[show abstract][hide abstract] ABSTRACT: Using a novel multilocus DNA marker analysis method, we studied the population genetic structure of Trypansoma brucei stocks and derived clones isolated from animal and rhodesiense sleeping sickness patients during a national sleeping sickness control program in Mukono district, Uganda. We then performed a cladistic analysis to trace relationships and evolution, using stocks and clones recovered from geographically and temporally matched hosts, including inter-strain comparisons with T. b. gambiense stocks and clones. Our results show that while there was close genetic relatedness among parasite populations from the same geographical region, micro-heterogeneities exist between different stocks. Data are presented that indicate that not every human sleeping sickness focus may be associated with a particular human-infective trypanosome strain responsible for long-term stability of the reference focus. We provide evidence of genetic sub-structuring among type 1 T. b. gambiense stocks, which has potentially important implications for molecular epidemiology of T. brucei.
Infection Genetics and Evolution 10/2003; 3(3):165-74. · 2.77 Impact Factor
[show abstract][hide abstract] ABSTRACT: There have been tremendous advances in our knowledge of trypanosome biology, yet many aspects remain unclear. Currently, the genome of Trypanosome brucei is being sequenced and this, with other genome-wide analysis methods, could provide novel insights into the parasite and facilitate the development of effective controls. An important new challenge investigators face is how to exploit the information in studying a parasite with so many genetic peculiarities. Here, we summarize our current understanding of molecular genetics of T. brucei and attempt to link genome analysis to the prospects for identifying possible targets for vaccines, novel drugs and specific diagnostics. The value of newly developed genotyping approaches in accelerating these processes is discussed.
Trends in Microbiology 08/2003; 11(7):322-9. · 8.43 Impact Factor
[show abstract][hide abstract] ABSTRACT: The pathogenic trypanosomes Trypanosoma equiperdum, T. evansi as well as T. brucei are morphologically identical. In horses, these parasites are considered to cause respectively dourine, surra and nagana. Previous molecular attempts to differentiate these species were not successful for T. evansi and T. equiperdum; only T. b. brucei could be differentiated to a certain extent. In this study we analysed 10 T. equiperdum, 8 T. evansi and 4 T. b. brucei using Random Amplified Polymorphic DNA (RAPD) and multiplex-endonuclease fingerprinting, a modified AFLP technique. The results obtained confirm the homogeneity of the T. evansi group tested. The T. b. brucei clustered out in a heterogenous group. For T. equiperdum the situation is more complex: 8 out of 10 T. equiperdum clustered together with the T. evansi group, while 2 T. equiperdum strains were more related to T. b. brucei. Hence, 2 hypotheses can be formulated: (1) only 2 T. equiperdum strains are genuine T. equiperdum causing dourine; all other T. equiperdum strains actually are T. evansi causing surra or (2) T. equiperdum does not exist at all. In that case, the different clinical outcome of horse infections with T. evansi or T. b. brucei is primarily related to the host immune response.
[show abstract][hide abstract] ABSTRACT: Restriction enzyme-detectable polymorphisms have been used for assessing genetic differences and generating informative genetic markers. The most detailed fingerprinting analyses have been obtained using the AFLP (amplified fragment length polymorphism) technique, which accesses subsets of polymorphisms at one or two restriction sites. To combine increased discriminatory power with the stringency of polymerase chain reaction amplification, it would be beneficial to access additional independent restriction sites per analysis, and to amplify subsets of DNA restriction fragments with only one pair of oligonucleotide primers. We have now developed a unique approach that permits the simultaneous use of four or more endonucleases in combination with one pair of adapters/primers, and applied it to genotype 21 trypanosome populations to subspecific level. The approach takes advantage of the fact that some endonucleases create cohesive ends that are compatible with the overhang sites created by other endonucleases. We demonstrate the greater resolution of identifiable polymorphic fragments over the conventional ligation-mediated restriction analysis method, and discuss the value of the approach as a tool for fine genetic mapping of Trypanosoma brucei. Finally, we propose use of the method for fine characterisation and for identifying co-dominant genetic markers in a variety of other taxa.