Abstract:

Trypanosoma brucei is a tsetse fly-transmitted protozaon pathogen of the genus Trypanozoon
that causes sleeping sickness or Human African Trypanosomiasis (HAT) in man and nagana
in animals. HAT is invariably fatal unless treated and causes huge economic losses to the
affected communities. Therefore HAT management is of great importance in improving the
livelihoods of such communities. HAT management involves accurate diagnosis and
chemotherapy. However, HAT is a neglected disease with respect to treatment as well as
diagnostics. In this thesis I set out to address some of the problems related to HAT diagnostics
and chemotherapy. This work comprises five (5) chapters. Chapter 1.0 which is the
Introduction: gives a general overview on Trypanosoma brucei and HAT. Chapters 2.0, 3.0
and 4.0, comprise the results. Chapter 2.0 is an aspect of HAT diagnostics while chapters 3.0
Serodiagnostic evaluation of Tandem Repeat (TR) Proteins in Trypanosoma brucei. In the
absence of laboratory equipement, serology is the main stay for diagnosis of HAT under field
conditions. Trypanosoma brucei TR proteins were retrieved and identified in silico and genes
of selected proteins with expression profiles in the bloodstream form of the parasite were
amplified and expressed in E. coli. The recombinant proteins were purified and evaluated
using ELISA. Preliminary results obtained show that the selected proteins cannot be used as
antigens for HAT diagnosis since they do not distinguish between sera from HAT patients
and that from healthy individuals. Chapter 3.0: Mechanisms of drug resistance in
Trypanosoma brucei is divided into three parts i) Combined contribution of TbATl and
aminopurine transporter
responsible for uptake of trypanocides like melarnophenyl arsenicals, pentamidine,
efflux transporter of the ABC type which contributes to melanophenyl arsenical resistance
TbMRPA to drug resistance in Trypanosoma brucei. TbATl is an
diminazene aceturate, isometamidium, cordycepin and tubercidin, while TbMRPA is an
and 4.0 are aspects of chemotherapy. Chapter 2.0 is Comparative genomics and
2 Abstract
when overexpressed. The aim of this work was to investigate what happens when both loss of
TbATl and gain in TbMRPA, coincide in the same cell. Results obtained show that the two
transporters independently contribute to arsenical resistance in Trypanosoma brucei (Chapter
3.0a). Part ii (Chapter 3.0b) and iii (Chapter 3.0c), investigate the phenomenon of crossresistance
between melanophenyl arsenicals and diamidines in Trypanosoma brucei. Drug
resistance to melarsoprol and pentamidine was independently induced in the laboratory strains
and the mechanisms involved were investigated. In chapter 3.0b the mechanism for crossresistance
is not known since only the melarsoprol-resistant isolate had lost the TbATl. In
chapter 3.0c however, cross-resistance is associated with the loss of the High Affinity
Pentamidine Transporter (HAPT1) activity. In Chapter 4.0: Comparative genomics of
metabolic networks from parasites and host, the aim was to determine how networks shrink
and to identify potential drug targets. In this work pathways constituting core metabolism of
different organisms were broken down into individual reactions and analysed in the
perspective of entire networks. Different properties of the networks were compared between
parasites and non-parasites. Two enzymes specific to Tiypanosoma brucei', methionine
synthase (EC2.1.1.14) and homocysteine methyltransferase (EC2.1.1.10) were knocked down
and knocked out, respectively and the generated clones were grown in methionine-free or
methionine-supplemented medium. Results obtained show that network integrity rather than
scale-freeness has acted as a selective principle for evolution of parasite metabolism.
Experiments on methionine synthase and homocysteine methyltransferase show that
Trypanosoma brucei is auxotrophic for methionine and that homocysteine methyltransferase
and methionine synthase are potential drug targets. Chapter 5.0 is the overall discussion and
outlook of all the work done and written in this thesis.