Mycobacterium tuberculosis has caused a resurgence in pulmonary disease in both developed and developing countries in recent times, particularly amongst people infected with the human immunodeficiency virus. The disease has assumed epidemic proportions in South Africa and in the Eastern Cape Province in particular. Of further concern is the isolation of increasing numbers of multiply drug resistant strains. Knowledge of the genetic capability of this organism is essential for the successful development of novel antibiotics and vaccines in an attempt to bring the global pandemic under control. Measurement of the genetic diversity of the organism may significantly contribute to such knowledge, and is of vital importance in monitoring epidemics and in improving treatment and control of the disease. This will entail answering a number of questions related to the degree of genetic diversity amongst strains, to the difference between urban and rural strains, and between drug resistant and drug sensitive strains, and to the geographical distribution of strains. In order to establish such baseline information, RAPD profiling of a large population of isolates from the western and central regions of the Eastern Cape Province was undertaken. A smaller number of drug resistant strains from a small area of KwaZulu-Natal were also analysed, with a view to establishing the genetic difference between strains from the two provinces. Cluster analysis, analysis of molecular variance and Geographical Information Systems technology were used to analyse the RAPD profiles generated. An unexpectedly high degree of genetic diversity was detected in strains from both provinces. While no correlation was seen between genetic diversity and either urban-rural situation or geographical location, a small degree of population structure could be correlated with drug resistance in the Eastern Cape. Furthermore, a significant degree of population structure was detected between strains from the two provinces, although this was still within the parameters for conspecific populations. Future work is necessary to further characterise strains from rural areas of both provinces, as well as from the eastern region of the Eastern Cape in an attempt to pinpoint the cause of the separation of the provincial populations.

Lic.-avh. (sammanfattning) Stockholm : Karolinska institutet, 2009.

Thesis (MSc)--Stellenbosch University, 2002. / ENGLISH ABSTRACT: Please see fulltext for abstract. / AFRIKAANSE OPSOMMING: Sien asb volteks vir opsomming.

Thesis (MScMedSc)--Stellenbosch University, 2015. / ENGLISH ABSTRACT: Drug resistance in Mycobacterium tuberculosis is an increasing global problem. Drug resistance is mostly caused by single nucleotide polymorphisms (SNPs) within the bacterial genome. This observed increase in global incidence of drug resistant tuberculosis (TB) has sparked the search for new anti-TB drugs and the repurposing of drugs that are currently used against other organisms or species of mycobacteria. One such repurposed drug, clofazimine (CFZ), is currently used for the treatment of leprosy, caused by Mycobacterium leprae. The mechanism of action of CFZ is not clear, but it is hypothesized that CFZ is reduced by a mycobacterial type II NADH oxidoreductase (NDH-2). The reduction of CFZ drives the production of reactive oxygen species (ROS) which is toxic to the pathogen. The aim of this study was to elucidate the mechanism of CFZ resistance. Towards this aim, spontaneous in vitro CFZ resistant mutants were selected, characterized and whole genome was used identify SNPs which may cause CFZ resistance. Mutations were identified in a transcriptional regulator encoded by Rv0678, fatty-acid-AMP ligase, or FadD28 (Rv2941) and glycerol kinase or GlpK (Rv3696c). Mutations in Rv0678 have previously been shown to play a role in both CFZ resistance and bedaquiline (BDQ) cross-resistance, while no link has been found between CFZ resistance and mutations in fadD28 and glpK. The