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1
Kupferberg, Eric David. "The expertise of germs : practice, language and authority in American bacteriology, 1899-1924." Thesis, Massachusetts Institute of Technology, 2001. http://hdl.handle.net/1721.1/8669.
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Thesis (Ph. D.)--Massachusetts Institute of Technology, Program in Science, Technology and Society, 2001."February 2001."
Includes bibliographical references (v. 2, p. 631-784).
This thesis traces the development of American bacteriology during the first quarter of the twentieth century. While bacteriology experienced a period of rapid growth, an enduring disciplinary anxiety equally characterized the field. In particular, bacteriologists feared increasing specialization and conceptual fragmentation. Leading practitioners repeatedly worried that their science constituted a collection of unrelated techniques, carried out in the service to other practical endeavors without the benefit of an underlying theory or unifying language. I suggest that the sources of bacteriology's rapid professional growth equally accounted for this sense of conceptual impoverishment and disciplinary privation. Typically, bacteriologists focused on what bacteria did rather than what they were in any biological sense. The first three chapters provide a comprehensive survey of the institutional contexts bacteriology (e.g., medical schools, public health laboratories, water sanitation works, dairies, land-grant colleges, and agricultural experiment stations). For the most part, bacteriologists studied bacteria only so far as to isolate, identify and eliminate pathogens. Dairy and soil bacteriologists, however, sought to distinguish productive types of bacteria, and render those forms more active, a direction that led them to consider a range of phenomena and organisms normally occluded by the practices of medical, public health, and sanitary bacteriology.
(cont.) The final three chapters of the dissertation trace the attempts of American bacteriologists to render their science less fragmented and more biological, focusing in particular on the actions of the Society of American Bacteriologists (SAB). Established in 1899, the SAB endeavored to bridge the divergent interests and practices of American bacteriologists. Through its inclusive membership, ecumenical leadership, diverse meeting programs, and society journal, the SAB served as an organizational exploration of those shared aspects of the discipline. Furthermore, the SAB issued a comprehensive chart for the identification of unknown cultures. While never endorsed as its official methods, the chart soon formed the basis of undergraduate and graduate training, while it guided research programs and published papers. In addition, the serial revisions of the chart led bacteriologists to consider many fundamental aspects of bacteria. Lastly, the SAB struggled to reform bacterial systematics. At the time of the SAB's founding, bacteriology languished under a state of taxonomic chaos, with each specialty offering its own system of naming and grouping bacteria. Believing that this linguistic fragmentation precluded the emergence of a unified discipline, the SAB overhauled bacterial systematics, arranging bacteria according to their detailed morphology, physiology, and likely evolutionary histories.
(cont.) While the SAB's taxonomy did not find immediate adherents, it did become authoritative by way of the classroom and laboratory. The SAB issued a new comprehensive determinative guide, the Bergey's Manual of Determinative Bacteriology, which incorporated the SAB's scheme. As the Bergey's Manual became ubiquitous to laboratory practice and course instruction, American bacteriologists unwittingly adopted a broader range of considerations ...
by Eric D. Kupferberg.
Ph.D.
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Sangodeyi, Funke Iyabo. "The Making of the Microbial Body, 1900s-2012." Thesis, Harvard University, 2014. http://dissertations.umi.com/gsas.harvard:11692.
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This dissertation examines how the relationship between microbes and the human body has been reconfigured over the course of the twentieth century and into the first decades of the twenty-first century. It presents a counter-narrative to the ways in which we have tended to view microbe-human relations to make sense of the emergence of twenty-first century microbial selves by focusing on the normal microbiota.History of Science
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Fuqua, Andrew. "Characterization of the Broad-spectrum Inhibitory Capability of Alcaligenes faecalis and A. viscolactis against Potential Pathogenic Microorganisms." Digital Commons @ East Tennessee State University, 2020. https://dc.etsu.edu/honors/546.
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The recent rise of multidrug resistant microorganisms has grown from an isolated concern to a massive public health crisis. It has become imperative that scientists look for new ways to combat this issue. Due to the selective pressures of competition, bacteria and other microbes possess a host of defenses and weapons designed to exploit vulnerabilities in other microorganisms. Consequently, the study of these systems and microbial interactions has much to reveal in the search for novel antimicrobial treatments. Previous research from our laboratory has discovered that both Alcaligenes faecalis and Alcaligenes viscolactis, two rarely studied and generally non-virulent bacteria, exert a microbicidal effect on Candida albicans and Staphylococcus aureus, two pathogenic and frequently drug-resistant organisms. In this study, we confirmed that these effects are via a live-cell, contact-dependent mechanism and showed that both Alcaligenes species inhibit S. aureus at the attachment phase of biofilm growth. Additionally, we found that A. faecalis and A. viscolactis target Gram-positive bacteria outside the genus Staphylococcus and certain Gram-negative species as well as Candida glabrata. This study also provides novel evidence of a putative Type VI Secretion System in both Alcaligenes species, which may explain their antimicrobial phenotype. Despite efforts to identify the genetic elements involved via mutagenesis, the mechanism of these interactions remain elusive due to the difficulty of gene transfer in these organisms. We hope these results will increase current knowledge of Alcaligenes’ capabilities and genetic composition as well as establish the groundwork for future efforts to discover its inhibitory system and mechanisms.
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Liu, Yunhao. "Structural and biochemical analysis of HutD from Pseudomonas fluorescens SBW25 : a thesis submitted in fulfilment of the requirements for the degree of Master of Science in Molecular Biosciences at Massey University, Auckland, New Zealand." Massey University, 2009. http://hdl.handle.net/10179/1074.
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Pseudomonas fluorescens SBW25 is a gram-negative soil bacterium capable of growing on histidine as the sole source of carbon and nitrogen. Expression of histidine utilization (hut) genes is controlled by the HutC repressor with urocanate, the first intermediate of the histidine degradation pathway, as the direct inducer. Recent genome sequencing of P. fluorescens SBW25 revealed the presence of hutD in the hut locus, which encodes a highly conserved hypothetical protein. Previous genetic analysis showed that hutD is involved in hut regulation, in such a way that it prevents overproduction of the hut enzymes. Deletion of hutD resulted in a slow growth phenotype in minimal medium with histidine as the sole carbon and nitrogen source. While the genetic evidence supporting a role of hutD in hut regulation is strong, nothing is known of the mechanism of HutD action. Here I have cloned and expressed the P. fluorescens SBW25 hutD in E. coli. Purified HutD was subjected to chemical and structural analysis. Analytic size-exclusion chromatography indicated that HutD forms a dimer in the elution buffer. The crystal structure of HutD was solved at 1.80 Å (R = 19.3% and Rfree = 22.3%) by using molecular replacement based on HutD from P. aeruginosa PAO1. P. fluorescens SBW25 HutD has two molecules in an asymmetric unit and each monomer consists of one subdomain and two ß-barrel domains. Comparative structural analysis revealed a conserved binding pocket. The interaction of formate with a highly conserved residue Arg61 via salt-bridges in the pocket suggests HutD binds to small molecules with carboxylic group(s) such as histidine, urocanate or formyl-glutamate. The hypothesis that HutD functions via binding to urocanate, the hut inducer, was tested. Experiments using a thermal shift assay and isothermal titration calorimetry (ITC) analysis suggested that HutD binds to urocanate but not to histidine. However, the signal of HutD-urocanate binding was very weak and detected only at high urocanate concentration (53.23 mM), which is not physiologically relevant. The current data thus does not support the hypothesis of HutD-urocanate binding in vivo. Although the HutD-urocanate binding was not confirmed, this work has laid a solid foundation for further testing of the many alternative hypotheses regarding HutD function.
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Brunson, Debra Nickole. "Loss of outer membrane porins in clonally related clinical isolates of Klebsiella pneumoniae modifies the bacteria; resulting in altered resistance to phagocytosis by macrophages." UNF Digital Commons, 2017. http://digitalcommons.unf.edu/etd/724.
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Klebsiella pneumoniae is an opportunistic pathogen responsible for lobar pneumoniae, liver abscess, and septicemia. Clinical isolates are found to be extended spectrum beta lactamase positive with differential expression of the two classical porins, OmpK35 and OmpK36. Porin loss is associated with increased minimum inhibitory concentrations of beta lactam, cephalosporin, and carbapenem antibiotics that target the peptidoglycan. However, little is known about how porin loss affects other aspects of the cell envelope. The focus of this study was to characterize clinical isolates exhibiting differential porin expression and determine if the cumulative changes altered the resistance to phagocytosis by macrophages. The results support the hypothesis that porin loss significantly impacts the overall cell envelope composition, which in turn alters interactions with macrophages.
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Baxter, Brooke E. "Cloning and Expression of C-terminal Fragment of TonB from Rhizobium leguminosarum ATCC 14479." Digital Commons @ East Tennessee State University, 2017. https://dc.etsu.edu/honors/410.
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The TonB-ExbB-ExbD complex is essential for the siderophore mediated acquisition of iron by Gram negative bacteria. The system provides energy from the proton motive force to the outer membrane in order for the iron siderophore complex to enter into the cell. The main protein involved in energy transduction, TonB, has been extensively studied in the species Escherichia Coli. It has been determined that the protein consists of 239 amino acids. In comparison, however, the TonB of Rhizobium leguminosarum consists of 457 amino acids with the same conserved regions. What is in question, therefore, is how the additional amino acids effect the structure of the C-terminal region of the protein and how such information can give insight into the way in which the proton motive force functions to provide energy to the outer membrane receptor. The protein regions of R. leguminosarum TonB chosen for study were 120 and 248 amino acids from the C-terminal end. Genomic DNA was isolated, primers were designed for each fragment, and polymerase chain reactions were performed. After appropriate restriction enzyme digestion, each DNA fragment was ligated into the plasmid pET-17b and then transformed into Escherichia Coli BL21 (DE3). Successful transformation of the 120 amino acid fragment was followed by expression via IPTG induction & extraction of protein. Afterwards, a T7-tag affinity column was attempted to collect the protein for analysis; however, a sufficient amount of protein was not eluted. The procedure will be repeated for obtaining sufficient protein for crystallization or NMR spectrometric analysis.
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Garcia-Moreno, Pamela K. "Mycobacterium tuberculosis inhibitors: action and resistance." FIU Digital Commons, 2018. https://digitalcommons.fiu.edu/etd/3893.
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Tuberculosis, an infectious disease caused by Mycobacterium tuberculosis, has been a global health problem for years. The emergence of drug resistance in this organism generates the necessity of exploring novel targets and developing new drugs. Topoisomerases are enzymes found in all kingdoms of life responsible for overcoming the topological barriers encountered during essential cellular processes. The genomes of mycobacteria encode only one type IA topoisomerase (MtopI), which has been validated as a novel TB drug target. The goal of this study is to obtain new information on the mechanism and resistance of endogenous and synthetic inhibitors of MtopI.
Rv1495 is a M. tuberculosis toxin that belongs to the MazEF family (MazE is the antitoxin and MazF is the toxin), with endoribonuclease activity. Rv1495 (MazF homolog in M. tuberculosis) toxin has been shown to interact directly with the C-terminal domain of MtopI for mutual inhibition. In this study the interaction of Rv1495 with the positively charged C-terminal tail in Mtop I is reported. This new information is useful for rational design and discovery of antibiotics for mycobacteria.
Ethacridine, an FDA approved drug has shown activity against MtopI. In this project we studied the mechanisms of resistance associated with this drug as well the use of Ethacridine in combination with Moxifloxacin, to potentiate the bactericidal effect of this current second line drug for TB treatment. Results from sequencing of the genomic DNA isolated from the resistant mutants suggested the involvement of the Holliday-junction Ruv resolvase. Further studies showed that co-treatment with Ethacridine can enhance the moxifloxacin-mediated killing of M. smegmatis.
FP-11g, a novel fluoroquinophenoxazine inhibitor of bacterial topoisomerase I, has shown promising activity against M, tuberculosis. We explored the bactericidal activity and resistance mechanisms associated to FP-11g using M. smegmatis as model organism. Additionally, the inhibitory effect of FP-11g was also evaluated on M. abscessus. FP-11g at concentration 4X MIC showed complete bactericidal activity against M. smegmatis after 24 hours. Inhibitory activity against M. abscessus was also observed. Results from sequencing of the genomic DNA isolated from the M. smegmatis resistant mutants revealed mutations in genes associated with general drug resistance.
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Radomski, Nicolas. "Sources des mycobactéries non-tuberculeuses dans les bassins versants." Phd thesis, Université Paris-Est, 2011. http://pastel.archives-ouvertes.fr/pastel-00669399.
