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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
261

Identification & molecular characterisation of a novel recA from Mycobacterium tuberculosis

Nair, Shamila 13 July 2017 (has links)
No description available.
262

From care inside the laboratory to the world beyond it: a multispecies ethnography of TB science towards growing a decolonised science in South Africa

Shain, Chloë-Sarah 19 April 2023 (has links) (PDF)
This anthropological research began with curiosity about human relationships with microbes. Inside the contained environment of a Biosafety Level 3 laboratory at a South African university-based tuberculosis research division, the fieldwork focused on the relationships between scientists and Mycobacterium tuberculosis − the pathogenic bacterium that causes the disease tuberculosis (TB). These deadly bacteria were cared for and nurtured by women scientists. This care extended to the cells and various species with which they worked. Moreover, this care moved beyond the scope of their immediate scientific research projects and well beyond the laboratory. Care was also central to how the participants conducted their scientific research and themselves in the world. This long-term, qualitative ethnographic research weaves together many layers of care in biomedical scientific research, highlighting that scientific research is a deeply personal, caring and subjective practice. The natural and the social are not − and can never be − mutually exclusive. Boundaries between mind/body, subject/object, human/nonhuman, researcher/researched, subjectivity/objectivity and science/society are porous. Acutely aware of the socio-political moment in which this research was embedded, these findings are put into conversation with South African student calls to decolonise science that emerged alongside the #RhodesMustFall student movement. In particular, the focus is on a 2016 meeting about decolonising science at the University of Cape Town where students argued for connection between the university and the community, science and society and the world of academia and the world of Africans. Implicit was the need for science to be relevant to Africans and deeply complex African social formations and problems. The care by women scientists that was observed inside the laboratory and beyond it speaks volumes to cultivating a more caring science and caring institutions of science that connect the laboratory to the world in which it exists in meaningful, relevant and impactful ways. I demonstrate how the participants embodied a decolonised science, and that what they cared about and how they acted upon those cares could serve as important guides for decolonising science and scientific institutions. This research provides important contributions to the field of science and technology studies (STS), to anthropological research on TB and to the conversation on decolonising science in South Africa.
263

The immunological role of cell wall components from diverse Mycobacterium tuberculosis clinical isolates in regulating HIV-1 replication in human macrophages

Ndengane, Mthawelanga 11 September 2023 (has links) (PDF)
Human immunodeficiency virus type 1 (HIV-1) and Mycobacterium tuberculosis (Mtb) coinfection remains a major global health threat. Both pathogens synergistically drive pathogenesis of the other. The risk of developing active tuberculosis (TB) is increased in people living with HIV-1, even in those receiving antiretroviral therapy (ART), whilst TB was responsible for 15 % of HIV-related deaths in 2020. Mtb co-infection increases the likelihood of transcriptionally activating HIV-1 replication potentially due to bioactive Mtb lipids engaging macrophage surface receptors, thus triggering signaling pathways which activate human transcriptional factors (hTF) and production of inflammatory cytokines capable of activating HIV-1 transcription. This work investigated the hypothesis that clinical Mtb strains with single nucleotide polymorphisms (SNP) in lipid-metabolizing genes, required for cell wall lipid biosynthesis, differentially affect HIV-1 replication and human macrophage inflammatory response during Mtb-HIV-1 co-infection in vitro. Monocyte derived macrophages (MDM) were the predominant model used to investigate this phenomenon. Infections, in the presence or absence of HIV-1 co-infection, were performed using either lineage 2 or lineage 4 clinical strains with non-synonymous SNP in polyketide synthase 2 (pks2) required for sulfolipid 1 (SL-1) biosynthesis and compared to control infections using phylogenetically close clinical strains without the SNP of interest and canonical lineage 2 and 4 laboratory strains (H37RvP1939/T605, CDC1551WT and HN878WT). Secreted cytokines and chemokines were measured in supernatant (SN) by Luminex. The effect of Mtb on HIV-1 viral production was assessed by measuring HIV-1 Gag p24 in the SN of co-infected MDM or SN of HIV-1 infected MDM incubated with conditioned media from Mtb-infected MDM. The influence of Mtb on HIV-1 transcriptional activity was measured using a transgenic cell line (TZM-bl) with Luciferase reporter under HIV-1 long terminal repeat (LTR) expression. The impact of incubating TZM-bl cells in Mtb-induced conditioned media before or after HIV-1 infection was assessed. One pair of phylogenetically close clinical strains with and without a pks2 SNP of interest (EX30Q1939/A605 and MRC16P1939/A605) with interesting lipid and inflammatory phenotypes, and H37RvP1939/T605 as a lineage 4 control, were subject to single nucleotide mutagenesis using recombineering to either revert SNP of interest to match the alleles of H37Rv or introduce the SNP of interest into the control strains. The wild-type and mutant strains were used in a trans-well assay to infect MDM in the presence of HIV-1 co-infection in the top chamber, while simultaneously mimicking the bystander effect of cytokine-mediated HIV-1 regulation in the bottom chamber which was only infected with HIV-1. Results demonstrate there was increased cytokine production by MDM infected with MRC16P1939/A605 in both the presence and absence of HIV-1 co-infection compared to its phylogenetically close paired strain EX30 Q1939/A605. The data shows that there was no difference in LTR activity in TZM-bl cells co-incubated with inflammatory environment between the strains of interests, however co-incubation of TZM-bl cells with Mtb-induced inflammatory environment generally increased LTR activity during HIV-1; a proxy for HIV-1 replication. In the trans-well co-infection assay, a significant positive association between production of HIVp24 and secretion of CCL2 was observed, whilst IL-1β secretion showed a significant negative relationship with the production of HIVp24, with donor variability in baseline cytokine production also associated with the extent of HIVp24, CCL2, IL-1β and IL-8 production. Introduction of the pks2 T605A SNP into H37RvP1939/T605 and reversion in EX30Q1939/A605T significantly modified their inflammatory phenotype. Together these results support the hypothesis that Mtb clinical stains with genetic variation in cell wall lipid biosynthesis impacts the inflammatory milieu and, subsequently, HIV-1 replication during co-infection. The outcome of Mtb-HIV co-infection is therefore not homogenous but contingent on the phenotype of infecting Mtb strain and individual.
264

