First report of the presence of Trypanosoma evansi in dogs from Paraguay applying molecular techniques

Main Article Content

Oscar Daniel Salvioni Recalde
Stefanía Fraenkel
María José Tintel
Verónica Paola Arze
Natalia Ramírez Centurión
Miriam Rolón
Celeste Vega Gómez


Surra is a disease caused by the hemoflagellate pathogen Trypanosoma evansi which affects a wide variety of mammals. The only cases reported of the presence of T. evansi in Paraguay were identified in samples from horses and capybaras and they were detected by light microscopy in the 19th century. The aim of this study is to report three autochthonous cases of canine trypanosomiasis caused by T. evansi, for the first time in the country, using molecular techniques and their application for the differential diagnosis of trypanosomatids species. The technique implemented was real-time PCR-HRM, amplifying a fragment of the hsp70 gene, using a pair of primers initially used to discriminate Leishmania species. This is the first report on the usage of these primers to detect T. evansi through HRM analysis, which allows the differentiation of trypanosomatids species simultaneously, making it an efficient tool for differential diagnosis.

Article Details

How to Cite
Salvioni Recalde, O. D. ., Fraenkel, S. ., José Tintel, M., Arze, V. P. ., Ramírez Centurión, N. ., Rolón, M. ., & Vega Gómez, C. . (2021). First report of the presence of Trypanosoma evansi in dogs from Paraguay applying molecular techniques. Brazilian Journal of Veterinary Medicine, 43(1), e001920.
Case report


Aquino, L. P. C. T., Machado, R. Z., Alessi, A. C., Santana, A. E., Castro, M. B., Marques, L. C., & Malheiros, E. B. (2002). Hematological, biochemical and anatomopathological aspects of the experimental infection with Trypanosoma evansi in dogs. Arquivo Brasileiro de Medicina Veterinária e Zootecnia, 54(1), 8-18. http://dx.doi. org/10.1590/S0102-09352002000100002.

Aquino, L. P., Machado, R. Z., Alessi, A. C., Marques, L. C., de Castro, M. B., & Malheiros, E. B. (1999). Clinical, parasitological and immunological aspects of experimental infection with Trypanosoma evansi in dogs. Memorias do Instituto Oswaldo Cruz, 94(2), 255-260. PMid:10224539.

Aref, M., Yasin, S. M., Bahear, W., Ghulam, Z., Hastie, L., Dennison, T., Schauwers, W., Büscher, P., & Deborggraeve, S. (2013). Canine Trypanosoma evansi infection in Afghanistan. Veterinary Parasitology, 197(3-4), 638-641. PMid:23993635.

Bachmann, A. O. (2012). A catalog of the types of Stratiomyidae, Tabanidae, Asilidae sensu lato, and Syrphidae (Insecta, Diptera) deposited in the Museo Argentino de Ciencias Naturales, Buenos Aires. Revista Del Museo Argentino de Ciencias Naturales, Nueva Serie, 14(1), 97–123.

Banowary, B., Dang, V. T., Sarker, S., Connolly, J. H., Chenu, J., Groves, P., Ayton, M., Raidal, S., Devi, R., Vanniasinkam, T., & Ghorashi, S. (2015). Differentiation of Campylobacter jejuni and Campylobacter coli Using Multiplex-PCR and High Resolution Melt Curve Analysis. PLoS One, 10(9), e0138808. pone.0138808. PMid:26394042.

Bono Battistoni, M. F., Orcellet, V., Peralta, J. L., Marengo, R., Plaza, D., Brunini, A., Ruiz, M., Widenhorn, N., Sanchez, A., Monje, L., & Cignetti, L. (2016). First report of Trypanosoma evansi in a canine in Argentina. Veterinary Parasitology, Regional Studies and Reports, 6, 1-3. PMid:31014521.

Borst, P., Fase-Fowler, F., & Gibson, W. C. (1987). Kinetoplast DNA of Trypanosoma evansi. Molecular and Biochemical Parasitology, 23(1), 31-38. PMid:3033499.

