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Acta Parasitologica

, Volume 60, Issue 2, pp 211–217 | Cite as

Molecular diagnosis of natural fasciolosis by DNA detection in sheep faeces

  • Silvana Carnevale
  • María Laura Pantano
  • Laura Kamenetzky
  • Jorge Bruno Malandrini
  • Claudia Cecilia Soria
  • Jorge Néstor Velásquez
Article

Abstract

Fasciolosis is an important parasitic zoonosis considered the most important helminth infection of ruminants in tropical countries. The aim of this study was to develop a PCR assay for the sensitive and specific detection of F. hepatica in formalin preserved sheep faeces. A 405-bp fragment of the cytochrome c oxidase subunit 1 gene of F. hepatica was amplified from stool samples of infected sheep. The PCR assay showed a detection limit of 20 pg of F. hepatica DNA. No cross-reactions were observed with samples containing coccidian oocysts or gastrointestinal nematodes eggs. Our PCR technique showed to be effective for specific detection of F. hepatica infections in sheep.

Keywords

Fasciola hepatica PCR faeces 

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References

  1. Ai L., Dong S.J., Zhang W.Y., Elsheikha H.M., Mahmmod Y.S., Lin R.Q., Yuan Z.G., Shi Y.L., Huang W.Y., Zhu X.Q. 2010. Specific PCR-based assays for the identification of Fasciola species: their development, evaluation and potential usefulness in prevalence surveys. Annals of Tropical Medical Parasitology, 104, 65–72. DOI: 10.1179/136485910X12607012 373713CrossRefGoogle Scholar
  2. Anderson N., Luong T.T., Vo N.G., Bui K.L., Smooker P.M., Spithill T.W. 1999. The sensitivity and specificity of two methods for detecting Fasciola infections in cattle. Veterinary Parasitology, 83, 15–24CrossRefGoogle Scholar
  3. Arias M.S., Pineiro P., Sanchez-Andrade R., Suarez J.L., Hillyer G.V., Diez-Banos P., Paz-Silva A., Morrondo P. 2013. Relationship between exposure to Fasciola hepatica in roe deer (Capreolus capreolus) and cattle extensively reared in an endemic area. Research in Veterinary Science, 95, 1031–1035. DOI: 10.1016/j.rvsc.2013.07.027CrossRefGoogle Scholar
  4. Ashrafi K., Valero M.A., Panova M., Massoud J., Mas-Coma S. 2006. Phenotypic analysis of adults of Fasciola hepatica, Fasciola gigantica and intermediate forms from the endemic region of Gilan, Iran. Parasitology International, 55, 249–260CrossRefGoogle Scholar
  5. Boch J., Supperer, R. 1988. Parasitología en Medicina Veterinaria. Editorial Hemisferio Sur, Buenos Aires, ArgentinaGoogle Scholar
  6. Boero J.J. 1976. Parasitosis Animales. EUDEBA, Buenos Aires, ArgentinaGoogle Scholar
  7. Bretagne S., Guillou J.P., Morand M., Houin R. 1993. Detection of Echinococcus multilocularis DNA in fox faeces using DNA amplification. Parasitology, 106, 193–199CrossRefGoogle Scholar
  8. Carnevale S., Rodriguez M.I., Guarnera E.A., Carmona C., Tanos T., Angel S. 2001. Immunodiagnosis of fasciolosis using recombinant procathepsin L cystein proteinase. Diagnostic Microbiology and Infectious Diseases, 41, 43–49CrossRefGoogle Scholar
  9. Carnevale S., Velasquez J.N., Labbe J.H., Chertcoff A., Cabrera M.G., Rodriguez M.I. 2000. Diagnosis of Enterocytozoon bieneusi by PCR in stool samples eluted from filter paper disks. Clinical and Diagnostic Laboratory Immunology, 7, 504–506PubMedPubMedCentralGoogle Scholar
  10. Castro E., Freyre A., Hernandez Z. 2000. Serological responses of cattle after treatment and during natural re-infection with Fasciola hepatica, as measured with a dot-ELISA system. Veterinary Parasitology, 90, 201–208CrossRefGoogle Scholar