Catching a glimpse: the visualization of Mycobacterium tuberculosis from TB patient bioaerosols

Dinkele, Ryan 08 June 2023 (has links) (PDF)
Transmission between hosts is crucial for the success and survival of the obligate human pathogen and aetiological agent of tuberculosis (TB), Mycobacterium tuberculosis (Mtb). Despite this, little is known about how and when Mtb is aerosolized nor the key metabolic and morphological determinants driving successful transmission. To address these knowledge gaps, my doctoral research sought to develop a microscopic method for the detection of aerosolized Mtb following liquidcapture within the respiratory aerosol sampling chamber (RASC). This was achieved through the combination of the mycobacterial cell wall probe, 4-N,Ndimethylamino-1,8-naphthalimide-trehalose (DMN-tre), with the arraying of bioaerosol samples on bespoke nanowell devices amenable to fluorescence microscopy. With this method, a median of 14 live Mtb bacilli (range 0-36) were detected in 90% of confirmed TB patients following 60 minutes of bioaerosol sampling. Three distinct DMN-tre staining patterns were identified among aerosolized Mtb, strongly suggestive of metabolic heterogeneity. Moreover, a low proportion of patients produced Mtb in small clumps. These observations highlight the advantages of using microscopy over conventional culture- or molecular-based techniques for probing the metabolic and morphological characteristics of aerosolized Mtb. Applying this method in a second study, we sought to understand how and when Mtb is aerosolized. To this end, we aimed to compare the aerosolization of Mtb and total particulate matter from patients with TB during three respiratory manoeuvres: tidal breathing (TiBr), forced vital capacity (FVC), and cough. Although total particle counts were 4.8-fold greater in cough samples than either TiBr or FVC, all three manoeuvres returned similar rates of positivity for Mtb. No correlation was observed between total particle production and Mtb count. Instead, for total Mtb counts, the variability between individuals was greater than the variability between sampling manoeuvres. Finally, when modelled using 24-hour breath and cough frequencies, our data indicate that TiBr might contribute more than 90% of the daily aerosolized Mtb among symptomatic TB patients. Assuming the number of viable Mtb organisms detected provides a proxy measure of patient infectiousness, this method suggests that TiBr is a significant contributor to TB transmission. In developing a novel platform for the detection of aerosolized Mtb, this work has suggested the need to re-examine old assumptions about Mtb transmission.
265

MOLECULAR ANALYSIS OF MYCOBACTERIUM COMMUNITY SHIFTS IN SOIL DURING THE BIODEGRADATION OF POLYCYCLIC AROMATIC HYDROCARBONS