Brun, R., Hecker, H., & Lun, Z. R. (1998). Trypanosoma evansi and T. equiperdum: Distribution, biology, treatment and phylogenetic relationship (a review). Veterinary Parasitology, 79(2), 95-107. S0304-4017(98)00146-0. PMid:9806490.

Colpo, C. B., Monteiro, S. G., Stainki, D. R., Colpo, E. T. B., & Henriques, G. B. (2005). Infecção natural por Trypanosoma evansi em cães. Ciência Rural, 35(3), 717-719.

Correa-Salgado, A. M., Pacheco de Araujo, F. A., & Cañón-Franco, W. A. (2010). Infección Natural por Trypanosoma evansi en Canino, Manizales - Colombia : Caso clínico. Revista Ibero-latinoamericana de Parasitología, 69(1), 98-100.

Desquesnes, M. (2004). Livestock Trypanosomoses and their vectors in Latin America (174 p.). World Organization for Animal Health.

Desquesnes, M., Dargantes, A., Lai, D. H., Lun, Z. R., Holzmuller, P., & Jittapalapong, S. (2013a). Trypanosoma evansi and surra: A review and perspectives on transmission, epidemiology and control, impact, and zoonotic aspects. BioMed Research International, 321237, 321237. PMid:24151595.

Desquesnes, M., Holzmuller, P., Lai, D. H., Dargantes, A., Lun, Z. R., & Jittaplapong, S. (2013b). Trypanosoma evansi and surra: A review and perspectives on origin, history, distribution, taxonomy, morphology, hosts, and pathogenic effects. BioMed Research International, 2013, 1-22. PMid:24024184.

Dwight, Z., Palais, R., & Wittwer, C. T. (2011). uMELT: Prediction of high-resolution melting curves and dynamic melting profiles of PCR products in a rich web application. Bioinformatics (Oxford, England), 27(7), 1019-1102. PMid:21300699.

Eberhardt, A. T., Monje, L. D., Zurvera, D. A., & Beldomenico, P. M. (2014). Detection of Trypanosoma evansi infection in wild capybaras from Argentina using smear microscopy and real-time PCR assays. Veterinary Parasitology, 202(3-4), 226-233. PMid:24636712.

Elmassian, M. (1901). Mal de caderas. In Conférence Faite au Conseil National d’Hygiène le 19 mai. Anales de la Universidad Nacional: Vol. 16. Universidad Nacional.

Elmassian, M. (1902). Mal de caderas, flagelosis paresiante de los equideos. Revista de la Sociedad Médica Argentina, 10(53), 122-148.

Elmassian, M., & Migone, L. E. (1903). Sur le mal de caderas ou flagellose parésiante des équidés sud-américains. Annales de l’Institut Pasteur, 17(4), 241-267.

El-Metanawey, T. M., El-Beih, N. M., El-Aziz, M. M., Hassanane, M. S., & El-Aziz, T. H. (2009). Comparative studies on diagnosis of Trypanosoma evansi in experimentally infected goats. Global Veterinaria, 3, 348-353.

Ereqat, S., Nasereddin, A., Al-Jawabreh, A., Al-Jawabreh, H., Al-Laham, N., & Abdeen, Z. (2020). Prevalence of Trypanosoma evansi in livestock in Palestine. Parasites & Vectors, 13(1), 21. 020-3894-9. PMid:31931864.

Fernández, D., González-Baradat, B., Eleizalde, M., González-Marcano, E., Perrone, T., & Mendoza, M. (2009). Trypanosoma evansi: A comparison of PCR and parasitological diagnostic tests in experimentally infected mice. Experimental Parasitology, 121(1), 1-7. PMid:18848544.

Fowler, M. E. (2010). Parasites. In M.E. Fowler. Medicine and surgery of Camelids (chap. 8, pp. 231-269). Wiley- Blackwell.

Franciscato, C., Lopes, S. T. A., Teixeira, M. M. G., Monteiro, S. G., Wolkmer, P., Garmatz, B. C., & Paim, C. B. (2007). Cão naturalmente infectado por Trypanosoma evansi em Santa Maria, RS, Brasil. Ciência Rural, 37(1), 288-291.

