Molecular identification and drug sensitivity of African trypanosome stabilates from livestock in Lamu County, Kenya

Abstract

Animal African Trypanosomiasis (AAT) causes economic losses estimated at US $5 billion per annum. Records from the veterinary department indicate high levels of trypanocidal drugs use in Lamu County. The objective of this study was to identify the trypanosome species causing infections in domestic animals in Lamu using parasitological and molecular techniques and to determine their drug sensitivity in white Swiss mice. Fifteen trypanosome stabilates and 92 whole blood samples collected from parasitologically negative animals were randomly retrieved from cryobank freezer at Biotechnology Research Institute (KALRO-BioRI), Kenya, and characterized. Human serum resistance associated (SRA) gene present in T. b. rhodesiense was used to differentiate T. brucei positive stabilates. Four stabilates identified as T. b. brucei were used for drug sensitivity study. Groups of six white Swiss mice were infected with these stabilates and treated with single doses of Homidium Bromide (1mg/kg), Isometamidium Chloride (1mg/kg) and Diminazene Aceturate (20mg/kg). Pathogenicity and virulence determination for one drug sensitive and two drug resistant stabilates was also carried out. Changes in packed cell volume (PCV), parasitaemia and body weights of mice were monitored. Results showed that 10/15 (67%) trypanosome stabilates and 13/92 (14%) whole blood samples from cattle, donkeys and goats were positive by PCR. Positive T. congolense 5/23 (22%) gave a product size of 700bp using ITS1 primers. Brucei group 7/23 (30%) and T. vivax 11/23 (48%) amplicons were 480bp and 250bp, respectively. Identified trypanosome stabilates were T. b. brucei (7), T. vivax (2) and T. congolense Savannah (1). Whole blood PCR profiles revealed T. vivax (9) and T. congolense Savannah (4). Trypanosoma b. brucei, T. vivax and T. congolense Savannah were the etiological agents for AAT in donkeys. In contrast, T. vivax and T. congolense Savannah caused the disease in cattle, and T.b.brucei in goats within Lamu County. KETRI 4028 stabilate was sensitive to Isometamidium and Diminazene. KETRI 4032, KETRI 3985 and KETRI 3984 stabilates were resistant to the three drugs used in this study. Importance of including molecular and parasitological methods when carrying out epidemiological disease surveys was highlighted. The results also indicate presence of more T. b. brucei sub populations circulating in Lamu that exhibit multiple resistance to Homidium, Isometamidium and Diminazene and presence of about 25% of sub population of T. b. brucei in Lamu that may be sensitive to Isometamidium and Diminazene and recommends that use of Homidium should be discouraged while use of Diminazene and Isometamidium should be used only in cases that proper diagnosis of the disease has been done.