Cheung, Pui Yi 11 October 2001 (has links)
No description available.
266

The foraging ecology of banded mongooses (Mungos mungo): Epidemiological and human-wildlife conflict implications

Laver, Peter Norman 11 June 2013 (has links)
Free-ranging banded mongooses (Mungos mungo) in northeastern Botswana are infected by a novel Mycobacterium tuberculosis complex pathogen, M. mungi, which putatively infects mongooses through lesions in the skin (often the planum nasale) from an environmental reservoir. To understand the epidemiology of the yearly and highly seasonal outbreaks of M. mungi in this population of banded mongooses, researchers need to understand what factors influence banded mongoose exposure to M. mungi and banded mongoose susceptibility to M. mungi infection. Researchers have no baseline data on the behavioral ecology of this population of banded mongooses - such as home range dynamics, denning ecology, movement ecology, and foraging ecology, all of which may play a role in banded mongoose exposure to M. mungi. Further, researchers have highlighted the potential role of prolonged elevations of glucocorticoids in impairing cell-mediated immunity, which would play a significant role in determining susceptibility to a mycobacterium such as M. mungi, however, researchers have no data on the endocrinology of banded mongooses. Finally, researchers have not detected M. mungi infection in any other population of banded mongooses. Our study population has a gradient of troops (social groups) that vary from troops with extremely close association with humans in a town, to troops associated with humans at tourist lodges within the Chobe National Park, to troops with no discernible association with humans within the national park and surrounding forest reserve. Researchers have few data on how synanthropy (living with humans) affects banded mongoose behavioral ecology and no data on how synanthropy affects banded mongoose endocrinology. Researchers do not know whether or how the high level of synanthropy in this population of banded mongooses plays a role in the epidemiology of M. mungi outbreaks. Thus, we document here some aspects of banded mongoose home range dynamics, movement metrics, denning ecology and foraging behavior for our study population in northeastern Botswana. We present a novel method for screening data from global positioning system (GPS) collars for large measurement error and we present a detailed home range study. We also document the spatio-temporal dynamics of glucocorticoid production among several banded mongoose study troops across our study site, using a non-invasive assay for fecal glucocorticoid metabolites, which we validated and also present here.  We tested to see which factors, including nutritional limitation, predation risk, and reproduction (and associated competition, agonistic encounters, and predation), best explained the variation in glucocorticoid production among our study troops over several years. We found that the metrics traditionally used to screen data from GPS collars, horizontal dilution of precision (HDOP) or fix dimension (2-D or 3-D), performed poorly relative to a new screening metric that we propose, the estimated elevation error (EEE). We propose that researchers use our screening method, which combines test data and a model-averaging information-theoretic framework that uses a priori candidate models of telemetry measurement error. Although we recommend including EEE in a priori candidate models, it may not describe telemetry error in other systems as well as it did in our own. Banded mongooses in our study population formed troops of a median of 13 adults (IQR: 11 to 21 adults) and these troops used home ranges of a median of 68 ha (IQR: 39 to 134 ha) with core areas of a median of 15 ha (IQR: 9 to 28 ha). These cores (statistically-clumped space use) occurred at a median volume contour of 66 % (IQR: 58 to 71 %). Synanthropic troops showed more clumped area use than apoanthropic troops (those living away from humans). Synanthropic troops also used man-made structures for den sites in SI{81}{percent} of nights, fed from refuse sites in 13 % of foraging observations, and drank from anthropogenic water sources in 78 % of drinking observations. From our conducted adrenocorticotropic hormone challenge, we detected valid increases in fecal glucocorticoid metabolite concentrations in mongoose feces using our four tested enzyme-immunoassays. An 11-oxoetiocholanolone assay detecting 11,17-dioxoandrostanes (11,17-DOA) performed best. Using this assay, we detected expected decreases in fecal glucocorticoid metabolite concentrations 48 h after administering dexamethasone sodium phosphate. We also validated this assay using biological events as challenges, in which captive mongooses showed higher fecal glucocorticoid metabolite concentrations during reproductive activity, agonistic encounters, and depredation events. The time delay of fecal glucocorticoid metabolite excretion approximately corresponded with food transit time, at a minimum of approximately 24 h. Fecal glucocorticoid metabolite metabolism was minimal up to 8 h post-defecation. Reproduction and its associated challenges dramatically increased glucocorticoid production, which otherwise remained low and stable in a captive troop with a constant food supply and lowered predation risk. Variation in glucocorticoid production in free-ranging banded mongooses was best explained by food limitation as described by current nutritional limitation (proportion of fecal organic matter), recent rainfall (which increases soil macrofauna availability), and access to concentrated anthropogenic food resources. Habitat differences in soil macrofauna density and reproductive events also explained variation in glucocorticoid production in free-ranging mongooses, but to a much lower degree. Predation risk, as measured by canopy cover (escape from aerial predators) and group size (decreased per capita vigilance) explained very little of the variation in glucocorticoid production. In the late dry season, banded mongooses in our population may face a "perfect storm" of nutritional limitation, agonistic encounters at concentrated food resources, aggressive evictions, estrus, competition for mates, parturition, and predation pressure on pups. We suspect that this prefect storm may push glucocorticoid responses into homeostatic overload and may impair cell-mediated immunity in banded mongooses. / Ph. D.
267

