The use of DNA hybridization and numerical taxonomy in determining relationships between Trypanosoma brucei stocks and subspecies

Abstract

The nuclear DNAs of 71 trypanosome stocks from different African countries, representative of the three Trypanosoma brucei subspecies, and one T. evansi stock, have been analysed by the combined use of restriction endonuclease digestion, gel electrophoresis and molecular hybridization with both trypanosome surface-antigen-specific and undefined genomic DNA probes. In contrast with T. brucei brucei and T. brucei rhodesiense stocks, all the T. b. gambiense stocks are characterized by a conserved, specific DNA band pattern, regardless of the probe. This allows T. b. gambiense to be non-ambiguously identified. On the contrary, T.b. brucei and T. b. rhodesiense, which could not be discriminated by the same criteria, both yield highly variable DNA band patterns. Our data confirm that domestic animals like pig, dog and sheep constitute a potential reservoir for T.b. gambiense. Using a numerical analysis of the DNA hybridization patterns we have measured the degree of similarity between the 72 trypanosome stocks. This investigation shows that all T.b. gambiense stocks are included in the same homogeneous population, while the stocks from the two other subspecies seem to be distributed in several heterogeneous groups, some of these showing correlation with the geographical origin of the trypanosomes. It is concluded that (i) T.b. gambiense stands out as a real subspecies that has undergone a distinct evolution relative to the 'non-gambiense' group, (ii) the alleged T.b. rhodesiense subspecies does not fit with any of the groups evidenced by our cladistic analysis and hence does not appear as a distinct subspecies and (iii) 'non-gambiense' trypanosomes are probably evolving much more rapidly than T.b. gambiense. Different aspects of trypanosome relationships and evolution are discussed.

Novel primer sequences for polymerase chain reaction-based detection of Trypanosoma brucei gambiense.

“…These strains were previously identified as being distinct from the conventional T. b. gambiense group 1, sharing molecular characteristics with Nigerian strains of T. b. brucei. 4,35,38,55 This initial characterization was based on cluster analysis of the RFLP pattern derived from the ribosomal non-transcribed spacer region. 35 Since the TgsGP-PCR targets a gene that is associated with NHS resistance, we subjected the Abba and Ligo strains to the SIIT.…”

“…[28][29][30][31][32][33][34][35][36][37][38][39][40][41][42] By running two subsequent TgsGP-PCRs on the same sample of spiked blood, a detection limit of 10 trypanosomes/ml of blood could be obtained. This is comparable with that reported with a two-run ESAG6/7 PCR or a repetitive nuclear DNA-based PCR used with blood samples from patients with sleeping sickness.…”

“…4 Within this context, various molecular techniques have been applied, such as isoenzyme analysis, restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA analysis (RAPD), karyotype analysis, polymorphism analysis within the sequences of the variant surface glycoprotein (VSG), mini-satellites and microsatellites, kinetoplast DNA, and an internal transcribed spacer 1 of rDNA. [28][29][30][31][32][33][34][35][36][37][38][39][40][41][42] These techniques often require prior expansion of trypanosome populations in laboratory animals or in culture medium and involve multiple and timeconsuming analytical steps. Taking into account the low isolation success rates observed for T. b. gambiense, and the fact that during primary isolation, and subsequent expansion, initially mixed-infection populations can be lost due to selection for the best growing one, the need for simplified techniques able to discriminate between the three T. brucei subspecies within the animal reservoir and the vector is obvious.…”

Expression of a Gene Encoding Trypanosoma congolense Putative Abc1 Family Protein is Developmentally Regulated

“…The diagnostic techniques have successively evolved through molecular biological techniques, such as DNA hybridization, PCR and loop-mediated isothermal amplification of DNA [11,16,19]. The diagnosis using PCR aims to identify the parasites at the species level using various target genes.…”

Les lents progrès du contrôle de la maladie du sommeil

“…Noireau et al (1989) ont identifié T. b. gambiense chez plusieurs espèces animales en République Populaire du Congo en utilisant l'hybridation de l'ADN des para sites associée à la taxonomie numérique (Paindavoine et al , 1986). Au Zaïre, Kageruka (1989) a isolé deux souches assi milables à T. b. gambiense chez la chèvre et le porc, parmi de beaucoup plus nombreux autres isolats de même mor phologie identifiés comme étant T. b. brucei.…”