  11. Chang Y.T., Loew G.H. 1994. Reaction mechanisms of formal dehyde with endocyclic imino groups of nucleic acid bases. Journal of the American Chemical Society, 11, 3548–3555CrossRefGoogle Scholar
  12. Choubisa S.L., Jaroli V.J. 2013. Gastrointestinal parasitic infection in diverse species of domestic ruminants inhabiting tribal rural areas of southern Rajasthan, India. Journal of Parasitic Diseases, 37, 271–275. DOI: 10.1007/s12639-012-0178-0CrossRefGoogle Scholar
  13. Cornelissen J.B.W.J., de Leeuw W.A., van der Heijden P.J. 1992. Comparison of an indirect haemagglutination assay and an ELISA for diagnosing Fasciola hepatica in experimentally and naturally infected sheep. Veterinary Quarterly, 14, 152–156CrossRefGoogle Scholar
  14. Cucher M.A., Carnevale S., Prepelitchi L., Labbe J.H., Wisnivesky-Colli C. 2006. PCR diagnosis of Fasciola hepatica in fieldcollected Lymnaea columella and Lymnaea viatrix snails. Veterinary Parasitology, 137, 74–82CrossRefGoogle Scholar
  15. Dinkel A., Von Nickisch-Rosenegk M., Bilger B., Merli M., Lucius R., Roming T. 1998. Detection of Echinococcus multilocularis in the definitive host: coprodiagnosis by PCR as an alternative to necropsy. Journal of Clinical Microbiology, 36, 1871–1876PubMedPubMedCentralGoogle Scholar
  16. Domke A.V., Chartier C., Gjerde B., Leine N., Vatn S., Stuen S. 2013. Prevalence of gastrointestinal helminths, lungworms and liver fluke in sheep and goats in Norway. Veterinary Parasitology, 194, 40–48. DOI: 10.1016/j.vetpar.2012.12.023CrossRefGoogle Scholar
  17. Duenngai K., Boonmars T., Sithithaworn J., Sithithaworn P. 2013. Diagnosis of early infection and post chemotherapeutic treatment by copro-DNA detection in experimental opisthorchiasis. Parasitology Research, 112, 271–278. DOI: 10.1007/s00 436-012-3134-0CrossRefGoogle Scholar
  18. Dumenigo B.E., Espino A.M., Finlay C.M., Mezo M. 2000. Kinetics of antibody-based antigen detection in serum and faeces of sheep experimentally infected with Fasciola hepatica. Veterinary Parasitology, 89, 153–161CrossRefGoogle Scholar
  19. Dumenigo B.E., Mezo M. 1999. Monoclonal antibody sandwich immunoassay detection of coproantigen to evaluate the efficacy of treatment in natural ovine fasciolosis. Research in Veterinary Science, 66, 165–167CrossRefGoogle Scholar
  20. El-Bahi M.M., Malone J.B., Todd W.J., Schnorr, K.L. 1992. Detection of stables diagnostic antigen from bile and feces of Fasciola hepatica infected cattle. Veterinary Parasitology, 45, 157–167CrossRefGoogle Scholar
  21. Gonzalez-Lanza C., Manga-Gonzalez Y., Del-Pozo-Carnero P., Hidalgo-Arguello R. 1989. Dynamics of elimination of the eggs of Fasciola hepatica (Trematoda, Digenea) in faeces of cattle in the Porma Basin. Spain Veterinary Parasitology, 34, 35–43CrossRefGoogle Scholar
  22. Gorman T., Valdes A., Fredes F., Ferreira A., Alcaino H. 1995. Reconocimiento antigenico de ovinos naturalmente infectados con Fasciola hepatica, monitoreado a traves de inmunoelectrotransferencia enzimatica (Westerm Blotting). Archivos de Medicina Veterinaria XXVII (2), 33–40Google Scholar
  23. Happich F.A., Boray J.C. 1969. Quantitative diagnosis of chronic fasciolosis. Australian Veterinary Journal, 45, 326–328CrossRefGoogle Scholar
  24. Hillyer G.V., Soler de Galanes M. 1991. Initial feasibility studies of the FAST-ELISA for the immunodiagnosis of fascioliasis. Journal of Parasitology, 77, 362–365CrossRefGoogle Scholar