Gene expression changes in macrophages infected with pathogenic M. tuberculosis and non-pathogenic M. smegmatis and M. bovis BCG

Mpongoshe, Vuyiseka 04 1900 (has links)
Thesis (MScMedSc)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: The current anti-TB drugs have had success in decreasing the number of deaths caused by TB, however, this success is limited by the emergence of drug resistant TB strains. Therefore, a novel TB therapy that limits the development of resistance has become necessary in an attempt to effectively control TB. The anti-TB drugs directly target mycobacterial enzymes, and potentiate the development of this resistance, and have therefore provided the rationale for this study. The aim was therefore to identify host macrophage genes that affect M. tb intracellular survival. The proposed alternative anti-TB therapy potentially involves the application of RNA interference (RNAi) and RNA activation (RNAa) biological processes that will target host genes, thereby inducing an indirect bactericidal effect. We hypothesized that macrophage genes that are differentially expressed by pathogenic and non-pathogenic mycobacterial species may be important in the regulation of M. tb intracellular survival. The lipid-rich mycobacterial cell wall is implicated in the excessive clumping of the mycobacterial cells in liquid culture. In order to minimize this, Tween 80 detergent was supplemented (mycobacteriaT). However, due to substantial evidence emphasising the detrimental effects of Tween 80 on the mycobacterial cell wall, mycobacteria were also cultured without Tween 80 (mycobacteriaNT), in order to investigate if the perturbed mycobacterial cell wall induced by Tween 80 affects the transcriptional response of macrophages. We endeavoured to develop a new method to culture mycobacteria without Tween 80 that will still generate single cells. We further hypothesized that the macrophage gene expression profile induced by mycobateriaNT differs from the response induced by mycobacteriaT. Differentiated THP-1 (dTHP-1) cells were infected with pathogenic and non-pathogenic mycobacteria (for 3 h, 24 h and 48 h with M. tb and M. bovis BCG, and 3 h and 8 h with M. smegmatis) cultured in the presence or absence of Tween 80. The expression of 12 macrophage genes, selected based on their involvement in the phagocytic pathway and autophagy, as well as their general involvement in the immune response, was determined by qRT-PCR and further analysed on the REST programme. The expression of each target gene was normalised relative to the expression of the reference gene (Beta actin). We observed that out of the 12 genes, TLR7 and VAMP7 were consistently downregulated in dTHP-1 cells infected with M. tbNT and upregulated in dTHP-1 cells infected with M. smegmatisNT. Their response to M. bovis BCG was inconsistent and not significantly different, and therefore could not be interpreted. Furthermore, CCL1 was upregulated by all the mycobacterial species. However, its expression was more pronounced in response to mycobacteriaNT, when compared to mycobacteriaT. Differential gene expression of TLR7 and VAMP7 in response to pathogenic and non-pathogenic mycobacteriaNT suggests that these 2 genes may be potential targets for RNAa-based anti-TB therapy, even though we could not conclude whether their response was specific to macrophages. In addition, the observed difference in the expression of CCL1 induced by mycobacteriaNT, compared to mycobacteriaT suggests that the perturbation caused by Tween 80 on the mycobacterial cell wall most likely affected the response of macrophages to infection with mycobacteria. Furthermore, this study has demonstrated a feasible method by filtration to generate single cells from mycobacteriaNT, which should be considered for future mycobacterial infection studies. / AFRIKAANSE