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L'eau et le sol sont considérés comme des sources potentielles de mycobactéries non-tuberculeuses (MNT). Parmi les infections humaines causées par les MNT d'origine environnementale, les infections pulmonaires et cutanées sont souvent décrites. Le manque de connaissances sur leur cycle de vie dans l'environnement requiert des outils analytiques, qui ne sont actuellement pas adaptés à ce type d'échantillons. Cette thèse vise donc premièrement à proposer des méthodes de quantification en bactériologie et en biologie moléculaire dans le but de déterminer les sources des MNT dans les bassins versants. Ainsi, la comparaison des méthodes d'isolement de MNT a montré que le traitement au chlorure de cetylpyridininium de l'eau suivi d'une culture en milieu riche supplémenté par un mélange d'antibiotiques (polymyxine B, amphotéricine, acide nalidixique, triméthoprime, carboxy-pénicilline) limitait la croissance des microorganismes interférents et éliminait moins de MNT que les autres méthodes comparées (Radomski et al. 2010, doi: 10.1128/AEM.00942-10). Bien que des espèces de MNT potentiellement pathogènes aient été isolées de l'eau de surface de la Seine en utilisant ces outils bactériologiques, la quantification des MNT ne s'est pas avérée reproductible. En conséquence, une méthode de quantification par polymérisation en chaîne en temps réel (qPCR) a été développée pour énumérer le genre Mycobacterium dans l'eau (Radomski et al. 2010, doi: 10.1128/AEM.02659-09). La nouvelle méthode développée, ciblant l'ARNr 16S, était plus spécifique que les autres méthodes qPCR publiées, ciblant un autre locus de l'ARNr 16S et le gène hsp65 (respectivement 100 % versus 44 % et 91 %). La comparaison des méthodes d'extraction d'ADN mycobactérien a montré que la lyse enzymatique combinée au bromure d'hexadécyltriméthylammonium était la procédure la plus efficace pour énumérer par qPCR les MNT dans des échantillons environnementaux. Ainsi, ces méthodes d'extraction d'ADN et de qPCR ont été utilisées pour étudier des sources de MNT dans des bassins versants. Dans un second temps, nous avons étudié trois sources potentielles de MNT : une ponctuelle et deux diffuses. Plus précisément, une station d'épuration (STEP) a été choisie comme source ponctuelle de MNT et a été étudiée en temps sec en fonction d'indicateurs de contamination fécale et des paramètres globaux habituellement contrôlés. Les MNT ont atteint 5,52×105±3,97×105 copies/L dans l'eau en entrée de STEP (84 % d'échantillons positifs), n'ont pas été détectées dans l'eau en sortie de STEP après décantation physico-chimique et biofiltration et ont été estimées à 1,04×106 ±1,75×106 copies/g dans les boues de STEP (50 % d'échantillons positifs). La plupart des MNT (98±2 %, correspondant à 2,45±0,78 log10) ont été éliminées par décantation physico-chimique et les MNT restantes (0,74×104 ±1,40×104 copies/L) ont été éliminées par biofiltration (53 % d'échantillons positifs). Ces résultats ont montré également que Mycobacterium, Escherichia coli et les entérocoques intestinaux possèdent des comportements significativement différents conduisant respectivement à trois modèles : hydrophobe, hydrophile et intermédiaire. Concernant les sources diffuses, la densité de MNT a été mesurée dans divers sols ruraux et urbains qui ont été caractérisés par différents paramètres physico-chimiques. Les densités de MNT les plus importantes ont été mesurées dans des sols de forêts tourbeuses (9,27×104±5,00×104 copies/g sec) et dans des sols faiblement urbanisés proches de marécages côtiers (1,71×106±2,85×106 copies/g sec) alors qu'aucune MNT n'a été détectée dans les autres types de sols étudiés. De plus, la densité de MNT a été significativement associée à des sols proches de zones acides et des teneurs fortes des sols en eau, matière organique et fer. Ces résultats suggéreraient que les MNT sont dépendantes de leur production intra et extracellulaire de chélateurs de fer et indiqueraient que les zones faiblement urbanisées pourraient être impactées par la proximité de marais acides. Afin d'étudier une autre source diffuse, les MNT et d'autres paramètres ont été mesurés lors d'événements pluvieux dans l'eau de surface de la Marne et de ses principaux affluents. Les densités de MNT ont été estimées à 2,16×105±2,36×105 copies/L dans environ 20 % des échantillons d'eau collectés, et elles ne différaient pas entre les zones péri-urbaines et rurales échantillonnées. Nos résultats ont montré que la pluviométrie et la durée de l'évènement expliquaient la diminution du nombre de MNT détectées dans l'eau de surface au cours de l'événement pluvieux de faible intensité (6,6 mm/h de pluviométrie cumulées en 5,5 h). Ces résultats ont souligné que certains affluents de la Marne pouvaient apporter des MNT en temps sec, mais qu'au cours de l'évènement pluvieux suivi les densités de MNT diminuaient.En guise d'amélioration à ces études appliquées, des réflexions sur les défis relatifs à la surveillance des microorganismes pathogènes dans l'environnement ont été explorées. En nous focalisant sur la MNT la plus pathogène, M. avium, nous avons discuté des défis de la détection et de l'énumération et proposé un guide d'adaptation des méthodes médicales aux échantillons environnementaux (Radomski et al. 2011, ed. A. Méndez-Vilas, Vol. 2). Ce guide se présente sous la forme d'un arbre de décision permettant de choisir les outils analytiques les plus appropriés pour surveiller les microorganismes pathogènes dans l'environnement. De plus, une stratégie in silico de comparaison de génomes bactériens totalement séquencés a été développée dans le but de décrire des nouvelles cibles de détection. L'analyse in silico des génomes totalement séquencés a permis de détecter 11 protéines présentant entre 80 % et 100 % de similarité dans les génomes mycobactériens et moins de 50 % de similarité dans les génomes non-mycobactériens des genres Corynebacterium, Nocardia et Rhodococcus. Sur la base d'alignements des séquences d'ADN de ces cibles potentielles, il a été possible de dessiner des amorces PCR et une sonde pour détecter le gène codant la sous-unité C de la synthase de l'adénosine triphosphate qui semble exclusivement conservée dans le génome mycobactérien. Le développement d'outils analytiques, en particulier la qPCR, a permis de montrer qu'une STEP éliminait efficacement les MNT et que le traitement des eaux usées est nécessaire pour préserver l'eau de surface de cette source ponctuelle de MNT. Il a été mis en évidence que les événements pluvieux diminuent la densité de MNT dans l'eau de surface et que les sols acides sont des sources naturelles majeures de MNT qui pourraient impacter des zones faiblement urbanisées en temps de pluie via le ruissellement. Concernant les réflexions sur la surveillance des microorganismes pathogènes dans l'environnement, l'arbre de décision des outils analytiques appropriés et la nouvelle stratégie in silico de détection de cibles moléculaires pourraient être appliqués pour l'étude d'autres microorganismes de l'environnement
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Pullen, Sheryl L. "In vitro activity of four fluoroquinolones on selected bacteria." Scholarly Commons, 1995. https://scholarlycommons.pacific.edu/uop_etds/2285.
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In 1990-1991, in a national surveillance study, and in 1991-1992, in a followup study, both by Thornsberry et al. (1993), ciprofloxacin data from various geographical and demographical institutions were collected. Several species of bacteria have shown resistance to ciprofloxacin and norfloxacin, but the degree of resistance to these drugs has not been reported for the Stockton area. To determine the extent of this resistance, Dameron Hospital antibiograms generated from 1990 to 1994 were reviewed and compared. Results of the comparison show that susceptibility among the Gram-negative isolates, with the exception of Providencia stuartii, Acinetobacter lwoffi, and to a lesser extent Aeromonas hydrophila, has changed very little. Consistent with the national surveys, resistance of Pseudomonas aemginosa has not changed appreciably during the five-year period.
Among the Gram-positive isolates that were tested against both ciprofloxacin for a five-year period (1990-1994) and norfloxacin for a three-year period (1992"' 1994), increased resistance was seen among strains of Staphylococcus aureus, S. epidermidis, S. haemolyticus, and Enterococcus jaecalis, but not among strains of Staphylococcus saprophyticus, Streptococcus pyogenes, and S. agalactiae. To determine whether resistance to one fluoroquinolone occurs also to other fluoroquinolones, several isolates of Gram-positive cocci and P. aeruginosa from the Gram-negative bacilli that showed resistance to either ciprofloxacin, norfloxacin, or both were selected from Dameron Hospital isolates and tested by the disk diffusion technique against ciprofloxacin, norfloxacin, ofloxacin, and lomefloxacin. The results indicate that differences do exist among these selected strains. Comparison of the invitro effectiveness of the various quinolones confirms that methicillin-resistant staphylococci (S. aureus, S. epidermidis, and S. haemolyticus) exhibit a higher degree of resistance to the four fluoroquinolones compared with the methicillin-susceptible strains of the same species. Resistance of the enterococci (Enterococcus jaecalis and E. jaecium) is also high. Generally, when the four fluoroquinolones were compared with each other, ofloxacin seemed to have better in vitro activity.
Resistance to the quinolones consists of two proposed mechanisms: ( 1) mutation of one or both of the structural genes of the A and B subunits of DNA gyrase and (2) decreased drug accumulation due either to lower uptake by the cell or enhanced effiux out of the cell. These mechanisms of resistance are reviewed.
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Presswood, Rachel Elizabeth. "Isolation of a Siderophore Produced by Methicillin-Resistant Staphylococcus aureus Strain H372." Digital Commons @ East Tennessee State University, 2010. https://dc.etsu.edu/etd/1728.
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Iron is necessary for many cellular processes such as the electron transport chain and gene regulation. However, most iron on earth is found in insoluble iron-hydroxide complexes. In addition, iron is tightly sequestered in the human body by proteins such as transferrin, making it unavailable for pathogens. In order to overcome these limitations bacteria have evolved siderophores. Siderophores are low molecular weight compounds that bind ferric iron with a high affinity. Staphylococcus aureus is an important human pathogen that is known to produce at least four siderophores, and these siderophores contribute to its virulence. S. aureus strain H372 was found to produce a siderophore that was a carboxylate type, hydrophilic, and contained ornithine. These properties were similar to the known siderophore staphyloferrin A. However, the probable molecular weight was 658, which is different from known staphylococcal siderophores.
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Barber, Thomas S. "Discovery and Characterization of an Antibiotic from the Soil Bacterium Bacillus sp." Digital Commons @ East Tennessee State University, 2010. https://dc.etsu.edu/etd/1777.
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Many important antibiotics have become nearly obsolete due to the rise of antibiotic resistant pathogens. Rhodococcus, an actinomycete related to the prolific antibiotic producing genus Streptomyces, harbors over 30 genes for secondary metabolism that could be involved in antibiotic production. Several antibiotics have already been reported for Rhodococcus, suggesting the genus may be a good source for new inhibitory compounds. Fifty four soil bacteria were isolated using enrichment culture techniques (including 37 Rhodococcus) and screened for antibiotic producers. BTHX2, a species of Bacillus was found to have activity against Micrococcus luteus and Rhodococcus erythropolis. BTHX2 has a 16S rDNA sequence 97% homologous to Bacillus licheniformis, and may be a new strain of B. licheniformis. The inhibitory substance produced by BTHX2 was and found to have a spectrum of activity against a broad Gram-positive bacteria and some fungus, and may have cytolytic activity.
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Cooke, Jennifer K. "Molecular Mechanism of Ferricsiderophore Transport via the Outer Membrane Receptor FhuA in Escherichia coli." Digital Commons @ East Tennessee State University, 2009. https://dc.etsu.edu/etd/1792.
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Iron is essential for life and growth in most organisms. Although it is abundant, iron exists mostly as insoluble iron-oxyhydroxide. Bacteria secrete siderophores to chelate iron and transport it into the cell via specific outer membrane receptors. The FhuA receptor protein transports ferrichrome, a siderophore produced by Ustilago sphaerogena. We determined the binding affinity of variants from the conserved 'lock region' of FhuA and also created and characterized variants of the highly conserved R452 to determine its role in ferrichrome transport. We hypothesize that during transport the plug domain of FhuA does not leave the barrel; rather it undergoes a conformational change to form a channel. We mutated selected amino acids to cysteine to form disulfide bonds to tether the plug, preventing its displacement or unfolding during transport. The tetra-cysteine mutant 72/615/109/356C was able to bind and transport radiolabeled ferrichrome. One double-cysteine mutant, 104/149C, was purified for crystallization.
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Pratt, Melanie Anne. "Iron Acquisition in Rhodococcus erythropolis Strain IGTS8: Characterization of a Mutant Strain that Over Produces Siderophore." Digital Commons @ East Tennessee State University, 2008. https://dc.etsu.edu/etd/2013.