Franke, C. R., Greiner, M., & Mehlitz, D. (1994). Investigations on naturally occurring Trypanosoma evansi infections in horses, cattle, dogs and capybaras (Hydrochaeris hydrochaeris) in Pantanal de Poconé (Mato Grosso, Brazil). Acta Tropica, 58(2), 159-169. PMid:7887341.

Goldschmidt, P., Degorge, S., Che Sarria, P., Benallaoua, D., Semoun, O., Borderie, V., Laroche, L., & Chaumeil, C. (2012). New Strategy for Rapid Diagnosis and Characterization of Fungal Infections: The Example of Corneal Scrapings. PLoS One, 7(7), e37660. PMid:22768289.

Gonzales, J. L., Chacon, E., Miranda, M., Loza, A., & Siles, L. M. (2007). Bovine trypanosomosis in the Bolivian Pantanal. Veterinary Parasitology, 146(1-2), 9-16. PMid:17374452.

Greif, G., Faral-Tello, P., Scardoelli Vianna, C., Hernandez, A., Basmadjian, Y., & Robello, C. (2018). The first case report of trypanosomiasis caused by Trypanosoma evansi in Uruguay. Veterinary Parasitology, Regional Studies and Reports, 11, 19-21. PMid:31014612.

Hall, T. A. (1999). BioEdit: A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series, 41, 95-98.

Hernández, C., Alvarez, C., González, C., Ayala, M. S., León, C. M., & Ramírez, J. D. (2014). Identification of six New World Leishmania species through the implementation of a High-Resolution Melting (HRM) genotyping assay. Parasites & Vectors, 7(1), 501. PMid:25394418.

Herrera, H. M., Norek, A., Freitas, T. P. T., Rademaker, V., Fernandes, O., & Jansen, A. M. (2005). Domestic and wild mammals infection by Trypanosoma evansi in a pristine area of the Brazilian Pantanal region. Parasitology Research, 96(2), 121-126. PMid:15824901.

Hoare, C. A. (1965). Vampire bats as vectors and hosts of equine and bovine trypanosomes. Acta Tropica, 22(3), 204-216. PMid:4379528.

Hosseininejad, M., Shirani, D., Nabian, S., Nassiri, S. M., & Mazaheri, R. (2007). Trypanosoma evansi in three dogs in Iran. Comparative Clinical Pathology, 16(1), 69-71.

Jaimes-Dueñez, J., Triana-Chávez, O., Valencia-Hernández, A., Sánchez-Arévalo, D., Poche-Ceballos, A., Ortíz-Álvarez, J., & Mejía-Jaramillo, A. M. (2017). Molecular diagnosis and phylogeographic analysis of Trypanosoma evansi in dogs (Canis lupus familiaris) suggest an epidemiological importance of this species in Colombia. Preventive Veterinary Medicine, 139(Pt A), 82-89. PMid:28364836.

Joste, V., Kamaliddin, C., Kendjo, E., Hubert, V., Argy, N., & Houzé, S. (2018). Distinction of Plasmodium ovale wallikeri and Plasmodium ovale curtisi using quantitative Polymerase Chain Reaction with High Resolution Melting revelation. Scientific Reports, 8(1), 300. PMid:29321578.

Kagkli, D. M., Folloni, S., Barbau-Piednoir, E., Van den Eede, G., & Van den Bulcke, M. (2012). Towards a pathogenic escherichia coli detection platform using multiplex SYBR®green Real-Time PCR methods and high resolution melting analysis. PLoS One, 7(6), e39287. PMid:22761753.

Keatley, S., Botero, A., Fosu-Nyarko, J., Pallant, L., Northover, A., & Thompson, R. C. A. (2020). Species-level identification of trypanosomes infecting Australian wildlife by High-Resolution Melting-Real Time Quantitative Polymerase Chain Reaction (HRM-qPCR). International Journal for Parasitology. Parasites and Wildlife, 13, 261-268. PMid:33294365.