OPSOMMING: Die huidige anti-tuberkulose middels se sukses lê daarin dat dit die aantal sterftes verminder maar hierdie sukses word weer beperk met die ontstaan van middel-weerstandige M.tb stamme. Daarom is nuwe middels nodig wat die ontwikkeling van middel-weerstandigheid beperk in ʼn poging om effektiewe TB behandeling te bewerkstellig. Anti-tuberkulose middels teiken hoofsaaklik mycobakteriële ensiemsisteme en ontlok sodoende weerstandigheid in M.tb stamme en dit vorm die rasionale vir hierdie studie. Die doel was om gasheer makrofaag gene te identifiseer wat M.tb oorlewing intrasellulêr bewerkstellig. Die voorgestelde alternatiewe anti-TB behandeling sal dan behels die toepassing van RNA intervensie (RNAi) en RNA aktivering (RNAa) tegnologie wat gasheer selgene teiken (inaktiveer) en sodoende ʼn bakterisidiese respons induseer. Die kanse is skraal dat mycobakterieë weerstandigheid sal kan ontwikkel onder hierdie omstandighede. Ons hipotetiseer dus dat makrofaag gene wat differensieel uitgedruk word deur patogeniese en nie-patologiese mycobakteriële spesies belangrik mag wees vir die oorlewing van M.tb intrasellulêr. Die lipiedryke selwand van mycobakterieë word geïmpliseer in die oormatige sameklomping van die bakterieë in vloeistofkulture. Om hierdie effek te minimaliseer word Tween 80 normaalweg tot die medium gevoeg (mycobakterieëT). Maar weens genoegsame bewyse dat Tween-80 die selwand van bakterieë nadelig beïnvloed, is mycobakterieë ook in die afwesigheid van Tween 80 gekultureer (mycobakterieëNT) om te bepaal of die nadelige effek van Tween 80 op die selwand die transkripsionele respons in makrofage beïnvloed post-infeksie. Dit was daarom ook ons doelstelling om ʼn nuwe tegniek te ontwikkel om mycobakterieë te kultureer in die afwesigheid van Tween 80 wat ook enkelselle sal genereer vir beter gekontroleerde makrofaag infeksie. Ons hipotetiseer ook verder dat makrofaag geenuitdrukking-profiele verskil afhangende of infeksie gedoen is met mycobakterieë wat in die afwesigheid of teenwoordigheid van Tween 80 gekultureer is. Gedifferensieerde THP-1 (dTHP-1) was geïnfekteer met patogeniese en nie-patogeniese mycobakterieë (vir 3 h, 24 h en 48 h met M.tb en M.bovis BCG, en 3 h en 8 h met M.smegmatis) gekultureer in die teenwoordigheid en afwesigheid van Tween 80. Die uitdrukking van 12 makrofaag gene, geselekteer op grond van hul betrokkenheid in die fagositose meganisme en in outofagie asook hul betrokkenheid in die immuunrespons, is gekwantifiseer met qRT-PCR en daaropvolgens geanaliseer met die REST-program. Die uitdrukking van elke geen is genormaliseer relatief tot die uitdrukking van die verwysingsgeen (Beta actin). Daar is bevind dat van die 12 gene, TLR7 en VAMP7 deurlopend afgereguleer was in dTHP-1 selle geïnfekteer met M.tbNT en opgereguleer was in dTHP selle geïnfekteer met M.smegmatisNT. Selrespons met M.bovis BCG was onbeduidend en derhalwe kon geen gevolgtrekking hier gemaak word nie. Ook, CCL1 was opgereguleer met infeksie deur enige van die mycobakteriële spesies, maar CCL1 se uitdrukking was groter in respons tot mycobakterieëNT wanneer vergelyk word met respons tot mycobakterieëT. Differensiële geenuitdrukking van TLR7 en VAMP7 in respons tot patogeniese en nie-patogeniese mycobakterieëNT impliseer dat hierdie twee gene potensiële teikens kan wees vir RNAa-gebaseerde anti-TB behandeling, alhoewel ons nie kon beslis of hierdie respons spesifiek vir makrofage was nie. Ook, die verskille waargeneem in die uitdrukking van CCL1 geïnduseer deur mycobakterieëNT, vergeleke met mycobakterieëT, impliseer dat die steuring in die selwand veroorsaak deur Tween 80, heelwaarskynlik die respons van die makrofaag beïnvloed het. Hierdie studie beskryf ook ʼn filtrasiemetode om enkele mycobakteriële selle te genereer wat oorweeg moet word by toekomstige mycobakteriële infeksiestudies.
268

