Browsing by Subject "Bacterial Infections and Mycoses"
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Item A Novel sRNA Member of the Carbon Storage Regulatory System of Escherichia Coli(2002-12-01) Weilbacher, Thomas; Jerry SimeckaWeilbacher, Thomas S., A Novel sRNA Member of the Carbon Storage Regulatory System of Escherichi coli. Master of Science (Microbiology & Immunology), December, 2002, 57 pp., 2 tables, 12 illustrations, bibliography, 44 titles. Small untranslated RNAs (sRNAs) perform a variety of important functions in bacterial systems. The 245 nt sRNA of Escherichia coli K-12, CsrC, was uncovered using a genetic screen for genes that regulate glycogen biosynthesis. CsrC RNA binds multiple copies of CsrA, a protein that post-transcriptionally regulates central carbon flux, biofilm formation, and motility in E. coli. CsrC antagonizes the regulatory effects of CsrA, presumably by sequestering this protein. The discovery of CsrC is intriguing, in that a similar sRNA, CsrB, performs essentially the same function. Both of these sRNAs possess similar imperfect repeat sequences (18 in CsrB, 9 in CsrC), primarily localized in the loops of predicted hairpins, which may serve as CsrA binding elements. Transcription of csrC increases as the culture approaches the stationary phase of growth and is activated by CsrA and the response regulator UvrY. Complementation and in vitro transcription-translation experiments reveal that CsrA effects on csrC are mediated indirectly, through UvrY. Because CsrB and CsrC antagonize the activity of CsrA and are dependent on CsrA for their synthesis, a csrB null mutation causes a modest compensatory increase in CsrC levels and vice versa. An updated model for the signaling circuitry of the Csr system is discussed.Item A Study of the Effectiveness and Tolerability of Weekly Rifapentine/Isoniazid for Three Months Versus Daily Isoniazid for Nine Months for the Treatment of Latent Tuberculosis Infection(2004-11-01) Lemp, Jessie; Patricia Gwirtz; Walter McConathy; Richard EasomLemp, Jessie M. A Study of the Effectiveness and Tolerability of Weekly Rifapentine/Isoniazid for Three Months Versus Daily Isoniazid for Nine Months for the Treatment of Latent Tuberculosis Infection. Master of Science, November, 2004, 107 pp., 4 tables, 4 figures, references, 29 titles. The standard treatment for latent tuberculosis infection, nine months of daily isoniazid, is effective at preventing active tuberculosis; however, its full benefits are limited by non-adherence. A shorter intermittent regimen of rifapentine plus isoniazid once weekly for three months is equally effective as the standard regimen in animal models. This regimen facilitates the use of directly observed therapy, a method that significantly improves adherence. The Center for Disease Control is sponsoring Study 26 to test the effectiveness and tolerability the three-month rifapentine based regimen in latently infected persons with risk factors for progression to active tuberculosis. This thesis will describe the background rationale and methods for the clinical trial, and the internship experience.Item Epidemiologic Assessment of a Targeted Tuberulosis Screening and Treatment Program Based on Geographic and Molecular Clustering(2005-08-01) Moonan, Patrick KevinMoonan, Patrick K., Epidemiologic Assessment of a Targeted Tuberculosis Screening and Treatment Program Based on Geographic and Molecular Clustering. Doctor of Public Health (Disease Prevention and Control), August 2005, 93 pp., 12 tables, 7 illustrations, bibliography, 148 tables. One of the primary goals of the tuberculosis elimination strategy is to interrupt the transmission of mycobacterium tuberculosis (TB). The most effective way to accomplish this goal is to identify and treat individuals who have active tuberculosis. However, even in highly effected tuberculosis control programs, M. tuberculosis continues to be transmitted to others, largely because most transmission occurs before diagnosis and initiation of therapy. Under the current recommendations, testing should be targeted at specific high-risk populations. While a strategy of targeted testing and treatment of persons most likely to develop tuberculosis is attractive, it is uncertain how best to accomplish this goal. This is the first study to