PCR for the diagnosis of Trypanosoma species in livestock: sample preparation to increase sensitivity

Desquesnes Marc, Tresse Laurent. 1999. PCR for the diagnosis of Trypanosoma species in livestock: sample preparation to increase sensitivity. In : Proceedings of first symposium on new world trypanosomes = [Actes du 1er symposium sur les trypanosomes d'Amérique]. Vokaty S. (ed.), Desquesnes M. (ed.). Bridgetown : IICA, pp. 121-127. Symposium on New World Trypanosomes. 1, Georgetown, Guyana, 20 November 1996/22 November 1996.

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Autre titre : La réaction en chaîne de la polymérase (PCR) pour diagnostiquer les espèces animales Trypanosoma : préparation d'échantillons pour accroître la sensibilité

Abstract : #T. vivax# DNA detection polymerase chain reaction (PCR) appears to be an alternative for a species-specific diagnosis of #T. vivax# active infections in livestock. PCR highly sensitive when processed on purified DNA from DNA extraction with ethanol/chloroform, but this technique is too expensive and time-consuming to be applied for diagnosis. First evaluation of the PCR directly applied for the diagnosis of #T. vivax# in cattle indicated a low sensitivity about 10(3) parasites/ml. Sensitivity of several techniques of low cost were evaluated with determined parasitaemia. 22 blood samples containing known numbers of #T. vivax#/ml, were prepared by dilution of infected sheep blood into blood from a non infected sheep. Sensivity of the PCR was evaluated in the following blood sample preparations: heparinized blood, plasma, lysed blood, burly coat from haematocrit capillary tubes, pellet from plasma centrifugation, and DNA purified with a commercial ion exchange resin. Most often, crude heparinized blood inhibited the PCR. Sensitivity of PCR with plasma and lysed blood was low, around 450 parasites/ml. PCR on buffy coat was more sensitive, but products were sometimes hardly visible. Pellet of plasma centrifugation an original, fast and economic preparation, which presented a high sensity: 100% of the samples were positive when the parasitaemia was over 13 parasites/ml. DNA purification is slightly more time-consuming and expensive, since it procedes from several manipulations and the use of a kit, but it appeared to be the most sensitive technique among those investigated

Mots-clés Agrovoc : Trypanosoma vivax, PCR, Diagnostic, Plasma sanguin, Méthode

Classification Agris : L72 - Pests of animals
L73 - Animal diseases

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