Molecular Identification of Trypanosoma theileri and Biology of Trypanosomes
Tewodros Fentahun, Jan Paeshuyse
World's Veterinary Journal
Trypanosoma theileri (T. theileri ) is a non-pathogenic, cosmopolitan, and commensal protozoa of cattle. The main objective of the current study was to investigate the biology and feasibility of T. theileri as a model candidate for the discovery of a novel drug. In the present study, the isolates of T. theileri obtained from the Institute of Tropical Medicine (ITM) in SDM 79 were cultivated at 26oC. Eight experiments with different inoculum and different times were grown. The growth curve was
... otted to check the growth trends. The doubling time in the logarithmic phase was determined to be 17.43 hours. In addition, an experimental infection was done on a 3-month-old Holstein Friesian calf to isolate the blood-streaming shape; however, it was not successful after the blood buffy coat smear and PBMC culture in RPMI 1640 and HMI 9. Furthermore, the viability was determined by quantitative colorimetric Resazurin assay in 96-well fluorescence Microplates containing 0.4 to 2.4 mM of Resazurin. On the other hand, the response to Pentamidine (1-100 ng/mL) showed a strong negative correlation between the fluorescence signal and the highest Pentamidine concentration. IC50 was 9.25 ng/mL. Genomic DNA was extracted using the phenol-chloroform method. The gradient PCR amplification using T. theileri specific PCR (Tth625-PCR) primers was detected at 465 base pair (bp). In addition, the full-length 18S rDNA sequence was detected at 730 bp. In the silico analysis using common anti-trypanosome drug targets, no significant similarity could be found on either the DNA or the protein level. Nevertheless, homologous sequences have been identified among the drug targets for Ornithine decarboxylase. Therefore, the analysis might show the possibility of using T. theileri as a model for the search of new drugs once they have entire genome sequences. Analysis of the whole genome and transcriptome indicated a phylogenetic relationship between T. theileri and other pathogenic trypanosomes which can be the basis for novel drug development.