  25. Kozłowska-Łoj J., Łoj-Maczulska A. 2013. The prevalence of Fasciola hepatica L. infection in cattle in the Lublin province in the years 2009–2012. Annals of Parasitology 59, 207–208PubMedGoogle Scholar
  26. Kuk S., Yazar S., Cetinkaya U. 2012. Stool sample storage conditions for the preservation of Giardia intestinalis DNA. Memorias do Instituto Oswaldo Cruz 107, 965–968.Google Scholar
  27. Langley R.J., Hillyer G.V. 1989. Detection of circulating immune complexes by the enzyme-linked immunosorbent assay in sera from cattle infected with Fasciola hepatica. Journal of Parasitology, 75, 690–695CrossRefGoogle Scholar
  28. Lelievre S., Verrez-Bagnis V., Jerome M., Vaz S. 2010. PCR-RFLP analyses of formalin-fixed fish eggs for the mapping of spawning areas in the Eastern Channel and Southern North Sea. Journal of Plankton Research, 32, 1527–1539. DOI: 10.1093/plankt/fbq067CrossRefGoogle Scholar
  29. Locatelli A., Genchi C., Sartorelli P. 1984. Compendio di Parassitologia Veterinaria. Calderini, Bologna, ItaliaGoogle Scholar
  30. Mage C., Bourgne H., Toullieu J.M., Rondelaud D., Dreyfuss G. 2002. Fasciola hepatica and Paramphistomum daubneyi: changes in prevalences of natural infections in cattle and in Lymnaea truncatula from central France over the past 12 years. Veterinary Research, 33, 439–447CrossRefGoogle Scholar
  31. Maniatis T., Fritsch E.F., Sambrook J. 1989. Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, USAGoogle Scholar
  32. Martinez A., Martinez-Cruz M.S., Martinez F.J., Gutierrez P.N., Hernandez S. 1996. Detection of antibodies to Fasciola hepatica excretory-secretory antigens in experimentally infected goats by enzyme immunosorbent assay. Veterinary Parasitology, 62, 247–252CrossRefGoogle Scholar
  33. Martinez-Perez J.M., Robles-Perez D., Rojo-Vazquez F.A., Martinez-Valladares M. 2012. Comparison of three different techniques to diagnose Fasciola hepatica infection in experimentally and naturally infected sheep. Veterinary Parasitology, 190, 80–86. DOI: 10.1016/j.vetpar.2012.06.002CrossRefGoogle Scholar
  34. Martinez-Valladares M., Cordero-Perez C., Castanon-Ordonez L., Famularo M.R., Fernandez-Pato N., Rojo-Vazquez F.A. 2010. Efficacy of a moxidectin/triclabendazole oral formulation against mixed infections of Fasciola hepatica and gastrointestinal nematodes in sheep. Veterinary Parasitology, 174, 166–169CrossRefGoogle Scholar
  35. Martinez-Valladares M., Robles-Perez D., Martinez-Perez J.M., Cordero-Perez C., Famularo Mdel R., Fernandez-Pato N., Gonzalez-Lanza C., Castanon-Ordonez L., Rojo-Vazquez F.A. 2013. Prevalence of gastrointestinal nematodes and Fasciola hepatica in sheep in the northwest of Spain: relation to climatic conditions and/or man-made environmental modifications. Parasites and Vectors, 6, 282. DOI: 10.1186/1756-3305-6-282CrossRefGoogle Scholar
  36. Mas-Coma S., Bargues M.D., Valero M.A. 2005. Fascioliasis and other plant-borne trematode zoonoses. International Journal for Parasitology, 35, 1255–1278CrossRefGoogle Scholar
  37. Mas-Coma S., Valero, M.A., Bargues M.D. 2009. Fasciola, lymnaeids and human fascioliasis, with a global overview on disease transmission, epidemiology, evolutionary genetics, molecular epidemiology and control. Advances in Parasitology 69, 41–146. DOI: 10.1016/S0065-308X(09)69002-3CrossRefGoogle Scholar
  38. Mathis A., Deplazes P. 2006. Copro-DNA tests for diagnosis of animal taeniid cestodes. Parasitology International 55, S87–S90CrossRefGoogle Scholar
