The Neisseria Lipooligosaccharide-Specific Alpha-2,3-Sialyltransferase is a Surface-Exposed Outer Membrane Protein

Neisseria gonorrhoeae and Neisseria meningitidis express an similar to43-kDa alpha-2,3-sialyltransferase (Lst) that sialylates the surface lipooligosaccharide (LOS) by using exogenous (in all N. gonorrhoeae strains and some N. meningitidis serogroups) or endogenous (in other N. meningitidis serogroups) sources of 5\u27-cytidinemonophospho-N-acetylneuraminic acid (CMP-NANA). Sialylation of LOS can protect N. gonorrhoeae and N. meningitidis from complement-mediated serum killing and from phagocytic killing by neutrophils. The precise subcellular location of Lst has not been determined. We confirm and extend previous studies by demonstrating that Lst is located in the outer membrane and is surface exposed in both N. gonorrhoeae and N. meningitidis. Western immunoblot analysis of subcellular fractions of N. gonorrhoeae strain F62 and N. meningitidis strain MC58not subset of3 (an acapsulate serogroup B strain) performed with rabbit antiserum raised against recombinant Lst revealed;an similar to43-kDa protein exclusively in outer membrane preparations of both pathogens. Inner membrane, periplasmic, cytoplasmic, and culture supernatant fractions were devoid of Lst, as determined by Western blot analysis. Consistent with this finding, outer membrane fractions of N, gonorrhoeae were significantly enriched for sialyltransferase enzymatic activity. A trace of enzymatic activity was detected in inner membrane fractions, which may have represented Lst in transit to the outer membrane or may have represented inner membrane contamination of outer membrane preparations. Subcellular preparations of an isogenic lst insertion knockout mutant of N. gonorrhoeae F62 (strain ST01) expressed neither a 43-kDa immunoreactive protein nor sialyltransferase activity. Anti-Lst rabbit antiserum bound to whole cells of N. meningitidis MC58not subset of3 and wild-type N. gonorrhoeae F62 but not to the Lst mutant ST01, indicating the surface exposure of the enzyme. Although the anti-Lst antiserum avidly bound enzymatically active, recombinant Lst, it inhibited Lst (sialyltransferase) activity by only about 50% at the highest concentration of antibody, used. On the contrary, anti-Lst antiserum did not inhibit sialylation of whole N. gonorrhoeae cells in the presence of exogenous CMP-NANA, suggesting that the antibody did not bind to or could not access the enzyme active site on the surface of viable Neisseria cells. Taken together, these results indicate that Lst is an outer membrane, surface-exposed glycosyltransferase. To our knowledge, this is the first demonstration of the localization of a bacterial glycosyltransferase to the outer membrane of gram-negative bacteria

4-H club circular ; 025

Prepared by Miss Marion K. White, Extension Specialist in Nutrition, in collaboration with Miss Jane Hinote, Miss Margaret C. Huston and Miss Sara Chiles, Assistant State Club Agents, as a revision of the original material prepared by Miss Marion E. Dunshee, Extension Specialist in Nutrition."April, 1928" (Reprinted, January, 1930

Versatile silicon-waveguide supercontinuum for coherent mid-infrared spectroscopy

Infrared spectroscopy is a powerful tool for basic and applied science. The molecular spectral fingerprints in the 3 um to 20 um region provide a means to uniquely identify molecular structure for fundamental spectroscopy, atmospheric chemistry, trace and hazardous gas detection, and biological microscopy. Driven by such applications, the development of low-noise, coherent laser sources with broad, tunable coverage is a topic of great interest. Laser frequency combs possess a unique combination of precisely defined spectral lines and broad bandwidth that can enable the above-mentioned applications. Here, we leverage robust fabrication and geometrical dispersion engineering of silicon nanophotonic waveguides for coherent frequency comb generation spanning 70 THz in the mid-infrared (2.5 um to 6.2 um). Precise waveguide fabrication provides significant spectral broadening and engineered spectra targeted at specific mid-infrared bands. We use this coherent light source for dual-comb spectroscopy at 5 um.Comment: 26 pages, 5 figure

Differential effects of energy stress on AMPK phosphorylation and apoptosis in experimental brain tumor and normal brain