assess the use of geographic and molecular surveillance in guiding a targeted tuberculosis screening and treatment of active tuberculosis and latent tuberculosis infection that monitors potential transmission in a defined high risk geographic area. The results of this geographically targeted program demonstrate significant yield for discovering active cases, latent tuberculosis infection, and recent transmission (TST converters). In this setting, geographically targeted screening identified as many as 19.8 tuberculosis cases per 1,000 persons screened and as many as 292.4 latent tuberculosis infections per 1,000 persons screened. Additionally, successful treatment of these individuals reduced the number of both cases and latent infection identified. Over a three-year period the case detection rate, latent infection detection rate, and TST conversion rate was reduced by 335%, 171% and 285% respectively.Item Factors Associated with Multi-Drug Resistance among Patients with Streptoccus pneumoniae Ear Infections(2004-05-01) Mendoza, Belinda A.; Francisco Soto Mas; Chiehwen Ed Hsu; Antonio ReneMendoza, Belinda A., Factors Associated with Multi-Drug Resistance among Patients with Streptoccus pneumoniae Ear Infections. Master of Public Health (Social and Behavioral Sciences), May 2004, 27 pp., 6 tables, 1 figure, references, 9 titles. Clinical trials play an important role in the development of new medical treatments. The purpose of this study is to describe patients participating in a clinical trial and at analyze the socio-demographic characteristics of patients with susceptible and multi-drug resistant Streptococcus pneumoniae ear infections. At the conclusion of this study, a socio-demographic description of clinical trial participants was obtained and the results were slightly younger than patients with susceptible S. pneumoniae ear infections and were more likely to attend day care.Item Next-Generation Sequencing of Culture Negative Bronchoalveolar Lavage Reveals the Presence of Potentially Pathogenic Microorganisms(2015-08-01) Smith, Ashley D.; Michael Allen; Rance E. Berg; Harlan P. JonesPatients undergoing mechanical ventilation are at increased risk for developing nosocomial pneumonia. Traditionally, the diagnosis of pneumonia has relied on the identification of an etiologic agent by the hospital pathology lab via quantitative culture. A problem arises when a patient experiences clinical signs and symptoms of pneumonia, but culturing of bronchoalveolar lavage (BAL) fluid reveals only normal “respiratory tract flora” or results in “no growth”. I hypothesize that culture-negative, presumptive positive BAL is infected with pathogenic bacteria that are not being cultivated on traditional culture media. To investigate this hypothesis, culture-independent techniques were chosen to examine culture-positive and culture-negative BAL. Sanger and Ion Torrent Sequencing were used to verify that molecular techniques are able to identify the same pathogens the hospital lab finds within culture-positive BAL. Ion Torrent sequencing was then used to characterize the microbial community within culture-negative BAL. Cytokine assays were used to determine if culture-positive and culture-negative patients mount a similar immune response. By sequencing colonies picked from the same culture plates used by the hospital lab, I confirmed the presence of the same pathogens identified by the hospital. Ion Torrent sequencing was able to identify hundreds of genera in both the culture-positive and culture-negative BAL samples. However, no difference in 16S copy number was found between the groups. A group of culture-negative BAL with high diversity and similar bacterial communities were observed. These samples clustered together upon principal coordinates analysis, I believe this grouping may represent a core microbiome. Production of pro-inflammatory cytokines were found to be increased in samples that were dominated by a particular pathogen/pathogens as opposed to those that were more diverse and had lower cytokine measurements. This work highlights the advantages associated with using culture-independent techniques for the diagnosis of pneumonia specifically when traditional culturing techniques fail to identify an etiologic agent.Item Positive Regulation of Acetate