Konnai, S., Mekata, H., Mingala, C. N., Abes, N. S., Gutierrez, C. A., Herrera, J. R. V., Dargantes, A. P., Witola, W. H., Cruz, L. C., Inoue, N., Onuma, M., & Ohashi, K. (2009). Development and application of a quantitative real-time PCR for the diagnosis of Surra in water buffaloes. Infection, Genetics and Evolution, 9(4), 449-452. http:// PMid:19460309.

Lee, T.-H., Wu, T.-S., Tseng, C.-P., & Qiu, J. T. (2012). High-resolution melting molecular signatures for rapid identification of human Papillomavirus genotypes. PLoS One, 7(8), e42051. pone.0042051. PMid:22916117.

Lun, Z.R., & Desser, S. S. (1995). Is the broad range of hosts and geographical distribution of Trypanosoma evansi attributable to the loss of Maxicircle Kinetoplast DNA? Parasitology Today (Personal Ed.), 11(4), 131-133. http:// PMid:15275353.

Migone, L. E. (1910). Le role des carpinchos comme reservoir de virus dans la conservation du Mal de Caderas. Bulletin de la Société de Pathologie Exotique, 3, 524-525.

Morrone, J. J., & Coscarón, S. (1998). Biodiversidad de artrópodos argentinos: una perspectiva biotaxonómica. Ediciones Sur: Buenos Aires, Argentina.

Muñoz, K., & Chávez, A. (2001). Trypanosoma evansi Isolated from Capybara (Hidrochaeris hidrochaeris). Memorias do Instituto Oswaldo Cruz, 96(7), 945-946. PMid:11685258.

Njiru, Z. K., Constantine, C. C., Guya, S., Crowther, J., Kiragu, J. M., Thompson, R. C., & Dávila, A. M. (2005). The use of ITS1 rDNA PCR in detecting pathogenic African trypanosomes. Parasitology Research, 95(3), 186-192. PMid:15619129.

Nunes, V. L. B., & Oshiro, E. T. (1990). Trypanosoma (Trypanozoon) evansi in the coati from the Pantanal reigon of Mato Grosso do Sul State, Brazil. Transactions of the Society of Tropical Medicine and Hygiene, 84(5), 692. PMID: 2278072.

Ramírez-Iglesias, J. R., Eleizalde, M. C., Gómez-Piñeres, E., & Mendoza, M. (2011). Trypanosoma evansi: A comparative study of four diagnostic techniques for trypanosomosis using rabbit as an experimental model. Experimental Parasitology, 128(1), 91-96. PMid:21320490.

Sharma, P., Juyal, P. D., Singla, L. D., Chachra, D., & Pawar, H. (2012). Comparative evaluation of real time PCR assay with conventional parasitological techniques for diagnosis of Trypanosoma evansi in cattle and buffaloes. Veterinary Parasitology, 190(3-4), 375-382. PMid:22858228.

Silva, R. A. M. S., Arosemena, N. A. E., Herrera, H. M., Sahib, C. A., & Ferreira, M. S. J. (1995). Outbreak of trypanosomosis due to Trypanosoma evansi in horses of Pantanal Mato-grossense, Brazil. Veterinary Parasitology, 60(1-2), 167-171. PMid:8644453.

Stevens, J. R., Nunes, V. L. B., Lanham, S. M., & Oshiro, E. T. (1989). Isoenzyme characterization of Trypanosoma evansi isolated from capybaras and dogs in Brazil. Acta Tropica, 46(4), 213-222. 706X(89)90021-1. PMid:2571248.

Strickman, D. (1982). Notes on Tabanidae (Diptera) from Paraguay. Journal of Medical Entomology, 19(4), 399–402.

Woo, P. T. K. (1977). Salivarian trypanosomes producing disease in livestock outside of sub-Saharan Africa. In J. P. Kreier. Parasitic Protozoa (pp. 269-296). Academic Press.

Zampieri, R. A., Laranjeira-Silva, M. F., Muxel, S. M., Stocco de Lima, A. C., Shaw, J. J., & Floeter-Winter, L. M. (2016). High resolution melting analysis targeting HSP70 as a fast and efficient method for the discrimination of Leishmania species. PLoS Neglected Tropical Diseases, 10(2), e0004485. pntd.0004485. PMid:26928050.