novel, non-synonymous SNP identified in Rv0678 resulted in the replacement of an alanine residue with threonine at codon 84, which is located in the DNA binding domain. Virtual modelling of the mutated Rv0678 protein showed that the A84T mutation may influence DNA binding, possibly due to its proximity to the DNA binding domain. This mutation caused a change in hydrophobicity, which may influence binding to DNA. Previous studies showed that mutations in Rv0678 resulted in the upregulation of mmpL5, a putative efflux pump. However, the mechanism whereby CFZ resistance occurs via increased abundance of this efflux pump in the cell wall is not clear and needs further investigation. The cross-resistance between CFZ and BDQ, caused by mutations in Rv0678, is of concern and may influence the planning of anti-TB drug regimens for the future. The roles of the other two mutations identified in this study in CFZ resistance is also not clear and requires further investigation. Finally, the findings of this study support the role of Rv0678 in CFZ resistance thereby suggesting that this gene could be useful as a diagnostic marker to test for CFZ resistance in clinical isolates. / AFRIKAANSE OPSOMMING: Middelweerstandigheid in Mycobacterium tuberculosis is 'n wêreldwye toenemende probleem. Middelweerstandigheid word meestal veroorsaak deur enkel nukleotied polimorfismes (SNPs) in die bakteriële genoom. Hierdie toename in middelweerstandige tuberkulose (TB) het gelei tot die soektog na nuwe anti-TB-middels en die alternatiewe aanwending van middels wat tans teen ander organismes of spesies van mikobakterieë gebruik word. Een so 'n alternatiewe middel, clofazimine (CFZ), word tans gebruik vir die behandeling van melaatsheid wat veroorsaak word deur Mycobacterium leprae. CFZ se meganisme van werking is nie duidelik nie, maar dit word vermoed dat CFZ gereduseer word deur 'n mikobakteriële tipe II NADH oksidoreduktase (NDH-2). Die reduksie van CFZ dryf die produksie van reaktiewe suurstof spesies wat giftig is vir die patogeen. Die doel van hierdie studie was om die meganisme van CFZ weerstandigheid te ondersoek. Om hierdie doel te bereik was spontane in vitro CFZ weerstandige mutante gekies, gekarakteriseer en heel genoom volgorde bepaling is gebruik om SNPs te identifiseer wat CFZ weerstandigheid veroorsaak. Mutasies in Rv0678, 'n transkripsie reguleerder, vetsuur-AMP ligase, of FadD28 (Rv2941) en gliserol kinase of GlpK (Rv3696c) geïdentifiseer. Dit is al voorheen gevind dat mutasies in Rv0678 ‘n rol speel in beide CFZ weerstandigheid en bedaquiline (BDQ) kruis-weerstandigheid, terwyl geen verband gevind is tussen CFZ weerstandigheid en mutasies in fadD28 en glpK nie. Die nuwe, nie-sinonieme SNP, geïdentifiseer in Rv0678 het gelei to die vervanging van 'n alanien aminosuur met treonien by kodon 84, wat geleë is in die DNS bindings domein. Virtuele modellering van die gemuteerde Rv0678 proteïen het getoon dat die A84T mutasie DNS binding moontlik kan beïnvloed, as gevolg van sy nabyheid aan die DNS bindings domein. Hierdie mutasie veroorsaak 'n verandering in die hidrofobiese natuur, wat DNS binding kan beïnvloed. Vorige studies het getoon dat mutasies in Rv0678 lei tot die opregulering van mmpL5, 'n waarskynlike uitvloei pomp. Die meganisme waardeur CFZ weerstandigheid veroorsaak, deur ‘n groot aantal van hierdie uitvloei pompe in die selwand, is nie duidelik nie en moet verder ondersoek word. Die kruis-weerstandigheid tussen CFZ en BDQ, wat veroorsaak word deur mutasies in Rv0678, is van belang en kan die beplanning van anti-TB middel behandeling vir die toekoms beïnvloed. Die rolle van die ander twee mutasies, wat in hierdie studie geïdentifiseer is, in CFZ weerstandigheid is ook nie duidelik nie en vereis verdere ondersoek. Ten slotte, die bevindinge van hierdie studie steun die rol van Rv0678 in CFZ weerstandigheid en dit dui daarop dat hierdie geen gebruik kan word as 'n diagnostiese merker om vir CFZ weerstandigheid te toets in kliniese isolate.

