Molecular Identification of Trypanosoma theileri and Biology of Trypanosomes

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 plotted 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.

Isolation of a Trypanosome Related to Trypanosoma theileri (Kinetoplastea: Trypanosomatidae) from Phlebotomus perfiliewi (Diptera: Psychodidae)

The Trypanosoma theileri group includes several trypanosome species hardly distinguishable due to the lack of discriminating morphological characters. Trypanosomes belonging to this group have been isolated from different bovine, ovine, and cervids in Europe, Africa, Asia, and Americas. The principal vectors of the T. theileri group are considered tabanid flies; however, T. melophagium is transmitted exclusively by sheep keds. In 2016, 128 sand flies out of 2,728 trapped in Valsamoggia municipality, Italy, were individually dissected and an unknown trypanosome strain, named TrPhp1, was isolated from a female of the sand fly Phlebotomus perfiliewi. Sequence analysis placed this trypanosome in the T. theileri group with very high homology to other trypanosomes detected in European cervids. This is the first report of the T. theileri group isolation from a sand fly, and the possible role of this insect group in the trypanosome transmission cycle is discussed. Within the T. theileri group, the phylogenetic analysis distinguished several lineages, which, unfortunately, do not correspond with their host specificity and their taxonomic status remains ambiguous.

Characterization of tabanid flies (Diptera: Tabanidae) in South Africa and Zambia and detection of protozoan parasites they are harbouring

SUMMARYTabanids are haematophagous flies feeding on livestock and wildlife. In the absence of information on the relationship of tabanid flies and protozoan parasites in South Africa and Zambia, the current study was aimed at characterizing tabanid flies collected in these two countries as well as detecting protozoan parasites they are harbouring. A total of 527 tabanid flies were collected whereby 70·2% were from South Africa and 29·8% were from Zambia. Morphological analysis revealed a total of five different genera collected from the sampled areas namely:Ancala, Atylotus, Haematopota, PhilolicheandTabanus. DNA extracted from South AfricanTabanus parandTabanus taeniolatested positive for the presence ofTrypanosoma congolense(Savannah) andTrypanosoma theileriwhilst one member fromT. parwas positive forTrypanosoma bruceispecies. DNA extracted from Zambian tabanid flies tested positive for the presence ofBesnoitiaspecies at 1·27% (2/157),Babesia bigemina5·73% (9/157),Theileria parva30·11% (30/157) and 9·82% (14/157) forTrypanosoma evansi. This study is the first to report on relationship ofBabesiaandTheileriaparasites with tabanid flies. Further investigations are required to determine the role of tabanids in transmission of the detected protozoan parasites in livestock and wildlife in South Africa and Zambia.