scale bioreactor tests examining the effects of CO{sub 2} concentration levels and lighting levels on Nostoc 86-3 growth rates; Completion of the construction of a larger model-scale bioreactor to improve and expand testing capabilities and initiation of tests; Substantial progress on construction of a pilot-scale bioreactor; and Preliminary economic analysis of photobioreactor deployment. Plans for next quarter's work are included in the conclusions. A preliminary economic analysis is included as an appendix

ENHANCED PRACTICAL PHOTOSYNTHETIC CO2 MITIGATION

This quarterly report documents significant achievements in the Enhanced Practical Photosynthetic CO{sub 2} Mitigation project during the period from 10/2/2001 through 1/01/2003. As indicated in the list of accomplishments below our current efforts are focused on evaluating candidate organisms and growth surfaces, preparing to conduct long-term tests in the bench-scale bioreactor test systems, and scaling-up the test facilities from bench scale to pilot scale. Specific results and accomplishments for the first quarter of 2003 include: Organisms and Growth Surfaces: (1) Additional thermal features with developed cyanobacterial mats, which might be calcium resistant, were found in the West Thumb area of YNP. New samples were isolated and are being cultured in glass tubes. (2) We checked the motile ability of 8.2.1 Synechococcus s.c. (10) and 3.2.2 Synechococcus s.c. 6. It was found that unicellular isolates 8.2.1 Synechococcus s.c. (10) and 3.2.2 Synechococcus s.c. 1 are phototaxic. Isolate 3.2.2 Synechococcus s.c. 1 currently consists of two populations: one population appears to be positive phototaxic, and second population appears negative phototaxis to the same level of light. This means that the character of screen illumination should be uniform and reasonable for cyanobacterial cells. (3) The aeration of growth media with 5% CO{sub 2} in air stimulates cyanobacterial growth 10-20 times over that with air alone. It is possible the rate of the stimulation of cyanobacterial growth in CRF will be higher because cyanobacteria will be grown as a biofilm. We plan to increase the concentration to 15% CO{sub 2} in air. (4) We are continuing the organizing of our collection of the thermophilic cyanobacteria isolated from Yellowstone National Park. During this reporting period we transferred about 160 samples and discarded about 80 samples with weak growth in standard media as BG-11, D or DH. As result of this work we currently have 13 unialgal cultures of thermophilic cyanobacteria. (5) We are screening the cultures to measure the effects of Omnisil on the growth of 2.1 (III) Mastigicladus laminosum, 8.2.1 Synechococcus s.c.10, 1.2 s.c. 6 Chlorogleopsis spp. and 3.3.2 Synechococcus s.c.1. It was found to date that only 1.2 Chlorogleopsis s.c.6 was able to grow in batch culture in the presence of Omnisil. At the moment we have no explanation for the toxic effect of Omnisil, if in fact it is merely spun silica. Nonetheless, we started the selection of Omnisil resistant clones among several cyanobacterial isolates. This process may require several months. Bioreactor support systems and test facilities: (1) A series of tests was run to demonstrate that the initial mass of algae loaded into the CRF-2 system can be accurately determined based on a statistical sampling procedure. Results from the series of tests indicate that the sampling method can be used to reliably estimate the initial algae mass for a CRF-2 test with about 3% uncertainty. (2) Survivability tests for Sc1.2(2) on Omnisil using drilled-hole header inserts are currently underway. Numerous system problems have delayed the testing, but we are hopeful that the debugging of the system is now complete and the current test is proceeding well. Full test results will be provided in the next quarterly report. (3) Initial tests of both ''drilled hole'' and ''pressure shim'' inserts for the integrated wetting/harvesting screens were successful. Both designs showed good flow characteristics and there was no appreciable clogging noticed during and after the test. Test results for flow rate vs. pressure for three header insert designs showed that the performance of the 0.02 inch shim and the drilled hole design are very similar, and thickening the shim to 0.025 inch has the expected effect of reducing the flow at a given pressure for the shim design, but the basic pressure-flow relationship retains the same form. (4) Since the header inserts will likely be manufactured from stainless steel, leaching tests were carried out to find the tolerance of the organisms to SS 316. The organism SC1.2 was very tolerant to SS 316 and it had no effect on the growth of the organism. (5) The pilot-scale bioreactor construction and debugging is continuing on schedule. We are currently waiting for results from the CRF-2 before finalizing the system design