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Iron is an essential nutrient for most bacteria because enzymes like nitrate reductase and cytochromes use it as a cofactor. However, in most aerobic, neutral pH environments, iron is essentially insoluble and not easily available for bacteria to use. Many bacteria respond to this problem by releasing small organic compounds called siderophores that bind and effectively solubilize iron so that it can be transported into the cell for growth. The focus of this study was to learn more about the iron acquisition and especially the transport of iron by the soil bacterium Rhodococcus erythropolis. To fulfill this aim, mutant strains of the bacteria were screened for those that overproduce siderophore. Often, a bacterium will over produce siderophore to compensate for a defect in transport. One such mutant, R187-12, was further analyzed by cloning the region of the chromosome containing the defective gene responsible for over production of siderophore into a plasmid vector. The DNA sequence of this region was determined and analyzed for the presence of similar genes encoding transport proteins.
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Reynolds, Susan D. "The Cost of Mupirocin Resistance in Staphylococcus." Digital Commons @ East Tennessee State University, 2006. https://dc.etsu.edu/etd/2192.
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Control of antibiotic resistance in bacteria is based on the concept that resistance incurs a fitness cost in non-selective conditions. Fitness costs were assessed for low- and high-level mupirocin resistance in locally-derived Staphylococcus aureus and S. epidermidis. Costs of resistance were assessed in pure cultures by comparing growth curve characteristics and in mixed culture as the proportion of resistant cells surviving. Costs were not present in comparisons of growth rates among groups of naturally-occurring isolates from the different resistance categories. However, in S. aureus, growth rates within resistance categories differed by approximately 30 – 90%. Among near-isogenic pairs of strains, fitness costs ≥10% were present in three of eleven pairs under pure culture and in six of eleven pairs under competition in mixed culture. Differences in intrinsic growth rates could easily mask fitness costs of the magnitudes observed. Thus, clinical outcomes also depend on whether there is a mixed infection and if so, on the growth rates of strains present.
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Wright, William H. IV. "Isolation and Identification of the Siderophore "Vicibactin" Produced by Rhizobium leguminosarum ATCC 14479." Digital Commons @ East Tennessee State University, 2010. https://dc.etsu.edu/etd/1690.
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Siderophores are small, iron chelating molecules produced by many bacteria to help meet the iron requirements of the cell. Multiple metabolic functions require iron as it serves as a cofactor in many enzymes and cellular processes. However, in the presence of oxygen and at physiologic pH, iron forms insoluble ferric complexes that cause the nutrient to be unavailable to bacterial cells. Siderophores alleviate this limitation by chelating the ferric iron, rendering it soluble and available for uptake. One group of microorganisms known for their ability to produce siderophores is the rhizobia. These bacteria are characterized both by their formation of symbiotic relationships with leguminous plants and their ability to fix atmospheric nitrogen. Rhizobium leguminosarum ATCC 14479, which infects the red clover Trifolium pratense, was found to produce a trihydroxamate siderophore. Purification and chemical characterization identified this siderophore as Vicibactin that has been found to be produced by other rhizobial strains.
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Chinnam, Naga babu. "Inhibition of Escherichia coli ATP Synthase Using Bioflavonoids." Digital Commons @ East Tennessee State University, 2011. https://dc.etsu.edu/etd/1298.
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ATP synthase is the fundamental means of the cellular energy production in all organisms. Malfunction of ATP synthase is associated with multiple disease conditions. This enzyme is not only implicated in disease conditions but also likely to contribute in new therapies for multiple diseases by being a molecular target for several inhibitors. Bioflavonoids are a class of plant secondary metabolites known to exhibit antioxidants, chemopreventive, and chemotherapeutic properties. Their actual mode of action is not clear; however, some bioflavonoid are known to block the action of enzymes and other substances that promote the growth of cancer cells by binding to the multiple molecular targets in the body including ATP synthase. The most common dietary polyphenol resveratrol was shown to induce apoptosis via mitochondrial pathways and has chemopreventive properties against prostate cancer. Here we report the general inhibitory effects of dietary bioflavonoids on ATP synthase enzyme and intact E. coli cells.
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Barnard, Danielle. "Conjugative Transfer Pathways of High-Level Mupirocin Resistance and Conjugative Transfer Genes in Staphylococcus." Digital Commons @ East Tennessee State University, 2006. https://dc.etsu.edu/etd/2188.
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To combat widespread infections caused by Staphylococcus aureus, mupirocin was introduced at the Veterans Affairs Medical Center, Mountain Home, Tennessee. Soon after introduction, high-level mupirocin-resistance emerged. The rapid emergence was hypothesized to be due to conjugative transfer of the mupA resistance gene from S. epidermidis to S. aureus. Results have shown that transfer of high-level mupirocin-resistance from S. aureus donors commonly occurs. However, transfer from naturally-occurring S. epidermidis donors was not attainable. Staphylococcus epidermidis transconjugants, however, were capable of serving as donors. Further examination of non-transmissibility included PCR analysis of conjugative transfer genes (tra genes) in capable and non-capable donors. Results confirmed that capable donors possess full-length copies of selected transfer genes. Non-capable donors varied in the presence/absence of full-length copies of transfer genes, but none had all three genes. The genetic differences among non-capable donors suggest that non-transmissibility has arisen independently in different strains via gene deletions and recombinations.
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Borisova, Ralitsa Bogomilova. "Isolation of a Rhodococcus Soil Bacterium that Produces a Strong Antibacterial Compound." Digital Commons @ East Tennessee State University, 2011. https://dc.etsu.edu/etd/1388.
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Rhodococci are notable for their ability to degrade a variety of natural and xenobiotic compounds. Recently, interest in Rhodococcus has increased due to the discovery of a large number of genes for secondary metabolism. Only a few secondary metabolites have been characterized from the rhodococci (including 3 recently described antibiotics). Twenty-four new Rhodococcus strains were isolated from soils in East Tennessee using acetonitrile enrichment culturing and identified using 16S rRNA analysis. Forty-seven Rhodococcus strains were screened for antibiotic production using a growth inhibition assay. One strain, MTM3W5.2, had 90% similarity to the Rhodococcus opacus 16S rRNA gene sequence and produced a large zone of inhibition against R. erythropolis and a large number of closely related species. The antimicrobial compound produced by MTM3W5.2 had a large MW of 911.5452 Da and acts much like a bacteriocin but no amino acids were detected in this molecule based on TLC analysis.
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Brudecki, Laura Elaine. "Modulation of Alpha-Subunit VISIT-DG Sequence Residues Ser-347, Gly-351 and Thr-349 in the Catalytic Sites of Escherichia coli ATP Synthase." Digital Commons @ East Tennessee State University, 2010. https://dc.etsu.edu/etd/1773.
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Binding of inorganic phosphate (Pi) in ATP synthase catalytic sites is a crucial step for the synthesis of adenosine-5'-triphosphate (ATP). ATP is the fundamental means of cellular energy in almost every organism, and in order to gain insight into the regulation of ATP catalysis, critical amino acid residues responsible for binding Pi must be identified. Here, we investigate the role of highly conserved α-subunit VISIT-DG sequence residues αSer-347, αGly-351, and αThr-349 in Pi binding. Mutations αS347A/Q, αG351Q, αT349A/D/R, βR182A, and αT349R/βR182A were generated via site directed mutagenesis. Results from biochemical assays showed that αSer-347 is required for transition state stabilization and Pi binding whereas αGly-351 is only indirectly involved in Pi binding and most likely maintains structural integrity of the catalytic site. Results from preliminary experiments on αThr-349 mutants suggest that the residue may be involved in Pi binding; however, further investigation is required to fully test this hypothesis.
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Frost, Hannah. "Characterisation of the Group A Streptococcus M and M-like proteins as potential vaccine antigens." Doctoral thesis, Universite Libre de Bruxelles, 2020. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/305271.
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Group A Streptococcus (GAS) is a human specific bacterial pathogen which causes a variety of diseases and is responsible for significant mortality worldwide. GAS are known to interact with the immune system, notably by binding host serum proteins to the bacterial surface.Many of these binding functions are attributed to the GAS M protein, the archetypal GAS virulence factor, the substrate for GAS typing and the leading GAS vaccine candidate. The vast diversity of GAS strains has however made vaccine development challenging. We investigated the potential for cross-protective immunity within closely related strains in a clinical setting in Fiji. This study has shown that immunity to GAS skin infection was broader than previously believed and included some level of cross-opsonisation betweenGAS strains. The level of such cross-opsonisation was, however, variable among GAS lineages. We have also shown that the immunity to conserved M antigens was quite variable. These results inspired us to investigate other suitable vaccine antigens. We hypothesised that in cases where the M protein was less immunodominant, perhaps closely related M-like proteins played an important role in immunity. We therefore began global characterisation ofthe two major M-like proteins, called Mrp and Enn. A representative worldwide genomic collection including more than 2000 isolates originating from multiple continents and various clinical manifestations has been established collaboratively. We focused our analyses on theMga regulon which encodes M and M-like proteins. We found that mrp and enn genes were present in 85% of genomes suggestive of their importance to GAS survival and spread. We developed molecular definitions of the different genes families, clarifying nomenclature forthe worldwide reference laboratory at the American CDC. We established and validated an updated, more specific typing protocol for GAS which will reduce future misclassifications. We have also analysed the genetic linkage between M and M-like protein alleles and developed clusters of closely related protein sequences. By characterising this complex and variable genetic region, we provide a framework for future functional investigations. Finally, we began functional characterisation of Enn proteins by investigating the differential capacity of Enn proteins to bind to C4BP, an inhibitor of complement. Altogether our results suggest M-like proteins play an important role in GAS virulence and should not be neglected. These data support further functional analyses to better understand the contribution of M-like proteins to GAS infection.Doctorat en Sciences biomédicales et pharmaceutiques (Médecine)
info:eu-repo/semantics/nonPublished
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Dill, Brian D. "Identification of Chlamydial Iron-Responsive Proteins during Intracellular Growth." Digital Commons @ East Tennessee State University, 2008. https://dc.etsu.edu/etd/1955.
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Chlamydia trachomatis is an obligate intracellular bacterium and the most prevalent cause of bacterial sexually transmitted disease. Genital chlamydial infections, marked by chronic, intense inflammation, can lead to genital tissue scarring and infertility and is a contributing factor to development of pelvic inflammatory disease and ectopic pregnancy. Iron is required as a cofactor for numerous highly conserved pathways, and nearly all studied organisms rely on iron for growth. In response to iron restriction, the chlamydial developmental cycle arrests at the intracellular reticulate body stage, resulting in a phenomenon termed persistence. Persistence likely plays a role in chlamydial pathogenesis through the expression of virulence factors and antigens in addition to sustaining chronic infection; however, little is known concerning how chlamydiae respond to iron limitation at the molecular level, and no systems for iron acquisition have been identified in Chlamydia. This dissertation presents an investigation into the chlamydial response to iron restriction. Chlamydial heat shock protein 60 (cHsp60) has been implicated in development of the more severe disease sequelae and has been found to increase in expression following iron restriction; however, three cHsp60 homologues were identified following the sequencing of the chlamydial genome. Here, iron restriction is shown to increase expression of cHsp60-2 but not the two other homologs, cHsp60-1 or -3. Next, in order to investigate an alternate model for restricting iron availability to chlamydiae, a cell line with inducible expression of recombinant ferroportin, a eukaryotic iron efflux protein, was examined. Lastly, 10 chlamydial proteins differentially expressed during growth in iron-restricted host cells were identified by proteomic analysis of radiolabeled proteins followed by mass spectrometry analysis; transcripts encoding 5 iron responsive proteins were examined across a timecourse of infection and revealed increased transcript levels at 18 and/or 24 hours post infection. Together, these studies have examined the molecular response of chlamydiae to reduced iron availability and have underlined the importance for pathways involved in protection against oxidative damage and adaptation to stress.
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Jankovic, Dragana. "Direct selection and phage display of the Lactobacillus rhamnosus HN001 secretome : a thesis presented to Massey University in partial fulfillment of the requirements for the degree of Doctor of Philosophy." Massey University, 2008. http://hdl.handle.net/10179/869.
