Acta Tropica 75 (2000) 315–321 Comparative sensitivity of dot-ELISA, PCR and dissection method for the detection of trypanosome infections in tsetse flies (Diptera : Glossinidae ) Johnson O. Ouma a, *, Rachael A. Masake b , Daniel K. Masiga a , Shamshudeen K. Moloo b , James T. Njuguna b , Joseph M. Ndung’u a a Kenya Trypanosomiasis Research Institute (KETRI), P.O. Box 362, Kikuyu, Kenya b International Liestock Research Institute (ILRI), P.O. Box 30709, Nairobi, Kenya Received 25 October 1999; received in revised form 15 December 1999; accepted 7 January 2000 Abstract A visually read dot-enzyme linked immunosorbent assay (dot-ELISA) developed for the detection of trypanosomes in tsetse flies (Glossina spp.) was evaluated in the laboratory and under field conditions. In the evaluation, the fly dissection method was used as a standard technique and compared to the polymerase chain reaction (PCR). In laboratory studies, 133 and 126 tsetse flies were experimentally infected with different stocks of Trypanosoma brucei and T. congolense, respectively. Twenty-five days after infection, the flies were dissected and tested for the presence of trypanosomes using dot-ELISA and PCR. Dot-ELISA detected 98.4% of T. brucei and 94% of T. congolense infections in tsetse midguts, while PCR detected 97.6% of T. brucei and 96% of T. congolense tsetse midgut samples. For field evaluation of dot-ELISA, 700 tsetse flies were caught and screened for trypanosome infections by dissection. Seven of these (1%) had trypomastigotes in the midgut, 23 (3.3%) in the proboscis and none had trypanosomes in the salivary glands. All the flies with midgut infections also had trypanosomes in their proboscides. Five of the seven flies (71.4%) with midgut infections revealed by dissection, were also positive for T. congolense by the dot-ELISA and PCR techniques. Dot-ELISA detected T. congolense infections in an additional 86 (12.4%) of the 700 flies dissected. Of the 23 infections in the proboscis, 16 were T. iax. Dot-ELISA detected 13 of the 16 (81%) while PCR detected 15 of 16 (94%) T. iax infections. No T. brucei infection was detected by any of the methods in all the 700 tsetse flies examined. The results obtained from both the laboratory and field studies indicate that the dot-ELISA and PCR techniques are sensitive and species-specific in revealing trypanosome infections in tsetse flies. While dot-ELISA required a single test to detect T. congolense, several primer pairs were needed for PCR. The potential use of dot-ELISA as a tool for studying the epidemiology of trypanosomosis, while considering its field applicability and relatively lower cost is discussed. © 2000 Elsevier Science B.V. All rights reserved. Keywords: Dot-ELISA; Polymerase chain reaction (PCR); Trypanosoma brucei ; Trypanosoma congolense ; Trypanosoma iax ; Glossina pallidipes ; Glossina longipennis www.elsevier.com/locate/actatropica * Corresponding author. Tel.: +254-154-329604; fax: +254-154-32397. E-mail address: ketri@net2000ke.com (J.O. Ouma) 0001-706X/00/$ - see front matter © 2000 Elsevier Science B.V. All rights reserved. PII:S0001-706X(00)00065-6