ENHANCED PRACTICAL PHOTOSYNTHETIC CO2 MITIGATION

This is the first quarterly report of the project Enhanced Practical Photosynthetic CO{sub 2} Mitigation. The official project start date, 10/02/2000, was delayed until 10/31/2000 due to an intellectual property dispute that was resolved. However, the delay forced a subsequent delay in subcontracting with Montana State University, which then delayed obtaining a sampling permit from Yellowstone National Park. However, even with these delays, the project moved forward with some success. Accomplishments for this quarter include: Culturing of thermophilic organisms from Yellowstone; Testing of mesophilic organisms in extreme CO{sub 2} conditions; Construction of a second test bed for additional testing; Purchase of a total carbon analyzer dedicated to the project; Construction of a lighting container for Oak Ridge National Laboratory optical fiber testing; Modified lighting of existing test box to provide more uniform distribution; Testing of growth surface adhesion and properties; Experimentation on water-jet harvesting techniques; and Literature review underway regarding uses of biomass after harvesting. Plans for next quarter's work and an update on the project's web page are included in the conclusions

ENHANCED PRACTICAL PHOTOSYNTHETIC CO2 MITIGATION

This report documents significant achievements in the Enhanced Practical Photosynthetic CO{sub 2} Mitigation project during the period from 10/2/2001 through 10/01/2002. This report marks the end of year 2 of a three-year project as well as the milestone date for completion of Phase I activities. This report includes our current status and defines the steps being taken to ensure that we meet the project goals by the end of year 3. As indicated in the list of accomplishments below our current efforts are focused on evaluating candidate organisms and growth surfaces, preparing to conduct long-term tests in the bench-scale bioreactor test systems, and scaling-up the test facilities from bench scale to pilot scale. Specific results and accomplishments for the third quarter of 2002 include: Organisms and Growth Surfaces: (1) Test results continue to indicate that thermophilic cyanobacteria have significant advantages as agents for practical photosynthetic CO{sub 2} mitigation before mesophilic forms. (2) Additional thermal features with developed cyanobacterial mats, which might be calcium resistant, were found in YNP. (3) Back to back tests show that there is no detectable difference in the growth of isolate 1.2 s.c. (2) in standard and Ca-modified BG-11 medium. The doubling time for both cases was about 12 hours. (4) The cultivation of cyanobacteria in Ca-BG medium should proceed in the pH range between 7 and 7.4, but this suggestion requires additional experiments. (5) Cyanobacteria can be grown in media where sodium is present at trace levels. (6) Ca{sup 2+} enriched medium can be used as a sink for CO{sub 2} under alkaline conditions. (7) Cyanobacteria are able to generate cones of filaments on travertine surfaces. [Travertine is a mixture of CaCO{sub 3} and CaSO{sub 4}]. We hypothesize that SO{sub 4}{sup 2-} stimulates the generation of such cones, because they are not almost generated on CaCO3 surface. On the other hand, we know that plant gas contains elevated concentrations of SO{sub 4}{sup 2-}. We may speculate that the introduction of 11.2 isolate in CRF might significantly increase the productivity of such facility. It is possible that a higher colonization potential for the screens may allow a higher surface productivity than some of the other isolates. (8) The colonization of Omnisil surface is an auto-inducible and time-requiring process. (9) Omnisil coupons should be treated under pH control, preferably using KOH. Bioreactor support systems and test facilities: (1) The pilot-scale bioreactor construction and debugging is continuing on schedule. Tests of the ''natural'' lighting system have shown acceptable levels of illumination for the bioreactor screens using only collected sunlight. (2) Flow control inserts have been designed for the CRF-2 screens, which require header pipes for flow distribution and control. A staggered drilled-hole design and a thick shim design have both shown acceptable performance results (little to no clogging, uniform flow, ability to load algae on to the screen). They will both be tested in the CRF-2 to see which performs the best over long durations, and the best performing design will be used for the pilot scale bioreactor screens