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Bacteria communicate with their hosts in part via surface, secreted and transmembrane proteins (collectively the secretome) resulting in probiotic (beneficial) or pathogenic (harmful) outcomes to the host. Therapeutic benefits of probiotic bacteria have been shown previously, but the molecular mechanisms and the health-promoting effector components involved are still being elucidated. Some evidence suggests that probiotic bacteria can competitively adhere to intestinal mucus and displace pathogens. The adherence of probiotic bacteria to human intestinal mucus and cells appears to be mediated, at least in part, by secretome proteins. Secretome proteins-encoding open reading frames can be identified in bacterial genome sequences using bioinformatics. However, functional analysis of the translated secretome is possible only if many secretome proteins are expressed and purified individually. Phage display technology offers a very efficient way to purify and functionally characterise proteins by displaying them on the surface of the bacteriophage. While a phage display system for cloning secretome proteins has been previously reported it is not efficient for enrichment and display of Gram-positive secretome proteins. In this study a new phage display system has been developed and applied in direct selection, identification, expression and purification of Gram-positive Lactobacillus rhamnosus strain HN001 secretome proteins. The new phage display system is based on the requirement of a signal sequence for assembly of sarcosyl-resistant filamentous phage virions. Using this system 89 secretome open reading frames were identified from a library of only 106 clones, performing at least 20-fold more efficiently than the previously reported enrichment method. Seven of the identified secretome proteins are unique for L. rhamnosus HN001. A L. rhamnosus HN001 shot-gun phage display library was also constructed to capture proteins that mediate adhesion or aggregation, initial steps in establishing host-microbe contact or forming multicellular aggregates, both of which may lead to beneficial effects – colonisation of the gastro-intestinal tract and exclusion of pathogens. In search for proteins involved in adhesion, a L. rhamnosus HN001 shot-gun phage display library was screened against the human extracellular matrix component fibronectin commonly used as binding target by bacteria that colonise diverse tissues. This screen selected, instead of a fibronectin-binding protein, a protein that binds to avidin, used to immobilise biotinylated fibronectin. Affinity screening of the shot-gun library for binding to L. rhamnosus HN001 cells identified a secretome protein, Lrh33, as an HN001-cell surface binding protein. This protein contains two bacterial immunoglobulin-like domains type 3. Analysis of phage-displayed nested deletions of Lrh33 determined that the proximal (N-terminal) immunoglobulin-like domain is not sufficient for binding; only the constructs displaying both domains demonstrated binding to HN001. Lrh33 does not have any similarity to previously identified Lactobacillus-binding proteins and no match in the NCBI database (at a cutoff value of > e-13), hence it represents potentially a new type of bacterial auto-aggregation protein.
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Wan, Kanglin. "Diversité génétique et résistance aux médicaments anti-tuberculeux de Mycobacterium tuberculosis en Chine." Phd thesis, Université Paris Sud - Paris XI, 2007. http://tel.archives-ouvertes.fr/tel-00734513.
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Ce manuscrit décrit la distribution géographique des génotypes de Mycobacterium tuberculosis dans une large partie de la Chine, et l'étude d'une éventuelle corrélation avec la vaccination BCG et la résistance aux antibiotiques. La prévalence de la résistance a été analysée dans 10 provinces, et les mutations responsables de la résistance à la rifampicin et à l'INH ont été recherchées. La distribution des différents génotypes a été analysée par spoligotypage, par l'identification de délétions génomiques et par l'analyse du polymorphisme de répétitions en tandem (MLVA). Le génotype Beijing représente 55 à 93% des souches dans les provinces étudiées, avec un gradient allant du Sud vers le Nord de la Chine. Plusieurs autres familles sont identifiées, toutes appartenant à la clade dite " Moderne ". Les familles " Chine 2 " et " Chine 3 " qui représentent l'essentiel des autres souches sont rares en dehors de la Chine. Quelques souches de la famille CAS (Central Asia) sont également rencontrées dans une province et un ancêtre possible de la lignée Beijing dans une autre. La prévalence de la résistance aux antibiotiques a été mesurée parmi plus de 2000 isolats de 10 provinces. Le pourcentage de souches résistantes à au moins une drogue est de 45% et celui des souches multirésistantes est d'environ 29%. En Chine, à la différence d'autres pays, aucune corrélation n'est observée entre le génotype Beijing et la vaccination BCG. La distribution des mutations du gène rpoB a été déterminée ainsi que les mutations responsables de la résistance à l'INH. Aucune mutation n'est observée dans les gènes oxyR et ahpC. Cette étude est la première étude de grande envergure effectuée en Chine sur la diversité génétique de M. tuberculosis et sur les mécanismes de résistance aux antibiotiques.
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Copp, Clinton W. "Production and Degradation of 4-Ethylphenol in LACTOBACILLUS SP. pep8 Cultures and in Blended Swine Lagoon Enrichments." TopSCHOLAR®, 2012. http://digitalcommons.wku.edu/theses/1189.
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4-Ethylphenol (4-EP) is a malodorant of swine waste and is derived from a component of lignin called p-coumaric acid (p-CA). The production of 4-EP from lignin in swine waste is untested. Additionally, the effect of Fe (III) on 4-EP levels is unknown. Four experiments were performed to determine if Lactobacillus sp. pep8 cultures, as well as enriched swine lagoon slurries, could liberate p-CA from lignin and convert p-CA to 4-EP. Furthermore, it was tested if the addition of Fe (III) influences the conversion of p- CA to 4-EP.
Experiment 1 tested Lactobacillus sp. pep8 cultures to determine if the addition of 10 mM Fe (III) and 0.2% sulphite lignin to Lactobacillussp. pep8 cultures would stimulate production of 4-EP.
Experiment 2 tested the effect of 0.2% sulphite lignin and 10 mM Fe (III) on 4-EP production in the presence of enriched swine lagoon slurries. On day 0 there was no detectable 4‐EP, for either 0.2% sulphite lignin addition or the 10 mmol l‐1 Fe (III) additions.
Experiment 3 tested alternative forms of lignin, including 0.2% sulphite, indulin, or sigma lignin as potential source compounds for 4-EP production in enriched swine lagoon slurries. 4-EP produced in all three conditions are likely endogenous to the lagoon slurry additions.
Experiment 4 was designed to measure the degradation of exogenous 4-EP with varying final concentrations of 4-EP in enriched swine lagoon slurries. Data in Figure 7 indicate immediate degradation of 4-EP by day 5, however, by day 7 synthesis of 4-EP occurred until day 14 where 4-EP levels remained in a steady state.
Our results suggest that when both Lactobacillus sp. pep8 cultures and enriched swine lagoons are supplemented with p-CA, 4-EP is produced indicating that p-CA serves as a source of 4-EP. However, when supplemented with Fe (III) and/or sulphite, indulin, or sigma lignin, 4-EP production was not stimulated. This data indicates that, 4- EP production is not enhanced by the presence of Fe (III) in either Lactobacillus sp. pep8 cultures or in enriched swine lagoon slurries. Furthermore, lignin did not serve as a source of 4-EP.
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Vergnes, Mike. "Plasticité fonctionnelle et structurale chez Legionella pneumophila - Impact des protéines de type histone sur la virulence et génotypage par les séquences d'insertion." Phd thesis, Grenoble, 2010. http://tel.archives-ouvertes.fr/tel-00670047.
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Le genre Legionella regroupe des bactéries pouvant causer chez l'homme une pneumonie fatale dans 10% des cas, la légionellose. Elles sont capables de coloniser tous les réseaux d'eau. Le génome de ces bactéries révèle une forte plasticité génomique, aux niveaux fonctionnel et structural. La première partie de cette thèse analyse l'impact des protéines de type histone sur la régulation de la virulence chez L. pneumophila. Ces protéines structurent le chromosome bactérien et influencent l'expression génique. Des mutants des gènes codant les protéines Dps et IHF ont été obtenus chez L. pneumophila et analysés pour leur sensibilité aux stress et leurs propriétés de virulence. Ces deux protéines sont impliquées dans la régulation de la virulence chez Legionella. De plus, Dps permet de diminuer la sensibilité au stress oxydatif et IHF régulerait l'entrée dans l'état VBNC (viable but non-culturable), un état physiologique dans lequel les bactéries sont viables mais ne sont plus cultivables. La seconde partie vise à utiliser la plasticité structurale, notamment celle induite par les éléments génétiques mobiles IS, comme outil épidémiologique. A l'heure actuelle, les méthodes d'identification ne permettent pas de discriminer les isolats de même espèce. Afin d'éviter de nouvelles contaminations, il est impératif d'identifier rapidement et avec précision l'installation contaminée, à partir des prélèvements de patients comme élément comparatif. L'utilisation d'une méthode RFLP-IS a permis de mettre en évidence une IS particulière, ISLpn11, possédant un taux de discrimination de 80% au sein de la souche L. pneumophila Paris, qui est responsable de plus de 10 % des épidémies en Europe.
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Makhloufi, Kahina Maya. "Caractérisation d'une bactériocine produite par une bactérie lactique Leuconostoc pseudomesenteroides isolée du boza." Phd thesis, Université Pierre et Marie Curie - Paris VI, 2011. http://tel.archives-ouvertes.fr/tel-00678029.
Full textAbstract:
Les bactéries lactiques préviennent la contamination de produits alimentaires par des bactéries pathogènes en inhibant leur prolifération, essentiellement par la production d'acide lactique et de peptides antimicrobiens nommés bactériocines. Les bactériocines sont synthétisées par voie ribosomique. Elles sont actives contre des bactéries phylogénétiquement proches incluant des pathogènes tels que Listeria et Enterococcus. Notre étude a porté sur l'isolement et la caractérisation d'une bactériocine produite par la souche KM432Bz isolée d'une boisson fermentée des Balkans, le boza et que nous avons identifiée comme un Leuconostoc pseudomesenteroides. Des analyses structurales par spectrométrie de masse et dégradation d'Edman de la bactériocine produite par Ln. pseudomesenteroides KM432Bz ont montré que sa structure primaire était similaire à celles de deux bactériocines de classe IIa, les leucocines A et B et à la leucocine A-QU15 récemment découverte. Le système génétique impliqué dans la biosynthèse de cette leucocine a été identifié et analysé. Le gène codant la préleucocine KM432Bz est identique à ceux codant les préleucocines B et A-QU15. La leucocine produite par la souche KM432Bz présente un spectre d'activité dirigé contre des espèces proches de la souche productrice telles que Lactobacillus, Leuconostoc et Weissella ainsi que des espèces pathogènes appartenant aux genres Listeria, Enterococcus et Streptococcus avec des concentrations minimales inhibitrices comprises entre 0,08 et 10 μM. Par ailleurs, il a été montré que le facteur de transcription s54, impliqué dans la transcription de la mannose perméase du système phosphotransférase, est impliqué dans la sensibilité de Listeria monocytogenes à cette bactériocine.
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Harlow, Brittany E. "Changes to the Equine Hindgut Microflora in Response to Antibiotic Challenge." UKnowledge, 2012. http://uknowledge.uky.edu/animalsci_etds/12.
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Antibiotics are important to equine medicine, but can cause detrimental side-effects including reduced feed intake, allergic reactions, and diarrhea. Antibiotic-associated diarrhea (AAD) is attributed to disruption of the hindgut microflora, permitting proliferation of pathogenic microbes. The objectives were to evaluate the effects of antibiotics on beneficial fecal bacteria, AAD-associated pathogens, microbial species richness and fermentation. Horses were assigned to treatment groups: control (no antibiotics, n=6), trimethoprim-sulfadiazine (oral, n=6), or sodium ceftiofur (IM, n=6). Fecal samples were taken during adaptation (3 wk), antibiotic challenge (1 wk), and withdrawal (1 wk). Fecal cellulolytics decreased by >99% during challenge and did not recover during withdrawal (P < 0.0001). Lactobacilli decreased by >60% during challenge (P = 0.0453). Salmonella spp. increased 94% with trimethoprim-sulfadiazine challenge (P = 0.0115). There was no detectable Clostridium difficile during adaptation or in any control horse. C. difficile increased (P < 0.0001) when horses were challenged, and remained elevated 7 d after withdrawal. There was no effect of challenge on in vitro digestibility or microbial species richness as evaluated by denaturing gradient gel electrophoresis (P > 0.05). These results indicate that antibiotics can disrupt the normal flora and allow proliferation of pathogens, even without affecting digestibility and causing AAD.
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Tayou, Junior Kom. "Requirement of ßDELSEED-Motif of Escherichia coli F1FO ATP Synthase in Antimicrobial Peptide Binding." Digital Commons @ East Tennessee State University, 2011. https://dc.etsu.edu/etd/1260.
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F1FO ATP synthase is a membrane bound enzyme capable of synthesizing and hydrolyzing ATP. Lately, α-helical cationic peptides such as melittin and melittin related peptide (MRP) were shown to inhibit E. coli ATP synthase. The proposed but unconfirmed site of inhibition is βDELSEED-motif formed by the residues 380-386, located at the interface of α/β subunit of ATP synthase. This project was a mutagenic analysis of βDELSEED-motif residues to understand the binding mechanism and mode of action of peptide inhibitors. The study addressed 2 main questions: Are the antibacterial/anticancer effects of these peptides related to their inhibitory action on ATP synthase through interaction with the βDELSEED-motif? If so, which amino acid residues play critical role in peptide binding?
