Authors

SS Abouakkada²; A Deweir³; HP Price¹; N Labn²; SA henidy²;  ¹ School of Life Sciences, Keele University, UK;  ² Faculty of Veterinary Medicine, Alexandria University, Egypt;  ³ Faculty of Veterinary Medicine, Alexandria University, Egypt, Egypt

Discussion

Trypanosoma evansi is the causative agent of Surra, one of the most economically important veterinary diseases in the world. Emergence of drug resistance represents a great barrier to chemotherapeutic control against trypanosomosis. Drug resistance is usually caused by changes to the drug transporters of the parasites. In Egypt, the prevalence of drug-resistant trypanosomes in endemic regions remains poorly understood. In the present study, we aim to investigate molecular markers of drug resistance in the form of mutations in adenosine transporter P2 (TbAT1) and Aquaglyceroporin (AQP2) genes encoding drug transporters in different strains of T.evansi collected in Egypt. We have sequenced the TbAT1 and the AQP2 genes from six T.evansi field strains collected from camels from different localities in Egypt and one laboratory UK strain. Blood samples were collected from the animals for extraction of total genomic DNA was performed for amplification of 164 bp TBR1/2 primers. The seven TBR1/2 positive DNA samples were used for amplification of 1600 and 1416 bp encoding for adenosine transporter P2 gene and Aquaglyceroporin transporter gene, respectively. Purified PCR products were sequenced and BLAST analysis was performed to establish sequence identity to sequences accessed from GenBank.Results revealed that the seven tested DNA sampleswere positive to TBR1/2 primers. In addition, three strains representing South Sinai, Matrouh and Halaib had lost both AQP2 and TbAT1 genes. The other four strains were positive for AQP2 and TbAT1 genes and gave specific bands at 1416 and 1600 bp, respectively. Alignment results of AQP2 gene revealed no polymorphism at the nucleotide or amino acid levels at any of the tested strains. Alignment of AT1 gene sequences revealed substitution of TG nucleotide by CT at the position of 972 and 973 laboratory and Cairo1 strains, with insertion of two nucleotides, GC at the position of 974 and 975 that resulted in insertion of a new amino acid, Cysteine. Both strains carried the same point mutations. Cairo2 strain showed a substitution of one nucleotide T by C at the position of 972 and insertion of the two nucleotides GC at the same positions of laboratory and Cairo1 strains. This resulted in insertion of the amino acid, Glycine at this site. Moreover, substitution of nucleotide G by T at the position of 710 resulted in substitution of glycine amino acid at the position 237 with Valine. A Kom-hamada strain displayed no polymorphism neither at nucleotide nor amino acid levels. In conclusion, the data presented involved the detection of TbAT1 gene in 4 out of 7 strains of