Etude In vitro de Phospholipases mycobactériennes impliquées dans la virulence / In vitro study of micobactérial Phospholipases involved in virulence

Bakala n'goma, Jean-claude 26 March 2010 (has links)
Les phospholipases et en particulier les phospholipases C sont d'importants facteurs de virulence chez de nombreuses bactéries pathogènes (C. perfringens, B. Cereus et P. aeruginosa). Cependant, peu de choses sont connues sur l'implication de ces enzymes dans le processus de virulence des mycobactéries. Bien que l'étude des mutants des phospholipases C de M. tuberculosis dans un modèle d'infection chez la souris ait permis de proposer une implication de ces protéines dans la virulence de ce bacille, leurs propriétés biochimiques, leur mode d'action et leur rôle physiologique exact restent à élucider. Ce manque de données biochimiques sur les phospholipases mycobactériennes peuvent être attribuée à la difficulté à produire et à purifier des quantités importantes de ces enzymes. Dans le but de mieux caractériser le rôle physiologique des phospholipase mycobactériennes, l'objectif de ma thèse a été de mettre au point des conditions d'expression hétérologue permettant la production des phospholipases C mycobactériennes recombinantes (rPLC) dans différents systèmes d'expression (E. coli, Pichia pastoris et baculovirus/cellules d'insectes). Ces systèmes d'expression n'ayant pas donné des résultats satisfaisants, nous avons développé une méthode efficace d'expression de ces protéines en utilisant M. smegmatis.Ce système d'expression nous a permis de produire et de purifier les quate PLC (PLC-A, PLC-B, PLC-C et PLC-D) de M. tuberculosis et la PLC de M. Abscessus sous forme soluble et active. Nous avons pour la première fois montré que ces protéines purifiées avaient un effet cytotoxique sur les macrophages de souris en culture mais ne présentaient aucune activité hémolytique. en utilisant des marquages radioactifs, nous avons confirmé que l'effet cytotoxique observé était lité à l'hydrolyse des phospholipides des membranaires des cellules hôtes. Pour la première fois, nous avons pu confirmer que ces PLC sont directement impliquées dans le processus d'infection et de virulence.Un autre aspect de mon travail de thèse a concerné l'étude de deux autres protéines sécrétées par M. tuberculosis appartenant à la famille des cutinases : la Rv1984c et la Rv3452. Après les avoir produites et purifiées chez E. Coli, nouq avons montré que malgré ces deux protéines présentent 50% d'identité de séquence en acides aminés, elles ont des spécificités de substrat différentes et probablement un rôle physiologique différent. La Rv1984c est une lipase capacle d'hydrolyser des lipides à chaines moyennes, alors que la Rv3452 est une phospholipase de type A2 et est capable d'induire la lyse de macrophage de souris en culture. / Phospholipases, particularly phospholipases C, are important virulence factors in several pathogenic bacteria (C. perfringens, B. cereus, L. monocytogenese and P. aeruginosa). However, little is know on the involvement of thses enzymes in mycobacteria pathogenesis. Although study on M. tuberculosis phospholipases C mutants in a mouse aerosol model of infection gave rise to the contribution of these proteins in virulence process, but their exact biochemical properties, mechanism of action and physiological role remain to be elucidated. This lack of data on mycobacterial phospholipases is mainly due to the difficulty to produce and purify these enzymes in large scale.With the aim to better characterise the physiological role of mycobacterial phospholipases, the main challenge of my thesis was to develop an efficient method for expression and purification of recombinant mycobacterial phospholipases C. Since no satisfactory results have been obtained with standard expression systems (E. coli, Pichia pastoris and baculovirus / insect cells), we develop a robust expression technique for these proteins using M. smegmatis as expression system.This allowed us to produce and purify all four PLC (PLC-A, PLC-B, PLC-C and PLC-D) of M. tuberculosis and the PLC of M. abscessus in soluble and active form. For the first time, we have show, that purified proteins have cytotoxic effect on mouse macrophages but have not haemolytic activity. Using radiolabelled lipids, we have confirmed that this first direct evidence that PLC are involved in infection and virulence processes. Another aspect of my thesis work concerned the study of two other secreted proteins of M. tuberculosis belonging to the cutinase family : the Rv 1984c ant the Rv3452. Recombinant proteins obtains in E. coli were found to have distinct substrate specificities and most likely distict physiological role, despite showing 50% amino acids sequence identity. Rv1984c is a lipase and is able to hydrolyse lipids with medium chains lengthn whereas Rv3452 is type A2, phospholipase and i able to induce macrophage lysis.
269