<p>Abstract</p> <p>Background</p> <p>AMP-activated protein kinase (AMPK) is a known physiological cellular energy sensor and becomes phosphorylated at Thr-172 in response to changes in cellular ATP levels. Activated AMPK acts as either an inducer or suppressor of apoptosis depending on the severity of energy stress and the presence or absence of certain functional tumor suppressor genes.</p> <p>Results</p> <p>Here we show that energy stress differentially affects AMPK phosphorylation and cell-death in brain tumor tissue and in tissue from contra-lateral normal brain. We compared TSC2 deficient CT-2A mouse astrocytoma cells with syngeneic normal astrocytes that were grown under identical condition <it>in vitro</it>. Energy stress induced by glucose withdrawal or addition of 2-deoxyglucose caused more ATP depletion, AMPK phosphorylation and apoptosis in CT-2A cells than in the normal astrocytes. Under normal energy conditions pharmacological stimulation of AMPK caused apoptosis in CT-2A cells but not in astrocytes. TSC2 siRNA treated astrocytes are hypersensitive to apoptosis induced by energy stress compared to control cells. AMPK phosphorylation and apoptosis were also greater in the CT-2A tumor tissue than in the normal brain tissue following implementation of dietary energy restriction. Inefficient mTOR and TSC2 signaling, downstream of AMPK, is responsible for CT-2A cell-death, while functional LKB1 may protect normal brain cells under energy stress.</p> <p>Conclusion</p> <p>Together these data demonstrates that AMPK phosphorylation induces apoptosis in mouse astrocytoma but may protect normal brain cells from apoptosis under similar energy stress condition. Therefore, using activator of AMPK along with glycolysis inhibitor could be a potential therapeutic approach for TSC2 deficient human malignant astrocytoma.</p

RNAi Trigger Delivery into Anopheles gambiae Pupae

Citation: Regna, K., Harrison, R. M., Heyse, S. A., Chiles, T. C., Michel, K., & Muskavitch, M. A. T. (2016). RNAi Trigger Delivery into Anopheles gambiae Pupae. Jove-Journal of Visualized Experiments(109), 9. doi:10.3791/53738RNA interference (RNAi), a naturally occurring phenomenon in eukaryotic organisms, is an extremely valuable tool that can be utilized in the laboratory for functional genomic studies. The ability to knockdown individual genes selectively via this reverse genetic technique has allowed many researchers to rapidly uncover the biological roles of numerous genes within many organisms, by evaluation of loss-of-function phenotypes. In the major human malaria vector Anopheles gambiae, the predominant method used to reduce the function of targeted genes involves injection of double-stranded (dsRNA) into the hemocoel of the adult mosquito. While this method has been successful, gene knockdown in adults excludes the functional assessment of genes that are expressed and potentially play roles during pre-adult stages, as well as genes that are expressed in limited numbers of cells in adult mosquitoes. We describe a method for the injection of Serine Protease Inhibitor 2 (SRPN2) dsRNA during the early pupal stage and validate SRPN2 protein knockdown by observing decreased target protein levels and the formation of melanotic pseudo-tumors in SRPN2 knockdown adult mosquitoes. This evident phenotype has been described previously for adult stage knockdown of SRPN2 function, and we have recapitulated this adult phenotype by SRPN2 knockdown initiated during pupal development. When used in conjunction with a dye-labeled dsRNA solution, this technique enables easy visualization by simple light microscopy of injection quality and distribution of dsRNA in the hemocoel

Resident Gut Microbiota Mediates Exercise Capacity and Tissue Metabolomes in Mice

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The Burden of Cardiovascular Disease from Air Pollution in Rwanda

Background: Rwanda, like many countries in sub-Saharan Africa, is still relatively early in development. Industrialization and urbanization are major drivers of the county’s economic growth. Rwanda is also undergoing an epidemiological transition, from a pattern of morbidity and mortality dominated by infectious diseases to a pattern shaped by non-communicable diseases (NCDs). The rise in NCDs is due, in part, to increasing exposures to environmental hazards. These include emissions from the growing number of motor vehicles and toxic occupational exposures. Cardiovascular disease (CVD) is now an increasingly important cause of death in Rwanda, and ambient air pollution is a CVD risk factor of growing importance. Objectives: To quantify the burden of CVD attributable to air pollution in Rwanda and identify opportunities for prevention and control of air pollution and pollution-related disease. Methods: We relied on the 2019 Global Burden of Disease (GBD) study for information on levels, sources, and trends in household and ambient air pollution and the burden of pollution-related disease in Rwanda. Information on pollution sources was obtained from the Health Effects Institute State of Global Air 2019 report. Findings: An estimated 3,477 deaths (95% Uncertainty Interval [UI]: 2,500–4,600) in Rwanda in 2019 were attributable to air pollution-related CVD. Of these, 689 (UI: 283–1,300) deaths were from ambient air pollution-related CVD, while 2,788 (UI: 1,800–3,800) deaths were from household air pollution-related CVD. Conclusion: Rwanda is experiencing increased rates of disease and premature death from NCDs, including CVD, as the country grows economically. While household air pollution is still the top pollution-related cause of disease and premature death, rising levels of ambient air pollution are an increasingly important CVD risk factor. Recommendation: Actions taken now to curb rising levels of ambient air pollution will improve health, reduce CVD, increase longevity, and produce great economic benefit for Rwanda. The single most effective intervention against air pollution will be a rapid nationwide transition to renewable energy. We recommend additionally that Rwanda prioritize air pollution prevention and control, establish a robust, nationwide air monitoring network, support research on the health effects of air pollutants, and build national research capacity. The allocation of increased resources for rural and urban public health and health care will complement air pollution control measures and further reduce CVD. To incentivize a rapid transition to renewable energy in Rwanda and other nations, we recommend the creation of a new Global Green Development Fund