Metabolism and Motility by the RNA-Binding Protein CsrA in Escherichia coli(2000-08-01) Wei, Bangdong L.; Jerry Simecka; Ming-Chi Wu; Stephen R. GrantWei, Bangdong L., Positive Regulation of Acetate Metabolism and Motility by the RNA-binding Protein CsrA in Escherichia coli. Doctor of Philosophy (Biomedical Sciences), August, 2000, 118 pp., 5 tables, 19 illustrations, bibliography, 175 titles. The carbon storage regulatory (Csr) system consists of a small RNA-binding effector protein, CsrA, and non-coding RNA, CsrB. CsrA acts as a global regulator and modulates specific mRNA stability in Escherichia coli. It regulates central carbon metabolism, physiology, and cell surface properties on a broad scale. In this study, the regulatory roles of csrA in acetate metabolism and motility were examined. The csrA gene was demonstrated to positively regulate acetyl-CoA synthetase and isocitrate lyase, while it did not affect phosphotransacetylase, isocitrate dehydrogenase, or citrate synthase. As a result, growth of csrA rpoS mutant strains was very poor on acetate as a sole carbon source. Surprisingly, growth also was inhibited specifically by the addition of modest amounts of acetate to rich media. Cultures grown in the presence of ≥25 mM acetate consisted substantially of glycogen biosynthesis (glg) mutants, which were no longer inhibited by acetate. Several classes of glg mutations were mapped to known and novel loci. The TCA cycle intermediates or pyruvate, but not glucose, galactose or glycerol, restored growth and prevented the glg mutations in the presence of acetate. Furthermore, amino acid uptake was inhibited by acetate specifically in the csrA rpoS strain. Apparently, central carbon flux imbalance, inhibition of amino acid uptake, and a deficiency in acetate metabolism are combined to cause metabolic stress by depleting the TCA cycle. The csrA gene was essential for motility and flagellum biosynthesis. Further studies elucidated the molecular mechanism by which CsrA positively regulates flagellum synthesis. Purified recombinant CsrA protein, which was isolated as a ribonucleoprotein complex consisting of one single CsrB molecule and ~18 CsrA subunits, directly stimulated the coupled transcription-translation of flhDC::lacZ in S-30 extracts and bound specifically to the 5’ non-coding segment of flhDC mRNA in mobility shift assay. The steady state level of flhDC mRNA was higher and its half-life was ~3-fold greater in a csrA wild type versus a csrA::kanR mutant strain, as shown by RT-PCR. Thus, CsrA is able to stimulate flhDC gene expression by a post-transcriptional mechanism that resembles its function in repression.Item Predictors of Complicated Staphylococcus Aureus Bacteremia: A Retrospective Validation Study(2008-04-01) Krishnamurthy, Pramod; Fischbach, Lori; Cardarelli, Roberto; Coggin, Claudia S.Krishnamurthy, P., Predictors of Complicated Staphylococcus aureus Bacteremia (SAB): A Retrospective Validation Study. Master of Public Health (Epidemiology), April 2008, 57 pp, 9 tables, 1 illustration, bibliography, 39 titles. SAB often has a complicated clinical course and it is important to identify those at risk for complications to guide management. We conducted a validation study of a clinical prediction tool that uses a scoring system to predict the likelihood of developing complicated SAB. Chapter I is a review of background literature and rationale for our study. Chapter II has sections describing the study design, methods, eligibility criteria, statistical analysis and a summary of the results. We observed significantly higher complications among patients with SAB in our validation study. The prediction tool is not a valid predictor of complicated SAB and we recommend better prediction models to accurately predict complications of SAB.Item The Effects of Two Staphylococcal Global Regulators (agr and sar) on Acid Phosphatase production in Staphylococcus aureus(2003-05-01) Agouna-Deciat, Bahrka Olivier; Jerry Simecka; Michael SmithAgouna-Deciat, B. Olivier, The Effect of Two Staphylococcal Global Regulators (agr and sar) on Acid Phosphatase Production in Staphylococcus aureus. Master of Science (Molecular Biology and Immunology), May 2003, 75 pp., 3 tables, 14 illustrations, 16 titles. Staphylococcus aureus produces an extensive number of