Large Eddy Simulation of Highly Compressible Jets with Tripped Boundary Layers

In high-speed aircraft, supersonic jets used for propulsion can lead to very intense aerodynamically generated acoustic noise. Thus, there is a need to study the aerodynamic and aeroacoustic properties of highly compressible jets. In previous studies, several simulations of supersonic jets have been conducted. Unfortunately, the turbulence intensity at the nozzle exit was dependent on the internal geometry of the nozzle and could not be tuned. This is a pity given that, as shown experimentally and numerically for subsonic and supersonic jets, the boundary layer state of the jet affects the jet flow and noise. In this study, a boundary-layer tripping method permitting to obtain an initially turbulent supersonic jet is studied. The influence of the tripped jet boundary layers on the flow and acoustic fields of the jet is analyzed. The impact of nozzle-exit turbulence levels on the noise radiation and notably on the acoustic components specific to supersonic jets (screech noise, broadband shock-associated noise, mixing noise and Mach wave radiation) is discussed

Lysophosphatidylcholine as an adjuvant for lentiviral vector mediated gene transfer to airway epithelium: effect of acyl chain length

Extent: 11p.Background Poor gene transfer efficiency has been a major problem in developing an effective gene therapy for cystic fibrosis (CF) airway disease. Lysophosphatidylcholine (LPC), a natural airway surfactant, can enhance viral gene transfer in animal models. We examined the electrophysiological and physical effect of airway pre-treatment with variants of LPC on lentiviral (LV) vector gene transfer efficiency in murine nasal airways in vivo. Methods Gene transfer was assessed after 1 week following nasal instillations of a VSV-G pseudotype LV vector pre-treated with a low and high dose of LPC variants. The electrophysiological effects of a range of LPC variants were assessed by nasal transepithelial potential difference measurements (TPD) to determine tight junction permeability. Any physical changes to the epithelium from administration of the LPC variants were noted by histological methods in airway tissue harvested after 1 hour. Results Gene transduction was significantly greater compared to control (PBS) for our standard LPC (palmitoyl/stearoyl mixture) treatment and for the majority of the other LPC variants with longer acyl chain lengths. The LPC variant heptadecanoyl also produced significantly greater LV gene transfer compared to our standard LPC mixture. LV gene transfer and the transepithelial depolarization produced by the 0.1% LPC variants at 1 hour were strongly correlated (r2 = 0.94), but at the 1% concentration the correlation was less strong (r2 = 0.59). LPC variants that displayed minor to moderate levels of disruption to the airway epithelium were clearly associated with higher LV gene transfer. Conclusions These findings show the LPC variants effect on airway barrier function and their correlation to the effectiveness of gene expression. The enhanced expression produced by a number of LPC variants should provide new options for preclinical development of efficient airway gene transfer techniques.Patricia Cmielewski, Don S. Anson and David W. Parson

The Cell Adhesion Molecule “CAR” and Sialic Acid on Human Erythrocytes Influence Adenovirus In Vivo Biodistribution

Although it has been known for 50 years that adenoviruses (Ads) interact with erythrocytes ex vivo, the molecular and structural basis for this interaction, which has been serendipitously exploited for diagnostic tests, is unknown. In this study, we characterized the interaction between erythrocytes and unrelated Ad serotypes, human 5 (HAd5) and 37 (HAd37), and canine 2 (CAV-2). While these serotypes agglutinate human erythrocytes, they use different receptors, have different tropisms and/or infect different species. Using molecular, biochemical, structural and transgenic animal-based analyses, we found that the primary erythrocyte interaction domain for HAd37 is its sialic acid binding site, while CAV-2 binding depends on at least three factors: electrostatic interactions, sialic acid binding and, unexpectedly, binding to the coxsackievirus and adenovirus receptor (CAR) on human erythrocytes. We show that the presence of CAR on erythrocytes leads to prolonged in vivo blood half-life and significantly reduced liver infection when a CAR-tropic Ad is injected intravenously. This study provides i) a molecular and structural rationale for Ad–erythrocyte interactions, ii) a basis to improve vector-mediated gene transfer and iii) a mechanism that may explain the biodistribution and pathogenic inconsistencies found between human and animal models