The findings demonstrated that the βDELSEED-motif is the binding site of the above peptides on ATP synthase and Glutamate residues are more important in peptide binding than the Aspartate residues.
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Ball, Kelly. "A Modified Scheme for the Isolation and Enumeration of Bacteria in Municipal Sewage Sludge." TopSCHOLAR®, 1992. http://digitalcommons.wku.edu/theses/1884.
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Because of the potential health hazards associated with the use of sludge for agricultural purposes, Dudley et al (1980) published a scheme for the routine analysis of bacteria in municipal sewage sludge. In this study, the Dudley et al scheme (1980) was modified by updating some of the procedures. Aerobically digested sludge generated by the Bowling Green Wastewater Treatment Plant, Bowling Green, Kentucky, was analyzed using the modified scheme. Sludge samples were collected once every two months over a one-year period from October 1989 to August 1990.
Egg yolk-free tryptose sulfite cycloserine agar in conjunction with the revewrse CAMP test was used to assay for Clostridium perfringens. This procedure improved the one proposed by Dudley et al. (1980) by achieving a higher confirmation rate, reducing testing time, allowing for easier interpretation of results, and increasing accuracy.
Selective and differential media by Rippey and Cabelli (1979) were added to the scheme to isolate Aeromonas, Aeroomonas hydrophila and Aeromonas caviae were successfully isolated wand were identified using the system by Cunliffe and Adcock (1989) for speciating aeromonads.
Baird-Parker medium was compared to mannitol salt agar for effectiveness in isolating Staphylococcus from sludge. Statistical analysis showed Baird-Parker medium to be significantly more effective than mannitol salt agar. However, neither agar reduced background flora to acceptable levels. Staphylococcus isolates were subject to species identification by the API Staph Ident system (Analytab Products, Plainview, New York). Staphylococcus xylosus, Staphylococcus haemolyticus, and Staphylococcus epidermidis were found to be present in the sludge.
A procedure by Ottolenghi and Hamparian (1987) was employed to isolate Salmonella in sludge. No salmonellae were isolated over the one year period.
Over the year-long study, bacterial numbers, with the exception of Clostridium perfringens and the total aerobic count, fluctuated with variations in the aerobic digester temperature. Numbers decreased as temperature increased. Clostridium perfringens counts were the most consistent throughout the year and exceeded fecal coliform and fecal streptococci counts in five of the six samplings.
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Coquel, Anne-Sophie. "Dynamique de l'agrégation protéique chez la bactérie Escherichia coli." Phd thesis, INSA de Lyon, 2012. http://tel.archives-ouvertes.fr/tel-00778887.
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L'agrégation protéique joue un rôle clé dans la dégénérescence cellulaire et est notamment reliée à de nombreuses maladies humaines en lien avec le vieillissement telles que les maladies d'Alzheimer et Parkinson ou encore la maladie du prion. Chez la bactérie Escherichia coli, l'accumulation de dommages sous forme d'agrégats protéiques et leur ségrégation asymétrique au pôle ont permis de démontrer des signes de vieillissement chez cette bactérie. Cette thèse s'est concentrée sur l'étude de la dynamique spatiale des agrégats protéiques in vivo chez la bactérie E. coli. Les agrégats protéiques peuvent être classifiés comme corps d'inclusion dont on dit souvent qu'ils sont amorphes ou comme amyloïdes dont le niveau de structuration est très élevée par la présence de nombreux feuillets β. Combinant une double approche théorique et expérimentale, basée sur la modélisation et la microscopie time-lapse et microfluidique, nous avons étudié le mécanisme gouvernant le mouvement des agrégats protéiques et la transmission verticale d'agrégats de type prionoide sur plusieurs dizaines de générations. Nos résultats indiquent clairement que les agrégats protéiques sont régis par un mouvement Brownien de diffusion avec un coefficient de diffusion dépendant de la taille de la molécule. L'étude de protéinopathie amyloïde a démontré l'existence de lignages propageant deux types d'agrégats : globulaire ou en forme de "comet-like". Les lignées présentant les agrégats sous forme globulaire indiquent une augmentation de la taille des agrégats jusqu'à inhibition de la division cellulaire tandis que la forme "comet-like" est moins préjudiciable à la croissance. Nous avons également observé à faible fréquence des lignées avec un changement de type d'agrégat. A partir d'un agrégat gobulaire, des agrégats "comet-like" peuvent naître.
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Ngeny, Beverly C. "Effects of Lactobacillus rhamnosus Milk Isolate on the Production of Inflammatory Cytokines in Enterocytes." Digital Commons @ East Tennessee State University, 2016. https://dc.etsu.edu/etd/3027.
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In the gastrointestinal tract, probiotics have been shown to promote host immunity and to regulate immune signaling pathways. This study used Caco-2 cell line to examine the effects of a Lactobacillus rhamnosus isolate from “amabere amaruranu” a Kenyan traditional cultured milk, on the production inflammatory cytokines in enterocytes. Live Lactobacillus rhamnosus (MRS6AN), its cytoplasmic fraction (CF), filtered spent broth (FSB) or heat inactivated FSB (HIB) were used as treatments on differentiated Caco-2 cell monolayer in transwells. Cytokine content in the cell lysates, apical and basolateral supernatants were determined using ELISA. Caco-2 cell lysate treatments showed significantly increased anti-inflammatory TGF-β (ng/ml) levels on average about 100x more compared to the increase in pro-inflammatory IL-8 (pg/ml) levels. These levels were significantly reduced after inhibition of NF-κB. In conclusion, live Lactobacillus rhamnosus, its CF, FSB or HIB seemed to modulate the production of inflammatory cytokines in enterocytes partly via the NF-κB signaling pathway.
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Frazier, Ashley Denise. "An Investigation of Bacterial Ribonucleases as an Antibiotic Target." Digital Commons @ East Tennessee State University, 2012. https://dc.etsu.edu/etd/1417.
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Antibiotics have been commonly used in medical practice for over 40 years. However, the misuse and overuse of current antibiotics is thought to be the primary cause for the increase in antibiotic resistance.
Many current antibiotics target the bacterial ribosome. Antibiotics such as aminoglycosides and macrolides specifically target the 30S or 50S subunits to inhibit bacterial growth. During the assembly of the bacterial ribosome, ribosomal RNA of the 30S and 50S ribosomal subunits is processed by bacterial ribonucleases (RNases). RNases are also involved in the degradation and turnover of this RNA during times of stress, such as the presence of an antibiotic. This makes ribonucleases a potential target for novel antibiotics.
It was shown that Escherichia coli mutants that were deficient for RNase III, RNase E, RNase R, RNase G, or RNase PH had an increase in ribosomal subunit assembly defects. These mutant bacterial cells also displayed an increased sensitivity to neomycin and paromomycin antibiotics. My research has also shown that an inhibitor of RNases, vanadyl ribonucleoside complex, potentiated the effects of an aminoglycoside and a macrolide antibiotic in wild type Escherichia coli, methicillin sensitive Staphylococcus aureus, and methicillin resistant Staphylococcus aureus.
RNases are essential enzymes in both rRNA maturation and degradation. Based on this and previous work, the inhibition of specific RNases leads to an increased sensitivity to antibiotics. This work demonstrates that the inhibition of RNases might be a new target to combat antibiotic resistance.
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Methot, Pierre-Olivier. "Historical epistemology of the concept of virulence : molecular, ecological, and evolutionary perspectives on emerging infectious diseases in the 19th and 20th century." Thesis, University of Exeter, 2011. http://hdl.handle.net/10036/3494.
Full textAbstract:
This thesis focuses on the trajectory of the biomedical concept of virulence from 1880 until the present. Following the concept across disciplinary boundaries, from a longue durée history perspective, it explores how virulence was shaped through two distinct, although sometimes overlapping, “styles of reasoning”. Located at the intersection of several distinct research domains in biology and medicine, the concept of virulence provides, in addition, a window into the complex and changing relations between evolutionary biology and the health sciences (broadly construed) over the past two centuries. Moving back and forth between field experiments and the laboratory, this work examines, through the lens of historical epistemology, the emergence of what I call the molecular and the ecological styles, and their respective conceptual practices. It focuses on the ways in which these styles operationalize the distinction between virulent or avirulent organisms in sometimes opposite sense: Whereas in the molecular (or endogenous) style the expression of virulence is explained by properties of internal structures of the infectious agent (e.g. polysaccharide capsule, virulence gene, or pathogenicity island), the concept of virulence in the ecological (or exogenous) style reflects, in contrast, either a lack of adaptation between two species (avirulence hypothesis) or the existence of one or more ecological compromises between, say, the mode of transmission of a pathogen and its host’s recovery rate (trade-off model). Both styles can be said to originate in the medical bacteriology of the late-nineteenth century, but while the former grew mostly out of the work of Louis Pasteur and Robert Koch in Europe, the latter was primarily shaped by Theobald Smith in the United States. Nearly a century later, the introduction of the category of emerging infectious disease within public health discourses in the mid-1990s facilitated a rapprochement between the two styles that had, so far, remained apart. Employing the 1918–1919 influenza pandemic as an example in which to illustrate the trajectory of the molecular and the ecological approaches, the diversity of explanatory schemes developed to account for the pandemic’s exceptional virulence points toward an unresolved, and yet productive, epistemic tension between the two styles, on the one hand, and the intrinsic polarity of the concept of virulence itself, on the other.
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Nham, Toan. "Suivi in vivo et en temps réel du processus infectieux induit par Yersinia pestis." Phd thesis, Université Paris-Diderot - Paris VII, 2012. http://tel.archives-ouvertes.fr/tel-00731170.
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Après trois pandémies majeures responsables de millions de morts, la peste n'a pas encore disparu. Cette maladie est causée par la bactérie Yersinia pestis, dont les mécanismes de virulence sont encore mal compris. Le suivi d'infection de la peste bubonique chez la souris, méthode classique pour étudier le processus infectieux, requiert beaucoup d'animaux et de temps pour obtenir des résultats significatifs. L'imagerie in vivo et en temps réel par bioluminescence permet de suivre la progression du pathogène au cours du processus infectieux en observant les animaux de façon non invasive. Nous avons transformé la souche virulente CO92 avec le plasmide pEm7-luxCDABE et confirmé la production de bioluminescence in vitro et in vivo. Nous avons pu quantifier la charge bactérienne dans plusieurs organes colonisés sans sacrifier l'animal et établir le schéma de progression de la bactérie au cours de la maladie. Après formation d'un foyer infectieux au site d'injection, la colonisation du ganglion lymphatique inguinal drainant ce site a été observée. Nous avons démontré que la bactérie suit un trajet direct du ganglion lymphatique inguinal au ganglion lymphatique axillaire. L'étape suivante est la colonisation des organes filtrant le sang, puis survient la septicémie dans les phases terminales de la mort. Nous avons établi que la forte variabilité dans le processus infectieux était due au temps pendant lequel la bactérie était contenue au site d'injection. À partir du moment où les ganglions lymphatiques sont colonisés, la cinétique de progression est à la fois régulière et rapide ; la septicémie survient dans les deux jours, suivie de près par la mort.
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Dadi, Prasanna Keerthi. "Inhibition of Escherichia coli ATP Synthase by Polyphenols and Their Derivatives." Digital Commons @ East Tennessee State University, 2010. https://dc.etsu.edu/etd/1704.
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We have studied the inhibitory effect of natural and structurally modified polyphenols on Escherichia coli ATP synthase to test (I) if the beneficial dietary effects of polyphenols are related to their inhibitory actions on ATP synthase, (II) if inhibitory effects of polyphenolic compound could be augmented through structural modifications, and (III) if they can act as antimicrobial agent through their actions on ATP synthesis. X-ray crystal structures of polyphenol binding sites suggested that polyphenols bind at a distinct polyphenol binding pocket, at the interface of α,β,γ-subunits. We found that both natural and modified polyphenols inhibit E. coli ATP synthase to varying degrees and structural modifications resulted in augmented inhibition. Inhibition was reversible in all cases. Both natural and modulated compounds inhibited E. coli cell growth to varying degrees. We conclude that dietary benefits of polyphenols may be in part due to the inhibition of ATP synthase.
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Pratte, Zoe A. "Investigating the Driving Mechanisms Behind Differences in Bleaching and Disease Susceptibility Between Two Scleractinian Corals, Pseudodiploria Strigosa and Diploria Labyrinthiformis." FIU Digital Commons, 2015. http://digitalcommons.fiu.edu/etd/2217.