cell-surface associated proteins, extracellular proteins and enzymes that contribute to its virulence. The key to better preventative or curative approaches resides in identifying and targeting the very genes and their products that play major roles in the survival of the bacteria within the host and the establishment of diseases. Two well known regulatory loci, the accessory gene regulatory (agr) and the staphylococcal accessory regulator (sar), control the expression of most S. aureus genes that encode for its virulence factors. Other virulence gene regulators have recently been isolated. Over 40 proteins and enzymes produced by S. aureus have been identified and several of them have been linked to staphylococcal pathogenesis. In this study, we attempt to determine the role of agr and sar in the regulation of the production of a secreted staphylococcal acid phosphatase (Sap) suspected to contribute to virulence.Item The Impact of the Mycoplasma pulmonis MALP-2 Homologue on Disease Progression(2008-04-01) Spear, Marcia G.; Simecka, Jerry W.; Hodge, Lisa M.; Mathew, Porunelloor A.Spear, Marcia. The Impact of Mycoplasma pulmonis MALP-2 Homologue on Disease Progression. Master of Science (Biomedical Sciences), April 2008. 64 pp., 3 tables, 8 illustrations. Using Mycoplasma pulmonis, this project looked at a possible critical component in mycoplasma disease, the MALP-2 homologue lipoprotein. Studies demonstrated other lipoproteins besides the MALP-2 homologue were critical for in vivo disease progression and in vitro macrophage IL-6, IL-12, and TNF-α cytokine production. This trend was also seen human endothelial kidney (HEK) cells transfected with toll-like receptor 1 (TLR2) and the heterodimer TLR2/6. An increase in IL-8 cytokine production seen in all stimulated HEK cell lines, indicating the lipoproteins involved in cell interactions are TLR2 mediated. This project suggests the M. pulmonis MALP-2 homologue is not the main lipoprotein involved in disease progression and cell interactions, indicating the MALP-2 homologue may not be an ideal target for vaccines or antibiotics.Item The Role of Regulatory T Cells in Mycoplasma Respiratory Infection(2011-05-01) Odeh, Adam N.; Jerry SimeckaThe purpose of these studies was to examine the role of regulatory T cells (Tregs) in mycoplasma respiratory disease. Depletion of Tregs resulted in increased disease severity. Tregdepleted mice lost significantly more weight over the course of the experiment, and displayed a significantly higher incidence of both gross lung lesions and histological lung lesions at day 14 post-infection. Treg depletion resulted in increased cell infiltration into the lungs by day 14 postinfection, and significant increases in the serum levels of mycoplasma-specific antibodies. Treg depletion also led to an increase in the percentage of IL-13+ T cells in the LRNs, meaning that the immune response was skewed towards a Th2 phenotype. There were no differences observed in lung CFU. These data demonstrate that Tregs in mycoplasma respiratory disease play a role in inflammation and disease severity, but have no effect on bacterial clearance. Importantly, depletion of Tregs causes a Th2-directed shift in the immune response. Additional studies demonstrated that Tregs from mycoplasma-infected mice secreted IFN-γ or IL-17. IFN-γ + and IL-17+ Treg populations both preferentially expanded in response to M. pulmonis infection. Depletion of Tregs resulted in decreased secretion of IFN-γ and IL-17 by CD4+ non-Treg cells. Cocultures of Tregs and T helper cells from mycoplasmainfected mice secreted large amounts of IFN-γ and IL-17 when stimulated with mycoplasma membrane antigen. Levels of IL-4, IL-10, and IL-13 did not significantly increase in response to antigen. Together these studies demonstrate that mycoplasma respiratory disease is influenced by Tregs. These data further suggest that mycoplasma-specific Tregs include two unique subpopulations that express either IFN-γ or IL-17, and that these Tregs may promote the secretion of IFN-γ and IL-17 by T helper cells. This may represent a novel mechanism of Tregmediated immune suppression. This knowledge can assist in the development of treatments for mycoplasma respiratory disease and in the development of Treg-mediated therapies for a number of diseases.