Thesis (PhD)--Stellenbosch University, 2000. / ENGLISH ABSTRACT: Drug-resistant tuberculosis poses a threat to global tuberculosis control by the WHO DOTS strategy. Studies in the United States and Europe have shown (i) that drug-resistant tuberculosis is present in every country; (ii) that, by contrast to previous dogma, drug-resistant bacilli are virulent and can be transmitted, especially in institutional settings and to immunocompromised patients; and (iii) that the majority of cases arise by acquisition of drug resistance due to errors in the management of TB cases. (iv) Furthermore, it has been shown that the extremely high case fatality rates of the 1980s and early 1990s can be reduced by individualized, but costly treatment. However, the majority of drug-resistant TB cases reside in the developing world. Data on disease epidemics in less developed parts of the world are scarce. The aim of this thesis was to study the disease dynamics of drug-resistant TB in a developing country where TB is endemic. All cases of drug-resistant TB during a 5-year period in two communities with poor socioeconomic living conditions were included for this observational study. Three different methods were used: restriction fragment length polymorphism (RFLP), mutation detection analysis by dot-blot hybridisation technique and a Geographic Information System. Results of RFLP analysis and mutation detection analysis showed that community outbreaks of drug-resistant Mycobacterium tuberculosis strains occur, even without the involvement of immunocomprimised patients. Infection with a drug-resistant strain occurred in new patients (primary drug resistance) as well as in patients treated before (exogenous reinfection). Exogenous reinfection was also shown to be an important mechanism of recurrence after previous cure for drug-sensitive TB. Transmission of drug-resistant strains occurred more frequent in areas with lower socioeconomic living conditions. The relative contribution of transmission differed substantially between the group of multi drugresistant (two thirds of cases) and single-drug-resistant (no cases) cases, which probably reflects the prolonged infectiousness of multi drug-resistant cases. To stop the growing epidemic of multi drug-resistant TB, prevention of acquisition as well as transmission of drug-resistant tuberculosis will be required. This will only be possible in areas where a DOTS strategy is well functioning and with a modification of central elements of the standard DOTS mechanism: a "DOTS-plus" strategy. Early and accurate diagnosis of drug resistance is essential for effective management. Diagnosis based on two direct smear tests might have to be replaced by routine drugsusceptibility tests at diagnosis. Because the routine performance of phenotypic drugsusceptibility tests was inferior to the performance of genotypic tests, the development of an affordable commercial kit testing a limited number of mutations conferring resistance could be of great value in the global fight against multidrugresistant TB. Because of the importance of early diagnosis, selective active contact tracing for multidrug-resistant cases, additional to the routine passive contact tracing, could prove to be cost-effective. Individualized treatment regimens are effective in reducing the failure rate, mortality and probably transmission of multidrug-resistant TB. Multidrug-resistant tuberculosis is a problem confronting the efforts for global tuberculosis control. Efficient strategies to turn the tide exist, but international political commitment and financial support will be essential. / AFRIKAANSE OPSOMMING: Middel weerstandige