Interventions to prevent youth violence in Latin America: a systematic review

Objectives: This review aims to summarise evidence on the effectiveness of interventions to prevent youth violence in Latin America. Methods: A systematic search on 13 academic databases was conducted to locate studies evaluating a primary or secondary prevention intervention in Latin America. Studies could use any type of quantitative design to assess outcomes related to youth violence. A search of websites, references and citation searching was also carried out. The quality of each study was assessed. Results: Nine studies were identified. Most documented positive effects of the interventions on the perception of youth violence present in the community/school. Evidence was found of a reduction in homicides and juvenile crimes in three studies, two of which evaluated a community-based intervention. There were mixed results for the self-report of participation on violent acts. The majority of the studies lacked of a rigorous design. Conclusions: Most of the interventions had some promising results, including the reduction of homicides within communities. Community-based programmes were the most consistent regarding an effectiveness to prevent violence. However, the evidence for Latin America is still scarce and relies on non-rigorously designed studies

Search for new phenomena in final states with an energetic jet and large missing transverse momentum in pp collisions at √ s = 8 TeV with the ATLAS detector

Results of a search for new phenomena in final states with an energetic jet and large missing transverse momentum are reported. The search uses 20.3 fb−1 of √ s = 8 TeV data collected in 2012 with the ATLAS detector at the LHC. Events are required to have at least one jet with pT > 120 GeV and no leptons. Nine signal regions are considered with increasing missing transverse momentum requirements between Emiss T > 150 GeV and Emiss T > 700 GeV. Good agreement is observed between the number of events in data and Standard Model expectations. The results are translated into exclusion limits on models with either large extra spatial dimensions, pair production of weakly interacting dark matter candidates, or production of very light gravitinos in a gauge-mediated supersymmetric model. In addition, limits on the production of an invisibly decaying Higgs-like boson leading to similar topologies in the final state are presente

Identification and Characterization of Inhibitors of Human Apurinic/apyrimidinic Endonuclease APE1

APE1 is the major nuclease for excising abasic (AP) sites and particular 3′-obstructive termini from DNA, and is an integral participant in the base excision repair (BER) pathway. BER capacity plays a prominent role in dictating responsiveness to agents that generate oxidative or alkylation DNA damage, as well as certain chain-terminating nucleoside analogs and 5-fluorouracil. We describe within the development of a robust, 1536-well automated screening assay that employs a deoxyoligonucleotide substrate operating in the red-shifted fluorescence spectral region to identify APE1 endonuclease inhibitors. This AP site incision assay was used in a titration-based high-throughput screen of the Library of Pharmacologically Active Compounds (LOPAC1280), a collection of well-characterized, drug-like molecules representing all major target classes. Prioritized hits were authenticated and characterized via two high-throughput screening assays – a Thiazole Orange fluorophore-DNA displacement test and an E. coli endonuclease IV counterscreen – and a conventional, gel-based radiotracer incision assay. The top, validated compounds, i.e. 6-hydroxy-DL-DOPA, Reactive Blue 2 and myricetin, were shown to inhibit AP site cleavage activity of whole cell protein extracts from HEK 293T and HeLa cell lines, and to enhance the cytotoxic and genotoxic potency of the alkylating agent methylmethane sulfonate. The studies herein report on the identification of novel, small molecule APE1-targeted bioactive inhibitor probes, which represent initial chemotypes towards the development of potential pharmaceuticals