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Disease and bleaching are two conditions which commonly lead to coral death. Among coral species, susceptibility to disease and bleaching is variable, and Pseudodiploria strigosa tends to be diseased more than Diploria labyrinthiformis, while D. labyrinthiformis bleaches more readily. The focus of this dissertation was to investigate and compare multiple components of these two coral species, and identify how they may relate to disease and bleaching resistance. Compenetnts examined included the surface mucopolysacharide layer (SML) thickness, gene expression, microbial associates, and a white plague aquarium study. The SML thickness decresased with increasing temperature regardless of coral species, indicating that SML thickness does not likely play a role in differences between susceptablities of these two coral species. However, Diploria labyrinthiformis had a lower mortality rate at 31°C, had fewer differentially expressed genes assossiated with stress, and upregulated genes associated with innate immunity in the summer, all of which may contribute to its relative disease resistance. The bacterial associates of each coral species were also monitored. Differences between the two coral species were primarily caused by Clostridia, Gammaproteobacteria, and rare species which may contribute to the relatively higher disease susceptibility of P. strigosa. Lastly, an aquarium study suggested that a potential pathogen of the Roseobacter clade infects both D. labyrinthiformis and P. strigosa, and might be transmitted by the Cryptochiridae gall crab, indicating that potential disease vectors associated with these two coral species may also play a role in disease resistance and resilience.
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Hutson, Savannah. "The Effects of Farnesol, a Quorum Sensing Molecule from Candida albicans, on Alcaligenes faecalis." Digital Commons @ East Tennessee State University, 2020. https://dc.etsu.edu/honors/539.
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Quorum sensing molecules have become a recent focus of study to learn if and how they can be used, both on their own and in conjecture with current antimicrobial methods, as a means of bacterial control. One such quorum sensing molecule is the sesquiterpene alcohol, Farnesol, which is synthesized and released by the fungus, Candida albicans. In most in-vivo cases, our laboratory has shown that Alcaligenes faecalis overtakes C. albicans, preventing its growth. However, as a way to counteract this inhibitory effect, Farnesol may be one way that Candida has found to fight back. In this study, we focused on the inhibitory properties of Farnesol for growth and motility of A. faecalis, as well as, the molecule’s ability to prevent Alcaligenes from creating biofilms and/or degrading them once they have already been established. Our experiments show evidence that Farnesol is able to inhibit both the growth and motility of A. faecalis, and determination of the specific concentrations of Farnesol needed to see the largest effects on A. faecalis biofilms. Our hope is that in future studies, we will be able to add varying concentrations of the Farnesol to known and widely used antibiotics in order to increase the effectiveness of antibiotics against bacterial strains, both in the Alcaligenes genus and in other genus, that have previously been considered “antibiotic resistant”.
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Tatke, Gorakh Digambar. "Elucidating The Role of MifS-MifR Two-Component System in Regulating Pseudomonas aeruginosa Pathogenicity." FIU Digital Commons, 2016. http://digitalcommons.fiu.edu/etd/3002.
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Pseudomonas aeruginosa is a Gram-negative, metabolically versatile, opportunistic pathogen that exhibits a multitude of virulence factors, and is extraordinarily resistant to a gamut of clinically significant antibiotics. This ability is in part mediated by two-component systems (TCS) that play a crucial role in regulating virulence mechanisms, metabolism and antibiotic resistance. Our sequence analysis of the P. aeruginosa PAO1 genome revealed the presence of two open reading frames, mifS and mifR, which encodes putative TCS proteins, a histidine sensor kinase MifS and a response regulator MifR, respectively. This two-gene operon was found immediately upstream of the poxAB operon, where poxB encodes a chromosomal ß-lactamase, hinting at the role of MifSR TCS in regulating antibiotic resistance. However, loss of mifSR had no effect on the antibiotic resistance profile when compared to P. aeruginosa parent PAO1 strain. Subsequently, our phenotypic microarray data (BioLOG) and growth profile studies indicated the inability of mifSR mutants to grow in α-ketoglutarate (α-KG), a key tricarboxylic acid (TCA) cycle intermediate, as a sole carbon source. To date, very little is known about the physiology of P. aeruginosa when provided with α-KG as its sole carbon source and the role of MifS and MifR TCS in virulence. Importantly, in the recent years, α-KG has gained notoriety for its newly identified role as a signaling molecule in addition to its conventional role in metabolism. This led us to hypothesize that MifSR TCS is involved in α-KG utilization and virulence in P. aeruginosa. Using mifS, mifR and mifSR clean in-frame deletion strains, our study demonstrates that the MifSR TCS modulates the expression P. aeruginosa kgtP (PA5530) and pcaT (PA0229) genes encoding putative α-KG permeases. In addition, our study shows that the MifSR-regulation of these transporters requires functional sigma factor RpoN (σ54). Loss of mifSR in the presence of α-KG, resulted in differential regulation of P. aeruginosa key virulence determinants including biofilm formation, motility, cell cytoxicity and the production of pyocyanin and pyoverdine. Involvement of multiple regulators and transporters suggests the presence of an intricate circuitry in the transport of α-KG and its importance in P. aeruginosa survival. This is further supported by the α-KG-dependent MifSR regulation of multiple virulence mechanisms. Simultaneous regulation of multiple mechanisms involved in P. aeruginosa pathogenesis suggests a complex mechanism of MifSR action. Understanding the physiological cues and regulation would provide a better stratagem to fight often indomitable P. aeruginosa infections.
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De, Negri Rafaela. "EQUINE SERUM ANTIBODY RESPONSES TO STREPTOCOCCUS EQUI AND STREPTOCOCCUS ZOOEPIDEMICUS." UKnowledge, 2013. http://uknowledge.uky.edu/gluck_etds/13.
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Streptococcus zooepidemicus (Sz) and Streptococcus equi (Se) share 98% DNA sequence homology, but display different pathogenic properties. Infection by one organism does not cross-protect against the other. To better understand pathogenic differences between these organisms and gain information about which proteins are expressed in horses infected experimentally with Se, intrauterine Sz or naturally with respiratory Sz we compared antibody specificities of convalescent sera using ELISA. These comparisons were based on sets of 8 and 14 immunoreactive recombinant proteins of Se strain CF32 and Sz strain NC78, respectively. Sera from donkeys that were previously naturally affected with strangles and later developed Sz pneumonia secondary to an experimental influenza challenge were also included.
Serum antibody responses were quantitatively and qualitatively much greater following recovery from strangles than following respiratory Sz infection. Increased reactions to Se proteins IdeE2, Se75.3, Se46.8, Se18.9 and Se42.0 were observed for the majority of strangles sera but not for sera from respiratory Sz infection cases. Reactions of sera from Sz respiratory disease to Sz proteins varied greatly and were mostly to HylC and ScpC. Interestingly, sera of donkey recovered from Sz bronchopneumonia did not show increased antibody reaction to any of the proteins even though these donkeys had also recovered from clinical strangles 6 months previously. Only 1/5 mare with Sz placentitis presented increased serum antibody responses to MAP. In conclusion, adaptive immune responses to Se of horses with strangles are stronger and involve a greater number of proteins than adaptive immune responses to Sz infection of the lower respiratory tract.
In an effort to develop an improved vaccine against Se, modified live strain of EHV-1, RacH was constructed to express three recombinant antigens of Se SeM, IdeE and Se18.9. Two groups of 10 and 2 ponies were vaccinated intramuscularly or intranasally, respectively. Another group (n=6) vaccinated with empty RacH served as controls. Sera from 2/3 ponies from each vaccination groups and 1/2 serum from IN vaccinated ponies showed increased serum neutralizing antibodies to EHV-1. ELISA detected no significant increase in antibodies to proteins. Only one IM and IN vaccinated pony showed serum bactericidal activity post vaccination.
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AQBI, HUSSEIN F. "Preconditioning of the tumor microenvironment by means of low dose chemotherapies for an effective immunotherapy of breast cancer." VCU Scholars Compass, 2019. https://scholarscompass.vcu.edu/etd/6025.
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Breast cancer mortality is mainly due to distant recurrence of the disease arising from dormant tumor cells established by cancer therapies. Patients who initially respond to cancer therapies often succumb to distant recurrence of the disease. It is not clear why people with the same type of breast cancer respond to treatments differently; some escape from dormancy and relapse earlier than others. In addition, some tumor clones respond to immunotherapy while others do not. We investigated how autophagy plays a role in accelerating or delaying recurrence of neu overexpressing mouse mammary carcinoma (MMC) following adriamycin (ADR) treatment, and in affecting response to immunotherapy. We explored two strategies: 1) transient blockade of autophagy with chloroquine (CQ), which blocks fusion of autophagosomes and lysosomes during ADR treatment, and 2) permanent inhibition of autophagy by a stable knockdown of ATG5 (ATG5KD), which inhibits the formation of autophagosomes in MMC during and after ADR treatment. We found that while CQ prolonged tumor dormancy, but that stable knockdown of autophagy resulted in early escape from dormancy and recurrence. Interestingly, ATG5KD MMC contained an increased frequency of ADR-induced polyploid-like cells and rendered MMC resistant to immunotherapy. On the other hand, a transient blockade of autophagy did not affect the sensitivity of MMC to immunotherapy. Our observations suggest that while chemotherapy-induced autophagy may facilitate tumor relapse, cell-intrinsic autophagy delays tumor relapse, in part, by inhibiting the formation of polyploid-like tumor dormancy.
Although immunotherapy of breast cancer by means of anti-HER2 antibodies prolongs survival of breast cancer patients, disease recurrence remains a major challenge. On the other hand administration of human vaccines against infectious disease in a preventive setting or during latency/dormancy has been successful in offering a cure. Here, we sought to use adoptive immunotherapy (AIT) at the time of tumor dormancy in order to prevent progression of breast cancer. We used a low dose immunogenic chemotherapy by means of 5-FU, Adriamycin, and Cyclophosphamide (FAC) in order to stabilize tumor progression prior to AIT using autologous tumor-reactive lymphocytes. Low dose FAC established local tumor dormancy, inhibited distant tumor dormancy occurring long before distant metastasis, and induced predominate a Ki67- quiescent type of tumor dormancy, which is less susceptible to tumor immunoediting. Dormant tumor cells expressed the cell survival pathways, including the endothelin receptor/ligand (ETRA, ETRB and ET-1) and PD-L1, thereby protecting them from elimination by AIT. In addition, tumor-reactive CD8+ T cells also produced ET-1 as a survival ligand for ETRA positive tumor cells. A combination of AIT with the blockade of tumor cell survival pathways resulted in a significant improvement of AIT against tumor dormancy. We also showed that the inhibition Bcl-xL downstream of the tumor cell survival pathways is specifically effective against dormant tumor cells, suggesting a combination of AIT with small molecules inhibitors of Bcl-xL. Altogether, we showed that distant tumor dormancy is established long before distant recurrence of breast cancer, and that the expression of several tumor cell survival pathways in dormant cells protects them from immunotherapy. Our results suggest that immunotherapeutic targeting of tumor dormancy combined with the blockade of a common downstream cell survival pathway could prevent tumor progression and recurrence of the disease.
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Moussaoui, Louardi. "Applications de la spectrométrie de masse type MALDI-TOF à la bactériologie et à la distinction de variants génétiques." Phd thesis, Université de Strasbourg, 2012. http://tel.archives-ouvertes.fr/tel-00872251.
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L'objectif de mon travail fut de valider et d'optimiser la spectrométrie de masse de type MALDI-TOF pour l'identification et la classification d'un ensemble de bactéries pathogènes ou opportunistes chez l'homme, en enrichissant une base de données et en testant la robustesse de la méthode, afin d'obtenir une méthode rapide fixe et fiable d'acquisition de résultats. Les différents résultats obtenus ont permis la validation de la technique comme outil d'identification bactérienne fiable en routine. Elle permet désormais de caractériser les mélanges de deux bactéries voir même la différentiation d'espèces très proches comme les Shigella spp et E. coli. Nous avons montré que la technique sera encore améliorée par un outil supplémentaire de comparaison des souches pour une veille épidémiologique "en temps réel", sans investissement supplémentaire, en permettant plusieurs types d'économie. Elle apporte un gain réel dans la prise en charge du patient et le choix éclairé des antibiotiques testés pour l'antibiogramme. La technique peut aussi constituer un outil alternatif de sérotypage.
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Balasubramanian, Deepak. "Pseudomonas Aeruginosa AmpR Transcriptional Regulatory Network." FIU Digital Commons, 2013. http://digitalcommons.fiu.edu/etd/863.