INFECÇÃO PELO Mycobacterium tuberculosis ENTRE OS PROFISSIONAIS DA EQUIPE DE ENFERMAGEM, EM UM HOSPITAL DE DOENÇAS INFECCIOSAS, GOIÂNIA - GO. / Mycobacterium tuberculosis INFECTION AMONG PROFESSIONALS NURSING TEAM, AT A HOSPITAL FOR INFECTIOUS DISEASES, Goiânia - GO.

LOPES, Lilian Kelly de Oliveira 23 February 2006 (has links)
Made available in DSpace on 2014-07-29T15:04:41Z (GMT). No. of bitstreams: 1 dissertacao lilian.pdf: 281825 bytes, checksum: 488c9e5d58cb1a521b28a8592ee6de50 (MD5) Previous issue date: 2006-02-23 / According to the World Health Organization (WHO), an hundred million of individuals are infected by M. tuberculosis, annually. Health care workers play an important role to control of tuberculosis, but they are also at high risk for this infection. Then, the objectives of the present study were to evaluate the prevalence of M tuberculosis infection in nursing professionals from the Tropical Diseases Hospital in Goiânia City, State of Goiás, to analyze the factors associated to tuberculin skin test (TST) positivity and to determine the TB infection incidence density in susceptible professionals Initially, the prevalence and factors associated to TST were investigated in 128 eligible individuals. Further, susceptible professionals (n=32) were followed up during three years (2001-2004) to detect TST conversion. Of the total individuals investigated, 69.5% (IC 95%: 60.7-77.2) were positive to TST. Two occupational factors were independently associated to skin test positivity: duration of profissional activity longer than 5 years (Adjustd OR = 6.3; 95% CI: 1.5-26.2) and occupational contact with a person with pulmonary TB ≤ 2 years (Adjusted OR = 12.2; 95% CI: 1.2-106.3). Seven profissionals showed tuberculinic conversion during the three years of follow up, and an incidence density of 11.5 new conversions to 100 persons-year was detected. All of them had taken care of patients during the period of the study. Two individuals developed tuberculosis disease. The data of this study ratify the high risk of tuberculosis in nursing team, and highlight the importance of this infection as an occupational disease to nursing professionals of our region. / De acordo com a Organização Mundial de Saúde (OMS), cem milhões de pessoas são infectadas pelo M. tuberculosis, a cada ano. Os profissionais de saúde são importantes para o controle da tuberculose, mas também um grupo de risco elevado para esta infecção. Assim, o presente estudo teve como objetivos avaliar a prevalência da infecção causada pelo M. tuberculosis em profissionais de enfermagem de uma instituição especializada em doenças infecciosas, em Goiânia Go, analisar os fatores associados à positividade à prova tuberculínica nesta população e determinar a densidade de incidência da infecção pelo M. tuberculosis, nos profissionais susceptíveis. Inicialmente, verificou-se a prevalência e os fatores associados à positividade à PT. A seguir, os profissionais suscetíveis à infecção (n=32) foram acompanhados, por três anos (2001-2004), para detecção de conversão tuberculínica. Do total de profissionais investigados, 69,5% (IC 95%: 60,7- 77,2) foram positivos à PT. Dois fatores ocupacionais foram independentemente associados à positividade à PT: tempo de atividade profissional > 5 anos (OR ajustado = 6,3; IC 95%: 1,5-26,2) e último contato laboral com alguém com TB ≤ 2 anos (OR ajustado = 12,2; IC95%: 1,2-106,3). Sete profissionais apresentaram viragem tuberculínica, resultando em uma densidade de incidência de 11,5 novas conversões por 100 pessoas/ano. Todos desenvolviam atividades assistenciais, durante o período do estudo. Duas profissionais desenvolveram tuberculose doença. Os resultados, deste estudo, ratificam o elevado risco de tuberculose nos profissionais de enfermagem, e evidenciam a importância desta infecção como doença ocupacional para equipe de enfermagem de nossa região.
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A Novel Antigen From Mycobacterium Bovis BCG : Biochemical And Immunological Studies

Pawar, Santosh N 12 1900 (has links) (PDF)
No description available.

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