tuberkulose hou 'n bedreiging in vir globale tuberkulose kontrole deur die WGO DOTS strategie. Studies in die Verenigde State en Europa het getoon (i) dat middel weerstandige tuberkulose in alle lande voorkom; (ii) dat, in teenstelling met vorige dogma, middel weerstandige bakterieë virulent is en oorgedra kan word, veral in inrigtings en aan immuun-onderdrukte pasiënte; en (iii) dat die meeste gevalle ontstaan deur die verwerwing van middel weerstandigheid a.g.v. die foutiewe hantering van tuberkulose gevalle. (iv) Bykomend is getoon dat die ontsettende hoë mortaliteit syfers van die 1980s verlaag kan word deur geindividualiseerde, maar duur behandeling. Die meeste middel weerstandige tuberkulose gevalle woon egter in die ontwikkelende wêreld. Data oor siekte epidemies in minder ontwikkelde dele van die wêreld is skaars. Die doel van hierdie tesis was om die siekte dinamiek van middel weerstandige tuberkulose te bestudeer in 'n ontwikkelende land waar tuberkulose endemies is. Alle gevalle van middel weerstandige tuberkulose gedurende 'n 5-jaar periode in twee lae sosio-ekonomiese gemeenskappe, is in hierdie studie ingesluit. Drie verskillende metodes is gebruik: restriksie fragment lengte polimorfisme (RFLP), mutasie analise deur dot-blot hibridisasie en 'n Geografiese Inligting Stelsel. Resultate van die RFLP analise het getoon dat uitbrake van middel weerstandige Mycobacterium tuberculosis stamme in die gemeenskap voorkom, selfs sonder die aantasting van immuun-onderdrukte pasiënte. Infeksie met middel weerstandige stamme het voorgekom in nuwe pasiënte (primêre middel weerstandigheid) en ook in pasiënte wat reeds voorheen behandel is (eksogene herinfeksie ). Daar is ook gevind dat eksogene herinfeksie 'n belangrike meganisme was van herhaalde tuberkulose na vorige genesing van middel sensitiewe tuberkulose. Die oordrag van middel weerstandige stamme het meer dikwels voorgekom in areas met laer sosioekonomiese omstandighede. Die relatiewe bydrae van oordrag het merkwaardig verskil tussen multi-middel weerstandigheid (twee derdes van gevalle) en enkelmiddel weerstandigheid (geen gevalle). Dit weerspieël waarskynlik die verlengde periode van infektiwiteit van die multi-middel weerstandige gevalle. Die bekamping van die groeiende epidemie van multi-middel weerstandige tuberkulose, vereis die voorkoming van verworwe sowel as oorgedraagde middel weerstandige tuberkulose. Dit sal slegs moontlik wees in areas waar 'n DOTS strategie reeds goed funksioneer en met 'n aanpassing van die sentrale elemente van die roetine DOTS meganisme: 'n "DOTS-plus" strategie. Vroeë en akkurate diagnose van middel weerstandigheid is essensieël vir effektiewe hantering. Diagnose gebaseer op twee direkte sputum smeer toetse mag moontlik vervang moet word deur roetine middel sensitiwiteit bepalings by diagnose. Die roetine fenotipiese middel sensitiwiteit bepaling is gevind om minderwaardig te wees in vergelyking met die genotipiese toetse. Die ontwikkeling van 'n bekostigbare toetsstelsel wat die mees algemene mutasies vir middel weerstandigheid sal opspoor, kan van groot waarde wees in die stryd teen mutimiddel weerstandige tuberkulose. Aangesien vroeë diagnose so belangrik is, kan aktiewe kontak opsporing koste-effektief wees. Ge-individualiseerde behandelingskedules is effektief om die sukses van behandeling en oorlewing te verbeter, en moontlik ook om die oordrag van multi-middel weerstandige tuberkulose te verminder. Multi-middel weerstandige tuberkulose is 'n probleem vir die globale kontrole van tuberkulose. Effektiewe strategieë om die vloed te stuit, bestaan, maar politieke verbintenis en geldelike ondersteuning sal essensieël wees.

