Trypanosoma evansi: A comparison of PCR and parasitological diagnostic tests in experimentally infected mice D. Fernández a , B. González-Baradat a , M. Eleizalde a , E. González-Marcano b , T. Perrone b , M. Mendoza a, * a Universidad Nacional Experimental Simón Rodríguez, Instituto de Estudios Científicos y Tecnológicos (IDECYT), Centro de Estudios Biomédicos y Veterinarios, Apartado Postal 47925, Caracas 1041A, Venezuela b Laboratorio de Fisiología de Parásitos, Centro de Biofísica y Bioquímica, Instituto Venezolano de Investigaciones Científicas (IVIC), Apartado Postal 21827, Caracas 1020A, Venezuela article info Article history: Received 26 March 2008 Received in revised form 16 September 2008 Accepted 18 September 2008 Available online 26 September 2008 Keywords: Molecular and parasitological techniques Diagnosis Trypanosoma evansi Venezuela abstract Trypanosoma evansi is the causative agent of equine trypanosomosis, disease that affects horse’s produc- tivity and health. Parasitological and molecular methods are mostly used to detect the infection. The aim of this work was evaluate PCR sensitivity to detect T. evansi using the primers 21/22-mer, ITS1, ESAG 6/7 and TBR 1/2 designed from repetitive (multicopies) genomic sequences. The results were compare with two parasitological tests in mice, micro-haematocrite centrifugation technique and direct microscopic examination. The results shows (a) that the minimum amount of DNA from blood of highly parasitaemic mice that was detectable by PCR was 0.001 ng, using the ESAG6/7 and TBR1/2 primer, (b) using TBR1/2 primer for parasites purified could detect 0.000001 ng and (c) in the prepatent period PCR detect the presence of parasites earlier than parasitological techniques. Nevertheless, the percentage of detection for PCR varies depending on primer employed with 60% and 66% for ITS1 and 21/22-mer, and 80% for ESAG6/7 and TBR1/2. Consequently, TBR1/2 and ESAG6/7 were the best primers to monitor T. evansi infections in mice. For epidemiological application, such comparative evaluation should be made for detection of T. evansi in livestock such as horses. Ó 2008 Elsevier Inc. All rights reserved. 1. Introduction Trypanosomosis is a disease caused by a protozoan of the Try- panosomatidae family, mainly distributed in tropical areas around the world, which can affect either humans or animals. Animal Try- panosoma are of great concern for veterinary laboratory diagnosis especially in inter-tropical areas of Asia, Africa and Latin America, where most of the pathogenic Trypanosoma species are present depending on the geographical distribution of the parasites, and the various species potentially present in a host, or a vector (Des- quesnes and Dávila, 2002). In Latin America (from Venezuela to Argentina), Trypanosoma evansi causes equine trypanosomosis, dis- ease known as derrengadera or surra (Desquesnes, 2004). This par- asitic disease produces negative effects on health and even can cause the death of the animal, produce a decrease in the productiv- ity of horses used in livestock, which are indispensable for tradi- tional extensive cattle ranching. This illness generates great economical losses in affected areas, but very few data is available. In Argentina, for example, antibody detection has shown 20% of po- sitive animals in the Formosa region where approximately 57,000 horses are exposed (Monzón et al., 1995). In the Pantanal region of Brazil, about 13% of the horses are lost annually and no appropriate control is used (Seidl et al., 2001). In the Venezuelan savannah, it has been reported an 80% and 50% seroprevalence for T. evansi in horses (Reyna-Bello et al., 1998) and capybaras (Arias et al., 1997), respectively. In Latin America, animal trypanosomosis is mechanically trans- mitted by haematophagous insects. The period between inocula- tion of the parasites in healthy animals, and their detection in blood or tissue fluids, by direct observation, is called prepatency. Following the prepatent period, the disease will continue in two phases, an acute phase characterized by high levels of parasitaemia and noticeable clinical symptoms, and a chronic phase character- ized by low parasitaemia, which can lead to emaciation or become unapparent symptoms. In the last phase, the host may become a parasite reservoir. To prevent decreasing physical conditions lead- ing to death of the animals, it would be necessary a good knowl- edge of epidemiologic data, which includes availability of sensitive and specific diagnostic techniques capable of detecting the infectious agent particularly in the prepatency and/or the chronic phase of the disease. Animal trypanosomosis is a serious problem in most countries of Africa, Asia and Latin America (Duvallet et al., 1999; Dávila et al., 2003). Hence, the development and improvement of meth- ods to control this disease is of global importance. There are several diagnostic tests for trypanosome, which have both advantages and limitations. The use of a particular test is often determined by the 0014-4894/$ - see front matter Ó 2008 Elsevier Inc. All rights reserved. doi:10.1016/j.exppara.2008.09.013 * Corresponding author. Fax: +58 2126719289. E-mail address: memendoza@cantv.net (M. Mendoza). Experimental Parasitology 121 (2009) 1–7 Contents lists available at ScienceDirect Experimental Parasitology journal homepage: www.elsevier.com/locate/yexpr