  39. Naidich A., McManus D.P., Canova S.G., Gutierrez A.M., Zhang W., Guarnera E.A., Rosenzvit M. C. 2006. Patent and pre-patent detection of Echinococcus granulosus genotypes in the definitive host. Molecular and Cellular Probes, 20, 5–10CrossRefGoogle Scholar
  40. Ngategize P.K, Bekele T., Tilahun G. 1993. Financial losses caused by ovine fasciolosis in the Ethiopian highlands. Tropical Animal Health and Production, 25, 155–161.CrossRefGoogle Scholar
  41. O’Neill S.M., Brady M.T., Callanan J.J., Mulcahy G., Joyce P., Mills K.H., Dalton J.P. 2000. Fasciola hepatica infection downregulates Th1 responses in mice. Parasite Immunology, 22, 147–155CrossRefGoogle Scholar
  42. Olaechea F.V. 1994. Epidemiologia y control de Fasciola hepatica en la Argentina. In (Eds. A. Nari and C. Fiel) Enfermedades Parasitarias de Importancia Económica en Bovinos. Editorial Agropecuaria, Hemisferio Sur SRL, Montevideo, Uruguay, 213–232Google Scholar
  43. Olaechea, F.V. 1995. Fasciola hepatica en bovinos. El tema elegido del mes: Fasciola hepatica. INTA, Sistema Regional de Salud Animal (SIRSA), Estacion Experimental Agropecuaria Bariloche, Unidad de Salud Animal, Bariloche, ArgentinaGoogle Scholar
  44. Pfister K., Daveau C., Ambroise-Thomas P. 1984. Partial purification of somatic and excretory-secretory products of adult Fasciola hepatica and their application for the serodiagnosis of experimental and natural fascioliasis using an ELISA. Research in Veterinary Science, 37, 39–43CrossRefGoogle Scholar
  45. Rapsch C., Schweizer G., Grimm F., Kohler L., Bauer C., Deplazes P., Braun U., Torgerson P.R. 2006. Estimating the true prevalence of Fasciola hepatica in cattle slaughtered in Switzerland in the absence of an absolute diagnostic test. International Journal for Parasitology, 36, 1153–1158CrossRefGoogle Scholar
  46. Reichel M.P. 2002. Performance characteristics of an enzyme-linked immunosorbent assay for the detection of liver fluke (Fasciola hepatica) infection in sheep and cattle. Veterinary Parasitology, 107, 65–72CrossRefGoogle Scholar
  47. Richter B., Ladinig A., Nedorost N., Weissenbock H. 2010. A Taq-Man quantitative polymerase chain reaction assay for the detection of Lawsonia intracellularis in fecal and tissue samples from pigs. Journal of Veterinary Diagnostic Investigation, 22, 70–73. DOI: 10.1177/104063871002200112CrossRefGoogle Scholar
  48. Rickard L.G. 1995. Development and application of a dot-ELISA test for the detection of serum antibodies to Fasciola hepatica antigens in llamas. Veterinary Parasitology, 58, 9–15CrossRefGoogle Scholar
  49. Robles-Perez D., Martinez-Perez J.M., Rojo-Vazquez F.A., Martinez-Valladares M. 2013. The diagnosis of fasciolosis in feces of sheep by means of a PCR and its application in the detection of anthelmintic resistance in sheep flocks naturally infected. Veterinary Parasitology, 197, 277–282. DOI: 10. 1016/j.vetpar.2013.05.006CrossRefGoogle Scholar
  50. Rodriguez-Perez J., Hillyer G.V. 1995. Detection of excretorysecretory circulating antigens in sheep infected with Fasciola hepatica and with Schistosoma mansoni and Fasciola hepatica. Veterinary Parasitology, 56, 57–66CrossRefGoogle Scholar
  51. Rojo-Vazquez F.A., Meana A., Valcarcel F., Martinez-Valladares M. 2012. Update on trematode infections in sheep. Veterinary Parasitology, 189, 15–38CrossRefGoogle Scholar
  52. Santiago N., Hillyer G.V. 1988. Antibody profiles by EITB and ELISA of cattle and sheep infected with Fasciola hepatica. Journal of Parasitology, 74, 810–818CrossRefGoogle Scholar
  53. SENASA. 1998. Manual de Procedimientos para la Inspección Veterinaria, MP3. SENASA, ArgentinaGoogle Scholar