Se evaluó aplicando el Test de Hamilton a 90 pacientes, portadores de tuberculosis, en sus diferentes tipos, y que vienen recibiendo el tratamiento correspondiente a través del Programa Nacional de Control de la Tuberculosis en los establecimientos de Salud que conforman la Red Nº 05 Arequipa Sur, durante el año 2002, con el objetivo de determinar la relación existente entre TUBERCULOSIS y DEPRESIÓN ; para lo cual se utilizo el Inventario para la Depresión de Hamilton, y los datos obtenidos a través de una ficha en la cual figuran los datos clínicos y sociodemográficos más importantes de cada paciente. Se encontró que la muestra en estudio está conformada en su mayoría (83.34%) por pacientes cuyas edades fluctúan entre los 15 y 44 años, que son afectados mayormente los varones (62.22%), más del 50% tienen estudios secundarios, son solteros, tienen un trabajo independiente y cuentan con los servicios básicos de salubridad. El 77% de los enfermos es portador de la enfermedad a nivel pulmonar y reciben por tratamiento el esquema I, el 6% de los casos es portador de tuberculosis multidrogorresistente y por lo tanto reciben el esquema estandarizado de retratamiento. Presentan niveles más intensos de depresión (36.745%) aquellos pacientes que cuentan con estudios primarios (73.33%), también se encuentran más deprimidos los enfermos que tienen un trabajo en su domicilio (71,43%), asimismo se deprimen más los pacientes que provienen de la región Arequipa (62.22%) seguido de los provenientes de Puno (53.85%). La tuberculosis afecta en orden de frecuencia a: los pulmones, la pleura, los riñones, y el sistema linfático, de ellos se deprime con niveles más intensos los portadores de la enfermedad a nivel renal en un 100%, seguido de los enfermos pulmonares en un 38.23%. Los pacientes que reciben el esquema de tratamiento para tuberculosis multidrogorresistente alcanzan cuadros depresivos más severos (60%), seguido del grupo de pacientes que reciben el esquema I (39.14%).A mayor tiempo de duración del tratamiento los niveles de depresión tienen mayor intensidad, es así que los que reciben la terapia por más de seis meses lo están en el 60%. De esta manera se comprueba lo planteado en la hipótesis, es decir que existe relación directa entre la Tuberculosis y la depresión en la población estudiada.

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O Mycobacterium tuberculosis é o agente causador da tuberculose, doença responsável por 26% das mortes passíveis de prevenção no mundo todo. No Brasil, são notificados anualmente 85 mil casos novos, e estima-se que 50 milhões de pessoas estejam infectadas pelo M. tuberculosis. A tuberculose é considerada prioritária para o controle de doenças e agravos pelo Ministério da Saúde. Para esse controle, é fundamental disponibilizar métodos e recursos para o pronto diagnóstico laboratorial. Os métodos utilizados para o diagnóstico da doença são bacterioscopia, análise histológica ou cultivo do micro-oganismo a partir de amostras de escarro. A bacterioscopia apresenta baixa sensibilidade, e o resultado da cultura demanda um período de tempo de até oito semanas. A PCR é uma técnica de amplificação de ácidos nucléicos que tem se mostrado promissor instrumento para o diagnóstico da tuberculose. O M. tuberculosis é um micro-organismo que tem tropismo pelas células (micro-organismo intracelular), e pode estar presente nas células do trato respiratório e da mucosa bucal. O esfregaço bucal, ao contrário do escarro, é obtido de forma fácil, sem constrangimentos, por procedimento não invasivo, e oferece menores riscos de contaminação por outros micro-organismos. Foram analisadas 80 amostras de esfregaço bucal de pacientes com diagnóstico confirmado de tuberculose, das quais 78 (97,4%) tiveram resultado positivo na PCR. Esse resultado permite concluir que a aplicação da PCR em amostras de esfregaço bucal é um método efetivo e confiável para detecção do M. tuberculosis. / Mycobacterium tuberculosis is the causing agent of the tuberculosis, responsible illness for 26% of the prevention deaths in the entire world. In Brazil 85000new cases are notified annually, being esteem 50 million people contaminated by the M. tuberculosis. It is considered priority disease for the control of illnesses for the Health department. For this control, it has to be reliable methods and resources for the ready laboratorial diagnosis. Bacterioscopiv, histological analysis or culture of the microrganism from samples of sputum are techniques normally used. The limitation of these methods is low sensitivity and long-winded 8 weeks. The PCR is one technique of amplification of DNA, that if has shown promising instrument for the diagnosis of the tuberculosis. The M. tuberculosis is a microorganism that has affinity for the cells (intracellular microorganism) and can be present in the cells of the respiratory treat and the oral mucosa. Oral swab, in contrast of sputum, is easily taken, not invasive and offering lesser risks of contamination for other microorganisms. We analyze 80 samples of oral swab of patients with confirmed diagnosis of tuberculosis, of these, 78 (97,4%) had resulted positive in the PCR. We conclude that the oral swab use and the application of the PCR are an effective and trustworthy method for tuberculosis detention of the M. tuberculosis.