Book Reviews

Book reviews of: Hattiesburg: An American City in Black and White By William Sturkey. (Cambridge: Harvard University Press, 2019. Acknowledgements, illustrations, map, notes, index. Pp. 442. $29.95 cloth. ISBN 978-0-674-97635-1.) Confederate Generals in the Trans-Mississippi, Volume 3: Essays on America’s Civil War. Edited by Thomas E. Schott and Lawrence Lee Hewitt. (Knoxville: University of Tennessee Press, 2019. Maps, photos, notes, appendix, bibliography, index. Pp. xxiv, 374.$64.95 cloth. ISBN: 978-1-62190-454-0.) Mothers of Massive Resistance: White Women and the Politics of White Supremacy. By Elizabeth Gillespie McRae. (New York: Oxford University Press, 2018. Acknowledgements, Abbreviations, illustrations, notes, index. Pp. xiv, 343. $34.95 hardcover. ISBN: 978-0-19-027171-8.) Poll Power: The Voter Education Project and the Movement for the Ballot in the American South. By Evan Faulkenbury. (Chapel Hill: University of North Carolina Press, 2019. Acknowledgements, illustrations, notes, index. Pp. xi, 200.$90 cloth, $27.95 paper. ISBN: 978-1-4696-5131-6.) Let Us Make Men: The Twentieth Century Black Press and a Manly Vision for Racial Advancement. By D’Weston Haywood. (Chapel Hill: University of North Carolina Press, 2018. Acknowledgements, illustrations, map, notes, index. Pp. xi, 340.$55 cloth, $19.50 paper. ISBN: 978-1-4696-4338-0.) The Man Who Punched Jefferson Davis: The Political Life of Henry S. Foote, Southern Unionist. By Ben Wynne. (Baton Rouge, Louisiana State University Press, 2018. Acknowledgements, illustrations, notes, index. Pp. ix, 323.$47.50 cloth. ISBN: 978-0-8071-6933-9.) Desegregating Dixie: The Catholic Church in the South and Desegregation, 1945-1992. By Mark Newman. (Jackson: University Press of Mississippi, 2018. Acknowledgements, appendices, notes, bibliography, index. Pp. xvii, 455. $90 cloth,$30 paper. ISBN: 978-1-4968-1886-7.) The Loyal Republic: Traitors, Slaves, and the Remaking of Citizenship in Civil War America. By Erik Mathisen. (Chapel Hill: The University of North Carolina Press, 2018. Acknowledgments, illustrations, map, notes, index. Pp. xi, 219. $34.95 cloth. ISBN: 978-1-4696-3632-0.) Aberration of Mind: Suicide and Suffering in the Civil War-Era South. By Diane Miller Sommerville. (Chapel Hill: University of North Carolina Press, 2018. Acknowledgements, notes, bibliography, index. Pp. 448.$105 cloth, $34.95 paper. ISBN: 978-1-4696-4330-4.) Women’s War: Fighting and Surviving the American Civil War. By Stephanie McCurry. (Cambridge, Massachusetts: The Belknap Press of Harvard University Press, 2019. notes, acknowledgements, index. Pp ix, 297.$26.95 hardcover. ISBN: 978-0-674-98797-5.) Lines Were Drawn: Remembering Court-Ordered Integration at a Mississippi High School. Edited By Teena F. Horn, Alan Huffman, and John G. Jones. (Jackson: University Press of Mississippi, 2016. Acknowledgments, illustrations, map, notes, index. Pp. xi, 266. $35 hard back. ISBN: 978-1-62846-231-9.) Industrial Development and Manufacturing in the Antebellum Gulf South: A Reevaluation. By Michael S. Frawley. (Baton Rouge: Louisiana State University Press. 2019. ix, 256 pp. Cloth,$45.00, ISBN 978-0-8071-7068-7.) Integration Now: Alexander v. Holmes and the End of Jim Crow Education. By William P. Hustwit. (Chapel Hill: University of North Carolina Press, 2019. 8 halftones, 1 map, notes, bibl., index. 288 pp. $39.95, hardcover. ISBN: 978-1-4696-4855-2.

Evaluation of the Workplace Environment in the UK, and the Impact on Users’ Levels of Stimulation

The purpose of this study is to evaluate a number of recently completed workplaces in the UK. The first aim is to assess the impact of various aspects of the workplace environment on users’ levels of stimulation. The body of previous research undertaken into the workplace environment, identified the aspects to be investigated. Samples of employees from the sixteen businesses were surveyed to determine their perceptions of the workplaces. The results were entered into a regression analysis, and the most significant predictors of perceived stimulation identified. The data also revealed a dramatic reduction in staff arousal levels from mornings to afternoons. Thus, there is a second aim to determine whether changes to significant aspects of the workplace environment during the day can counteract the reduction in users’ stimulation. Two further workplaces were studied to enable changes to be made over a 12-week period. A sample of employees completed questionnaires, and semi-structured interviews revealed the reasons behind the results. It was found that provision of artwork, personal control of temperature and ventilation and regular breaks were the most significant contributions to increasing stimulation after lunch; while user choice of layout, and design and décor of workspaces and break areas, were the most significant aspects at design stage