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In Enterobacteriaceae, the transcriptional regulator AmpR, a member of the LysR family, regulates the expression of a chromosomal β-lactamase AmpC. The regulatory repertoire of AmpR is broader in Pseudomonas aeruginosa, an opportunistic pathogen responsible for numerous acute and chronic infections including cystic fibrosis. Previous studies showed that in addition to regulating ampC, P. aeruginosa AmpR regulates the sigma factor AlgT/U and production of some quorum sensing (QS)-regulated virulence factors. In order to better understand the ampR regulon, the transcriptional profiles generated using DNA microarrays and RNA-Seq of the prototypic P. aeruginosa PAO1 strain with its isogenic ampR deletion mutant, PAO∆ampR were analyzed. Transcriptome analysis demonstrates that the AmpR regulon is much more extensive than previously thought influencing the differential expression of over 500 genes. In addition to regulating resistance to β-lactam antibiotics via AmpC, AmpR also regulates non-β-lactam antibiotic resistance by modulating the MexEF-OprN efflux pump. Virulence mechanisms including biofilm formation, QS-regulated acute virulence, and diverse physiological processes such as oxidative stress response, heat-shock response and iron uptake are AmpR-regulated. Real-time PCR and phenotypic assays confirmed the transcriptome data. Further, Caenorhabditis elegans model demonstrates that a functional AmpR is required for full pathogenicity of P. aeruginosa. AmpR, a member of the core genome, also regulates genes in the regions of genome plasticity that are acquired by horizontal gene transfer. The extensive AmpR regulon included other transcriptional regulators and sigma factors, accounting for the extensive AmpR regulon. Gene expression studies demonstrate AmpR-dependent expression of the QS master regulator LasR that controls expression of many virulence factors. Using a chromosomally tagged AmpR, ChIP-Seq studies show direct AmpR binding to the lasR promoter. The data demonstrates that AmpR functions as a global regulator in P. aeruginosa and is a positive regulator of acute virulence while negatively regulating chronic infection phenotypes. In summary, my dissertation sheds light on the complex regulatory circuit in P. aeruginosa to provide a better understanding of the bacterial response to antibiotics and how the organism coordinately regulates a myriad of virulence factors.
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Ballihaut, Guillaume. "Détection et identification de sélénoprotéines par électrophorèse sur gel associée aux spectrométries de masse atomique et moléculaire." Phd thesis, Université de Pau et des Pays de l'Adour, 2007. http://tel.archives-ouvertes.fr/tel-00656273.
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Le sélénium est un élément trace connu pour son caractère essentiel au développement de nombreux organismes vivants mais également pour ses effets bénéfiques sur la santé humaine. Les recherches effectuées ces cinq dernières années ont renforcé l'idée que ces propriétés seraient dues à la synthèse de sélénoprotéines chez les êtres vivants. Les méthodes classiques d'analyse protéomique ne sont pas adaptées à l'identification ciblée de ces sélénoprotéines très minoritaires. Les travaux de cette thèse ont consisté à développer des méthodes complémentaires plus spécifiques et sensibles en vue de leur détection et de leur identification. Un étalon protéique sélénié stable a été produit pour développer ces méthodes. Le couplage de l'ablation laser et de la spectrométrie de masse couplée à un plasma induit (LA-ICP-MS) a été mis en oeuvre pour la détection des sélénoprotéines après une séparation par électrophorèse sur gel de polyacrylamide (PAGE). Le dispositif d'ablation laser conventionnel a ici été amélioré avec un laser femtoseconde à balayage très rapide permettant une détection plus sensible des sélénoprotéines dans les échantillons biologiques. Une fois détectées, les sélénoprotéines ont été identifiées en spectrométrie de masse moléculaire. Pour cela les sélénopeptides issus de la digestion enzymatique des sélénoprotéines sont d'abord repérés en nanochromatographie couplée à l'ICP-MS puis séquencés en nanochromatographie couplée à la spectrométrie de masse en tandem à ionisation électrospray (nanoHPLC-ESI-MS/MS). La procédure en LA-ICP-MS développée a notamment permis la détection de nouvelles sélénoprotéines chez la bactérie Desulfococcus multivorans. La procédure d'identification a été validée sur deux sélénoprotéines purifiées thiorédoxine réductase et glutathione peroxydase carboxyméthylée. La méthodologie développée contribuera à l'identification de nouvelles sélénoprotéines pour une meilleure compréhension des rôles physiologiques de ces molécules chez de nombreux êtres vivants.
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Bornot, Julie. "Caractérisation et quantification rationnelles de la physiologie de Deinococcus geothermalis par une approche de génie nutritionnel." Phd thesis, INSA de Toulouse, 2013. http://tel.archives-ouvertes.fr/tel-01073388.
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Les bactéries du genre Deinococcus sont des microorganismes qui présentent des propriétés remarquables de résistance aux conditions extrêmes telles que les radiations ionisantes, les stress oxydatifs, la dessiccation ou encore les températures extrêmes. L'exploitation de leurs capacités présente donc un réel intérêt pour les procédés biotechnologiques de production de métabolites d'intérêt, les biocarburants. Néanmoins, la physiologie et le métabolisme de ces microorganismes ont été très peu étudiés à ce jour. L'objectif de ce travail est d'identifier les exigences nutritionnelles d'une souche de déinocoques, Deinococcus geothermalis, dans le but de définir la composition d'un milieu synthétique permettant une croissance sans limitation. L'étude statistique de quarante huit formulations de milieux de culture a mis en évidence la variabilité qualitative et quantitative des compositions des milieux utilisés pour leur culture. Il ressort que l'ajout de facteurs stimulant la croissance, sous forme d'extrait de levure, est indispensable à la croissance non limitée de la souche Deinococcus geothermalis DSM 11300. En bioréacteur à 45 °C, sur un milieu complexe et substrat glucose, une concentration finale de 2,7 gMS.L-1 a été obtenue en six heures avec un taux de croissance égal à 0,65 h 1. Les résultats ont montré une variabilité intra-espèce importante ; parmi les trois souches de Deinococcus geothermalis étudiées, DSM 11300, DSM 11301 et DSM 11302, la souche Deinococcus geothermalis DSM 11302 présente la meilleure croissance en termes de nombre de générations, en milieu défini ou complexe. L'étude de l'influence des étapes de préparation de l'inoculum a permis de standardiser les conditions de préparation de l'inoculum préalablement aux cultures en bioréacteur. La souche Deinococcus geothermalis DSM 11302 présente une croissance exponentielle durant quatre heures en bioréacteur sur un milieu défini avec 10 g.L-1 de glucose ; le taux de croissance de 0,25 h-1 ne se maintient pas sur une durée plus longue. Le rendement de production de biomasse à partir du glucose atteint 0,25 gX.gGlc-1 soit 0,30 CmolX.CmolGlc-1. L'ajout de sources carbonées, sources soufrées, vitamines ou autres facteurs de croissance ne permet pas d'améliorer la croissance de la souche dans ces conditions. Sur ces bases, la quantification de la physiologie de Deinococcus geothermalis DSM 11302 a été étudiée lors d'une culture en mode discontinu alimenté avec une stratégie d'apport de deux substrats, l'extrait de levure et le glucose. Le mode de conduite a permis de révéler que le glucose n'est pas la source de carbone assimilée préférentiellement et que l'extrait de levure peut être consommé comme source azotée organique et/ou comme source carbonée. Les déinocoques sont caractérisés par un métabolisme principalement protéolytique : il a été possible de confirmer que l'extrait de levure est indispensable à la croissance non limitée de Deinococcus geothermalis DSM 11302. Avec cette stratégie d'alimentation co-substrats, 253 g d'extrait de levure et 26 g de glucose ont été ajoutés pour produire 99 gMS de biomasse soit 9,6 gMS.L-1 en six heures. Les vitesses spécifiques maximales de consommation des substrats, extrait de levure et glucose, atteignent respectivement 0,68 et 0,39 Cmol.CmolX-1.h-1. Le taux de croissance exponentiel maximal obtenu est égal à 1.05 h-1, meilleur résultat décrit à ce jour pour un déinocoque. Ces résultats contribuent ainsi à implémenter l'état des connaissances sur la physiologie et les exigences nutritionnelles de Deinococcus geothermalis et donnent accès aux valeurs de paramètres cinétiques et de rendements, données absentes de la littérature
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Khamisse, Elissa. "Etude du microbiote susceptible de persister sur les surfaces d'un atelier de la filière viande bovine." Phd thesis, AgroParisTech, 2012. http://pastel.archives-ouvertes.fr/pastel-00770326.
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Ce travail de thèse concerne l'étude de l'écologie microbienne d'un atelier de découpe de viande bovine, dans le but de mieux comprendre la persistance bactérienne, c'est-à-dire, la présence répétée d'un même clone bactérien pendant une longue période malgré l'application bien conduite et régulière du nettoyage et de la désinfection (N-D). Des prélèvements par " chiffonnages " multiples de surfaces d'équipements ont été réalisés lors de trois campagnes de prélèvement espacées les unes des autres d'au moins six mois. Les prélèvements ont été réalisés sur un tapis convoyeur en polychlorure de vinyle (PVC) et sur des machines éplucheuses en acier inoxydable avant et après N-D. Nous avons quantifié les cellules totales (les cellules vivantes et les cellules mortes) par PCR quantitative en temps réel (qPCR), les cellules viables par EMA-qPCR, et les UFC (provenant de cellules cultivables) par dénombrement après incubation à 25°C sur gélose tryptone soja. Les résultats montrent qu'avant N-D, les cellules totales (en moyenne 5,6 - exprimé en log10 cellules/cm2 - sur PVC et 4,7 sur acier inoxydable) sont plus nombreuses que les cellules viables (4,5 sur PVC et 4,4 sur acier inoxydable) lesquelles sont plus nombreuses que les UFC (3,8 sur PVC et 2,9 sur acier inoxydable). Le N-D entraîne moins d'une réduction décimale (RD) des populations à l'exception des UFC sur acier inoxydable qui subissent 1,5 RD en moyenne. Ce dernier chiffre s'explique par des forces d'adhésion faibles. L'étude de la diversité des bactéries cultivables montre que sur un total de 51 genres identifiés, 13 seulement sont retrouvés lors des trois campagnes de prélèvements. Les isolats de ces 13 genres représentent 75, 72 et 62% des isolats des campagnes1, 2 et 3 respectivement. Parmi ces isolats, les plus fréquents sont (par ordre décroissant du nombre d'isolats) : Pseudomonas, Staphylococcus, Microbacterium, Acinetobacter, Chryseobacterium, Psychrobacter et Kocuria. Le génotypage d'isolats de 3 genres majoritaires (Staphylococcus, Pseudomonas et Acinetobacter) montre qu'une seule souche, Staphylococcus equorum, est sans aucun doute persistante. L'ensemble de ces observations montrent que l'écosystème varie d'une campagne à une autre. Ces modifications de la diversité bactérienne reflèteraient les modifications de flores des viandes traitées dans l'atelier, qui ont des origines multiples. En outre, il apparaît que, contrairement à ce qui est généralement admis, les bactéries à coloration de Gram négative cultivables sont plus facilement inactivées par le N-D que les bactéries à coloration de Gram positive. L'étude de l'écosystème par PCR-DGGE a permis d'identifier sept genres bactériens et montre que les espèces dominantes sont toutes sous forme vivante, autrement dit, aucune des espèces dominantes n'a été détectée uniquement sous forme de cellules mortes. Sur les sept genres identifiés six sont des Gram - dont majoritairement les genres Acinetobacter, Pseudomonas et Psychrobacter. Cette dominance montre que le N-D permet une forte perte de cultivabilité des bactéries Gram - mais qu'une grande partie n'est pas détachée. La dominance des bactéries Gram - observée par PCR-DGGE masque les staphylocoques qui ne sont pas détectés alors qu'ils sont majoritaires parmi la flore cultivable. Seul un genre bactérien, Propionibacterium, est identifié par PCR-DGGE uniquement mais il n'est trouvé qu'à une seule campagne et uniquement sur l'acier inoxydable avant N-D. En conclusion, l'avancée majeure de ce travail est la mise en évidence qu'une proportion importante de bactéries survit après les opérations très poussées de N-D mais pour une période transitoire.
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Lee, Chi-An. "Distribution of Enterotoxigenic Clostridium perfringens Spores in U.S. Retail Spices." 2016. https://scholarworks.umass.edu/masters_theses_2/427.