  54. Serra-Freire N.M., Bordin E.L., Lessa C.S.S., Scherer P.O., Farias M.T., Malacco M.A., Correa T.C., Tschumi J.A. 1995. Reinvestigacao sobre a distribuicao da Fasciola hepatica no Brasil. A Hora Veterinária, 1, 19–21Google Scholar
  55. Skage M., Schander C. 2007. DNA from formalin-fixed tissue: extraction or repair? That is the question. Marine Biology Research, 3, 289–295. DOI: 10.1080/17451000701473942CrossRefGoogle Scholar
  56. Spithill T.W., Dalton J.P. 1998. Progress in development of liver fluke vaccines. Parasitology Today, 14, 224–228CrossRefGoogle Scholar
  57. Stoll N.R., Hausheer W.C. 1926. Concerning two options in dilution egg counting: small drop and displacement. American Journal of Hygiene, 6, 134–145Google Scholar
  58. Taylor S.M. 1989. Control of fasciolosis in the British Isles. Magyar Allartorv Lapja, 44, 651–655Google Scholar
  59. Telemann W. 1908. Eine methode zur erleichterung der auffindung von parasiteneiern in den faeces. Deutsche Medizinische Wochenschrift, 35, 1510–1511CrossRefGoogle Scholar
  60. Torgerson P., Claxton J. 1999. Epidemiology and control. In: (Ed. J. P. Dalton) Fasciolosis. CAB International Publishing, Wallingford, Oxon, UK, 113–149Google Scholar
  61. Tum S., Puotinen M.L., Skerratt L.F., Chan B., Sothoeun S. 2007. Validation of a geographic information system model for mapping the risk of fasciolosis in cattle and buffaloes in Cambodia. Veterinary Parasitology, 143, 364–367CrossRefGoogle Scholar
  62. Velasquez J.N., Osvaldo G.A., di Risio C., Etchart C., Chertcoff A.V., Perisse G.E., Carnevale S. 2011. Molecular characterization of Cystoisospora belli and unizoite tissue cyst in patients with Acquired Immunodeficiency Syndrome. Parasitology, 138, 279–286. DOI: 10.1017/S0031182010001253CrossRefGoogle Scholar
  63. Velasquez J.N., Carnevale S., Guarnera E.A., Labbe J.H., Chertcoff A., Cabrera M.G., Rodriguez M.I. 1996. Detection of the microsporidian parasite Enterocytozoon bieneusi in specimens from patients with AIDS by PCR. Journal of Clinical Microbiology, 34, 3230–3232PubMedPubMedCentralGoogle Scholar
  64. Velasquez J.N., Carnevale S., Oelemann W.M.R., Etchart C., Peralta J.M. 2005. Diagnosis of Enterocytozoon bieneusi by the polymerase chain reaction in archival fixed duodenal tissue. Enfermedades Infecciosas y Microbiología Clínica, 23, 218–220CrossRefGoogle Scholar
  65. Zimmerman G.L., Jen L.W., Cerro J.E., Farnsworth K.L., Wescott R.G. 1982. Diagnosis of Fasciola hepatica infections in sheep by an enzyme linked immunosorbent assay. American Journal of Veterinary Research, 43, 2097–2100PubMedGoogle Scholar

Copyright information

© Witold Stefański Institute of Parasitology, Polish Academy of Sciences 2015

Authors and Affiliations

  • Silvana Carnevale
    • 1
    • 2
  • María Laura Pantano
    • 1
  • Laura Kamenetzky
    • 3
  • Jorge Bruno Malandrini
    • 4
  • Claudia Cecilia Soria
    • 4
  • Jorge Néstor Velásquez
    • 5
  1. 1.Instituto Nacional de Enfermedades Infecciosas - ANLIS “Dr. Carlos G. Malbrán”Ciudad de Buenos AiresArgentina
  2. 2.Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET)Ciudad de Buenos AiresArgentina
  3. 3.Instituto de Microbiología y Parasitología Médica (IMPaM), UBA-CONICETCiudad de Buenos AiresArgentina
  4. 4.Facultad de Ciencias de la SaludUniversidad Nacional de CatamarcaSan Fernando del Valle de CatamarcaArgentina
  5. 5.Hospital Municipal de Infecciosas “Dr. Francisco Javier Muñiz”Ciudad de Buenos AiresArgentina

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