Thesis (PhD)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: Tuberculosis (TB) has become a global health epidemic affecting millions of people worldwide
with a high incidence in third-world countries. The emergence of multi-drug and extremely-drug
resistant M. tuberculosis strains together with the HIV/AIDS pandemic warrants the need for
new drugs or new drug combinations.
The folic acid synthesis pathway is one of the key pathways that are essential for the survival of
bacteria in general. Sulfonamides are a group of compounds that target folic acid synthesis,
particularly dihydropteroate synthetase, the first enzyme in the folate pathway. Some of these
sulfonamides were used during the introduction of chemotherapy for the treatment of TB in the
1930s, but had toxic side effects. Newer derivatives became safer, but were not employed again
for TB treatment. In a recent case study it was reported that the combination of trimethoprim-sulfamethoxazole
(Bactrim), which is used to treat various bacterial infections, such as urinary tract infections, had
activity against M. tuberculosis. In light of this and the fact that trimethoprim-sulfamethoxazole
is well tolerated by humans, we have investigated their antimycobacterial activity with particular
interest in the combinational effect of sulfamethoxazole and trimethoprim with the first-line
anti-TB drugs, Isoniazid, Rifampicin and Ethambutol against M. tuberculosis. Since sulfonamides
are known to produce oxidative stress, we also investigated the contribution of this factor to the
efficacy of sulfamethoxazole using a mycothiol deficient strain of M. tuberculosis, ΔmshA.
Though trimethoprim-sulfamethoxazole targets the folic acid pathway, we also investigated the possibility that trimethoprim-sulfamethoxazole may have other cellular targets and applied
proteomic analysis.
We have found that Trimethoprim-Sulfamethoxazole has activity against M. tuberculosis and
that Sulfamethoxazole is the active compound. However, our observation was that not all
sulfonamides are active against M. tuberculosis. In addition we observed that sulfamethoxazole
enhances the activity of Rifampicin against M. tuberculosis in a synergistic way. We also
observed that a mycothiol deletion mutant was more susceptible to Sulfamethoxazole compared
to the wild type strain CDC 1551. Through global protein expression profiling (Proteomics) we
were also able to show that sulfamethoxazole could also kill M. tuberculosis by oxidative stress
production as we identified oxidative stress responsive proteins that were differentially
regulated upon exposure to sulfamethoxazole. As trimethoprim-sulfamethoxazole is a registered
drug combination, inexpensive and widely available, we propose that this regimen could be used
in our fight against M. tuberculosis infection. / AFRIKAANSE OPSOMMING: Tuberkulose (TB) is ‘n globale gesondheidsprobleem wat miljoene mense wêreldwyd affekteer
met ‘n besoderse hoë voorkoms in die derdewêreld lande. Die voorkoms van multi-middel
weerstandige en uitersweerstandige M. tuberculosis stamme, tesame met die HIV/VIGS
pandemie, steun die erns vir die ontwikkeling van nuwe middels teen M.tuberculosis .
Die foliensuur sintesepad is essensieël tot die oorlewing van bakterieë in die algemeen. Vir
daardie rede is daar vele middels ontwerp om hierdie metaboliese pad te teiken. Die
sulfonamiedes is ‘n groep antibiotika wat foliensuursintese, spesifiek dihidropteroaatsintese,
die eerste ensiem in die foliensuursintese pad, teiken. Van hierdie sulfonamiedes is voorheen
in die 1930’s gebruik vir die behandeling van tuberkulose, maar het toksiese newe-effekte
getoon. Nuwe, minder toksiese derivate, is later ontwikkel maar is nooit vir TB behandeling
weer aangewend nie. In ‘n onlangse gevallestudie is daar gerapporteer dat die kombinasie
trimethoprim-sulfamethoxazole (TMP/SMX. Handelsnaam: Bactrim), wat normaalweg
gebruik word vir die behandeling van algemene bakteriële infeksies soos blaasinfeksies,
aktiwiteit teen M. tuberculosis getoon het. Na aanleiding hiervan en dat Bactrim veilig in
mense gebruik kan word, het ons die aktiwiteit van Bactrim komponente teen M. tuberculosis
bepaal en in die besonder die aktiwiteite van SMX en TMP in kombinasie met die eerstelinie
anti-tuberkulose middels Isoniasied, Rifampisien en Ethambutol. Aangesien sulfonamiedes ook oksidatiewe stres intrasellulêr genereer, het ons ook die bydrae van hiervan tot die
doeltreffendheid van SMX bepaal deur gebruik te maak van ‘n mycothiol-gemuteerde M. tuberculosis stam ( mshA). Omdat TMP/SMX die foliensuur-pad hoofsaaklik teiken het ons
ook die moontlikheid ondersoek dat SMX ander sellulêre teikens het en het ons proteomiese
(Proteomics) tegnieke hiervoor aangewend. Ons het gevind dat TMP/SMX aktiwiteit teen M.
tuberculosis toon en dat SMX die aktiewe komponent van Bactrim is teen M. tuberculosis .
Ons wys ook dat sulfonamiedes in die algemeen nie noodwendig ook aktiwiteit teen M.
tuberculosis toon nie. Ons het ook waargeneem dat SMX die aktiwiteit van rifampisien
bevorder en dat die twee middels saamwerk op ‘n sinergistiese wyse. Ons het ook getoon dat
oksidatiewe stres ‘n rol speel deurdat‘n mycothiol delesie-mutant meer vatbaar was vir SMX
in vergelyking met die wilde-tipe stam van M. tuberculosis (CDC1551). Met globale proteïenkartering
(Proteomics) het ons ook getoon dat SMX M. tuberculosis kan doodmaak deur
oksidatiewe stres te genereer omdat ons oksidatiewe stres reaktiewe proteïne geïdentifiseer
het wat differensieël gereguleer is gedurende blootstelling aan SMX. Aangesien Bactrim ‘n
reeds geregistreerde middel is, goedkoop is en geredelik beskikbaar is, stel ons voor dat
Bactrim moontlik geïnkorporeer kan word in die huidige behandeling van Tuberkulose.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/86743 |
| Date | 04 1900 |
| Creators | Macingwana, Lubabalo |
| Contributors | Wiid, Ian J. F., Baker, B., Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences, Molecular Biology and Human Genetics. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | English |
| Type | Thesis |
| Format | xii, 129, [4] pages : illustrations (some colour), colour maps |
| Rights | Stellenbosch University |
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