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246 samples of bulk and packaged spices from retail stores in the western, southeastern, southern, midwestern, and northeastern areas of the U.S. were examined for the presence of Clostridium -perfringens. Isolates were checked for the presence of the lecithinase gene (cpa) and enterotoxin genes (cpe) by PCR. Enterotoxin formation during sporulation was investigated using the Oxoid Toxin Detection Kit. Forty-three confirmed isolates (from 17% of total samples) were cpa-positive. Of those, 27 were cpe-positive. Together, levels of C. perfringens spores ranged from 3.6-2400/gm. The amount of enterotoxin in cell extracts ranged from 2-16 ng/ml. Some of the SEM images of isolated spore (# 78) and one plasmid-borne ent control (FD-153) showed an organized surface structure termed “candy-wrapper”. This extracellular structure remained after treatment with 0.1 % SDS for 1 hr, suggesting it was not composed of membrane debris from the mother cell. The D values of spores ranged from 1.19- 3.31 min. The addition of lysozyme in the plating medium elevated the recovery rate of heat-treated spores. The growth rate of a cocktail of spores from spices (# 31, # 32, # 45) between 4 to 5 hr after inoculation was determined with a doubling time of 6.82 min in hamburger. A cocktail of spores of plasmid-borne ent control showed an optimum growth rate between 5 to 8 hr after inoculation with doubling time of 15.98 min. However; spice isolate cocktail, plasmid-borne ent control cocktail (FD-5603 and FD-153), and a chromosome-borne ent control (NTCT 8239) were unable to germinate and outgrowth at 20oC. Inoculation in laboratory medium FTG indicated the same result as hamburger at 20oC. The ability of C. perfringens spores in spices to potentially survive cooking procedures can be followed by germination and growth of vegetative cells during improper cooling to levels associated with foodborne illness caused by this organism. Our results suggest that retail spices are potential vehicles of transmission of enterotoxin-positive C. perfringens.
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Rasolomboahanginjatovo, Hasina Santatriniaina. "Relations entre le statut utérin, les paramètres biochimiques du sérum et du liquide de lavage utérin et la production d’embryons chez les vaches laitières après surovulation." Thèse, 2011. http://hdl.handle.net/1866/5154.
Full textAbstract:
Le développement et la survie de l’embryon dépendent des nutriments fournis par les sécrétions utérines. Les objectifs de cette étude étaient de déterminer l’effet de la surovulation (SOV) sur la bactériologie et cytologie utérine et sur les paramètres biochimiques utérin et sérique et leurs effets sur le nombre d’embryons transférables (ET). Deux groupes de vaches Holstein (groupe I, non lactante, n=7 et groupe II, lactante, n=28) ont été respectivement induites en chaleur ou surovulées et ensuite inséminées. Au jour 7 du cycle œstral (J7) et lors du jour de la récolte (JR), un prélèvement individuel de sang et de liquide de lavage utérin a été fait pour l’analyse du statut bactériologique et cytologique de l’utérus et la mesure de la concentration de plusieurs paramètres biochimiques présélectionnés. Les embryons récoltés ont été évalués selon les critères de l’IETS. La SOV a donnée une moyenne de 7.39 ± 6.22 ovocytes/embryons dont 3.32 ± 4.81 ET. Il n’y avait pas de variation significative de la bactériologie et cytologie utérine des deux groupes entre J7 et JR. La concentration sérique de l’urée (P=0.0001), d’E2 (P=0.006); la concentration utérine du Glu (P=0.002), de Ck (p=0.0007), de LDH (P <0.0001), de PT (P=0.004), de P4 (P=0.008), de PGFM (P<0.0001) du groupe I et la concentration sérique de P4 (P<0.0001), de PGFM (P<0.0001); la concentration utérine de LDH (P=0.002), de PGFM (P<0.0001) du groupe II ont été significativement élevées à JR qu’à J7. La concentration utérine et sérique de l’urée (P<0.0001 et P<0.0001), de LDH (P<0.0001 et P=0.008), la concentration sérique de P4 (P=0.0002) et la concentration utérine de PT (P=0.0003) à JR du groupe II étaient différente du groupe I. Il n’y avait pas d’association entre la bactériologie et cytologie utérine et le nombre d’ET. Cependant, le nombre d’ET a été positivement corrélé avec la concentration sérique d’IGF-1 à J7 (r=0.45; P=0.001) et la concentration sérique de P4 à JR (r=0.43; P<0.05) et négativement corrélé avec la concentration utérine et sérique de PGFM à la fois à J7 (r=-0.54; P<0.005 et r=-0.67; P<0.001) et à JR (r=-0.48; P<0.01 et r=-0.57; P<0.002). Ces résultats suggèrent que la SOV induit des changements au niveau sérique et utérin qui affectent le nombre d’ET récoltés.The developing embryo is dependent on the nutrients provided by the oviduct and the uterine fluid. The objectives of this study were to determine the effect of SOV on uterine bacteriology and cytology, on serum and uterine biochemical parameters and consequently on the number of TE. Non-lactating (n=7) and lactating (n=28) Holstein cows were synchronized for estrus and superovulated respectively and were inseminated twice. Uterine bacteriology and cytology and various uterine and serum biochemical parameters were measured at day 7 of estrus cycle (D7, starting day of the SOV protocol) and at the designated day of embryo recovery (DER). Harvested embryos were evaluated according to IETS’s criteria. Superovulated cows produced an average of 7.39 ± 6.22 ova/embryos of which 3.32 ± 4.81 were TE. There were no significant variations of uterine bacteriology and cytology between D7 and DER within the two groups. Serum urea (P=0.0001), E2 (P=0.006); uterine Glu (P=0.002), Ck (P=0.0007), LDH (P<0.0001), TP (P=0.004), P4 (P=0.008), PGFM (P<0.0001) in group I and serum P4 (P<0.0001), PGFM (P<0.0001); uterine LDH (P=0.002), PGFM (P<0.0001) in group II were significantly higher at DER than at D7. At DER, group I was different to group II’ uterine and serum urea (P<0.0001 and P<0.0001), LDH (P<0.0001 and P=0.008), PGFM (P=0.002 and P=0.009), serum P4 (P=0.0002) and uterine TP (P=0.0003). There was no association between uterine bacteriology and cytology and the number of TE. However, TE was positively correlated with serum IGF-1 at D7 (r=0.45; P=0.001) and P4 at DER (r=0.43; P<0.05) and negatively correlated with both serum and uterine PGFM respectively at D7 (r=-0.54; P<0.005 and r=-0.67; P<0.001) and DER (r=-0.48; P<0.01 and r=-0.57; P<0.002). The present results infer that changes following SOV in both serum and uterine secretion may affect the number of TE.
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Ha, Stephanie A. "A Novel Periplasmic Protein involved in the Mannan Chain Elongation Step of Lipomannan and Lipoarabinomannan Biosynthesis in Mycobacterium smegmatis." 2017. https://scholarworks.umass.edu/masters_theses_2/466.
Full textAbstract:
Mycobacteria are atypical bacteria possessing unusual cell envelopes comprised of an outer membrane, covalently linked to an arabinogalacatan-peptidoglycan structure via waxy mycolic acids, in addition to the conventional inner membrane. This thick and highly impermeable cell envelope is a major deterrent to antibiotic treatment of clinically relevant mycobacterial pathogens, including Mycobacterium tuberculosis (Mtb), which infects a third of the world’s population and kills millions each year. Thus, the regulation of mycobacterial cell envelope biosynthesis is of great interest for the development of more effective therapeutics for treating Mtb infections. Using the model organism Mycobacterium smegmatis (M. smegmatis), we identified a novel protein, Spe2, with an unknown role in the biosynthesis of the cell envelope glycolipids lipomannan (LM) and lipoarabinomannan (LAM). Based on the observation that Δspe2 mutants produce truncated LM/LAM, I speculated Spe2 might enhance the elongation of these products. Here, I use biochemical assays to show Spe2 is localized to the periplasm where it can directly interact with the LM/LAM biosynthetic pathway. I further utilize a genetic approach to demonstrate that Spe2 acts at the stage in which the mannosyltransferase MptA mediates periplasmic LM elongation. Moreover, native polyacrylamide gel electrophoresis (PAGE) and co-immunoprecipitation techniques failed to reveal Spe2 protein binding partners. Together, these data suggest Spe2 is a periplasmic protein involved in regulating LM/LAM biosynthesis, perhaps through direct interactions with LM intermediates.
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Hagan, Elsina E. "A Multidisciplinary Approach to Food Safety Evaluation: Hummus Spoilage and Microbial Analysis of Kitchen Surfaces in Residential Child Care Institutions (rcci) in Massachusetts, U.S.A." 2011. https://scholarworks.umass.edu/theses/612.
Full textAbstract:
Food borne illnesses continues to be a public health challenge in the United States (U.S.); an estimated 9.4 million incident cases occurred in 2011. In view of this challenge we conducted two food safety studies; 1) related to product formulation (hummus spoilage challenge study) and 2) evaluating the microbial safety of domestic kitchen surfaces in Residential Child Care Institutions (RCCI pilot study).
Hummus is of Mediterranean origin but is currently eaten globally. This challenge study evaluates a variety of industrial hummus formulations (four in total, differing in pH and/or addition of a preservative (natamycin). Two batches were setup: batch 1; aseptically inoculated hummus with 100 CFU/g fungal isolates and batch 2; uninoculated hummus. Samples of both hummus batches were stored at both 20oC (10 days accelerated testing) and 4oC (84 days recommended temperature testing). Inoculated samples were analyzed for fungus, whiles both fungi and bacteria (standard plate count (SPC) and Lactococci) counts were done for uninoculated samples. Results indicate that accelerated testing inaccurately predicts fungal growth at 4oC in hummus, also fungal growth inhibition requires a pH ≤ 4.0 ± 0.2 and refrigeration.
Limited studies have specifically evaluated the prevalence of pathogenic bacteria in domestic kitchens in the U.S, for this reason we assessed the microbial safety of 6 RCCI locations in MA. Fifteen key food contact surfaces and dish washing sponges, if available at each RCCI facility were assessed for SPC, yeast and molds, total coliform and E. coli, Listeria sp and Salmonella sp. Microbiological assessments were conducted preceding and after a hazard analysis and critical control point (HACCP) food safety training and implementation at each location. Microbial growth varied by surface for each type of microorganism, wet surfaces had higher most probable number (MPN) counts. Compared to dry surfaces, wet surfaces had significantly higher mean total coliform counts. For both E. coli and total coliform, microbial load differed significantly by surfaces sampled (P = 0.0323 and 0.014) respectively. The surface and training interaction effect was highly significant for only E. coli (P = 0.0089). Training overall had no significant effect on reducing the microbial load on kitchen surfaces.
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"Schistosoma haematobium infection and uropathies in hyperendemic and hypoendemic communities in the extreme northern province of Cameroon: Ultrasonographic, chemical reagent strip and bacteriologic correlates of morbidity." Tulane University, 1990.
Find full textAbstract:
The present study, carried out in heavily infected communities in northern Cameroon, was designed to determine the prevalence of urinary tract lesions and their association with Schistosoma haematobium (S.h.) infection and to assess the validity functional, parasitologic and bacteriologic parameters in the identification of persons at high risk for these lesions Schistosoma haematobium infection status, morphologic changes in the urinary tract (as detected by ultrasonography), functional and bacteriologic changes in the urine were determined on 980 persons selected randomly after stratification by age, sex and intensity of infection. Inclusion of egg-free controls from a hypoendemic area further facilitated measurement of association of lesions with bilharzial infection Lesions of the lower urinary tract were very frequent in the study population living in hyperendemic communities. Bladder wall hypertrophies (78%), bladder wall irregularities (67%) and bladder wall tumor/s (34%) were observed. History of persistent hematuria, level of current S.h. egg output, and massive proteinuria ($<$99 mg/dl) were the best predictors of these lesions. Large bladder tumors were, however, also strongly associated with excretion of nitrites (bacterial metabolites) in the urine and/or presence of significant bacteriuria. On the other hand, S.h. infection was associated with increased prevalence of positive excretion of urinary nitrites (14% in infected versus 2% in controls) and with increased risk for bacteriuria among children 5-14 year old (8% in infected versus 3% in the controls) Congestion of the upper urinary tract (UUT) was less frequent (4% for Hydroureter and 14% for hydronephrosis) and more confined to bilharzian patients (Less than 1 percent of controls were affected). The group at the highest risk included persons with combined LUT hyperplasia and evidence of bacterial superinfection, for whom prevalence of UUT congestion was more than 10 times higher than in controls Pathologic changes in LUT or UUT were very rare in persons with neither past history of chronic hematuria nor with current S.h. infection, most of whom were found in the control group from the hypoendemic area Observation of circadian variation in both S.h. urinary egg output and related functional changes (hematuria, proteinuria) have lead field investigators to focus their attention on midday voided urine. The present study has shown that testing first morning urine specimens could provide additional information (nitrite and bacteriuria) shown to be of high predictive value for severe uropathies. (Abstract shortened by UMI.)acase@tulane.edu