Hostname: page-component-8448b6f56d-tj2md Total loading time: 0 Render date: 2024-04-16T19:10:10.797Z Has data issue: false hasContentIssue false
  • English
  • Français

Identification of taeniid eggs in the faeces from carnivores based on multiplex PCR using targets in mitochondrial DNA

Published online by Cambridge University Press:  09 February 2007

D. TRACHSEL ,
P. DEPLAZES  and
A. MATHIS
D. TRACHSEL
Affiliation:
Institute of Parasitology, Vetsuisse and Medical Faculty, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland
P. DEPLAZES
Affiliation:
Institute of Parasitology, Vetsuisse and Medical Faculty, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland
A. MATHIS*
Affiliation:
Institute of Parasitology, Vetsuisse and Medical Faculty, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland
*
*Corresponding author. Tel: +41 (0)44 6358536. Fax: +41 (0)44 6358907. E-mail: alexander.mathis@access.unizh.ch
Get access Rights & Permissions [Opens in a new window]

Summary

A multiplex polymerase chain reaction (PCR) was evaluated for the identification of morphologically indistinguishable eggs of the taeniid tapeworms from carnivores using primers targeting mitochondrial genes. The primers for Echinococcus multilocularis (amplicon size 395 bp) were species-specific as assessed by in silico analysis and in the PCR using well-defined control samples. The design of primers that specifically amplify DNA from E. granulosus or Taenia spp. was not possible. The primers designed for E. granulosus also amplified DNA (117 bp) from E. vogeli, and those designed for Taenia spp. amplified products (267 bp) from species of Mesocestoides, Dipylidium and Diphyllobothrium. Nevertheless, as our diagnostic approach includes the concentration of taeniid eggs by sequential sieving and flotation, followed by their morphological detection, this non-specificity has limited practical importance. Sequence analysis of the corresponding amplicon can identify most of the described E. granulosus genotypes. Taenia spp. can be identified by direct sequencing of the 267 bp amplicon, or, for most species, by restriction fragment length polymorphism (RFLP) analysis. The multiplex PCR was readily able to detect 1 egg (estimated to contain 7000 targets, as determined by quantitative PCR). Having been validated using a panel of well-defined samples from carnivores with known infection status, this approach proved to be useful for the identification of taeniid eggs from both individual animals and for epidemiological studies.

Keywords

Echinococcus granulosusE. multilocularisTaenia sppmultiplex polymerase chain reaction (PCR)genotypesfaecal samplesquantitative PCRmitochondrial genes
Type
Research Article
Information
Parasitology , Volume 134 , Issue 6 , June 2007 , pp. 911 - 920
Copyright
Copyright © Cambridge University Press 2007

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Abbasi, I., Branzburg, A., Campos-Ponce, M., Abdel Hafez, S. K., Raoul, F., Craig, P. S. and Hamburger, J. (2003). Copro-diagnosis of Echinococcus granulosus infection in dogs by amplification of a newly identified repeated DNA sequence. American Journal of Tropical Medicine and Hygiene 69, 324–330.CrossRefGoogle ScholarPubMed
Bowles, J., Blair, D. and McManus, D. P. (1992). Genetic variants within the genus Echinococcus identified by mitochondrial DNA sequencing. Molecular and Biochemical Parasitology 54, 165–173.CrossRefGoogle ScholarPubMed
Bowles, J. and McManus, D. P. (1993). NADH dehydrogenase 1 gene sequences compared for species and strains of the genus Echinococcus. International Journal for Parasitology 23, 969–972.CrossRefGoogle ScholarPubMed
Cabrera, M., Canova, S., Rosenzvit, M. and Guarnera, E. (2002). Identification of Echinococcus granulosus eggs. Diagnostic Microbiology and Infectious Disease 44, 29–34.CrossRefGoogle ScholarPubMed
Christofi, G., Deplazes, P., Christofi, N., Tanner, I., Economides, P. and Eckert, J. (2002). Screening of dogs for Echinococcus granulosus coproantigen in a low endemic situation in Cyprus. Veterinary Parasitology 104, 299–306.CrossRefGoogle Scholar
Corpet, F. (1988). Multiple sequence alignment with hierarchical clustering. Nucleic Acids Research 16, 10881–10890.CrossRefGoogle ScholarPubMed
Craig, P. S., Macpherson, C. N. and Nelson, G. S. (1986). The identification of eggs of Echinococcus by immunofluorescence using a specific anti-oncospheral monoclonal antibody. American Journal of Tropical Medicine and Hygiene 35, 152–158.CrossRefGoogle ScholarPubMed
Deplazes, P., Jimenez-Palacios, S., Gottstein, B., Skaggs, J. and Eckert, J. (1994). Detection of Echinococcus coproantigens in stray dogs of northern Spain. Applied Parasitology 35, 297–301.Google ScholarPubMed
Dinkel, A., von Nickisch-Rosenegk, M., Bilger, B., Merli, M., Lucius, R. and Romig, T. (1998). Detection of Echinococcus multilocularis in the definitive host: coprodiagnosis by PCR as an alternative to necropsy. Journal of Clinical Microbiology 36, 1871–1876.CrossRefGoogle ScholarPubMed
Dinkel, A., Njoroge, E. M., Zimmermann, A., Walz, M., Zeyhle, E., Elmahdi, I. E., Mackenstedt, U. and Romig, T. (2004). A PCR system for detection of species and genotypes of the Echinococcus granulosus-complex, with reference to the epidemiological situation in eastern Africa. International Journal for Parasitology 34, 645–653.CrossRefGoogle Scholar
Eckert, J. and Deplazes, P. (2004). Biological, epidemiological and clinical aspects of echinococcosis: a zoonosis of increasing concern. Clinical Microbiology Reviews 17, 107–135.CrossRefGoogle ScholarPubMed
Gasser, R. B. and Chilton, N. B. (1995). Characterisation of taeniid cestode species by PCR-RFLP of ITS2 ribosomal DNA. Acta Tropica 59, 31–40.CrossRefGoogle ScholarPubMed
Gasser, R. B., Zhu, X. and Woods, W. (1999). Genotyping Taenia tapeworms by single-strand conformation polymorphism of mitochondrial DNA. Electrophoresis 20, 2834–2837.3.0.CO;2-F>CrossRefGoogle ScholarPubMed
Haag, K. L., Araujo, A. M., Gottstein, B., Siles-Lucas, M., Thompson, R. C. and Zaha, A. (1999). Breeding systems in Echinococcus granulosus (Cestoda; Taeniidae): selfing or outcrossing? Parasitology 118, 63–71.CrossRefGoogle ScholarPubMed
Hegglin, D., Ward, P. I. and Deplazes, P. (2003). Anthelmintic baiting of foxes against urban contamination with Echinococcus multilocularis. Emerging Infectious Diseases 9, 1266–1272.CrossRefGoogle ScholarPubMed
Kapel, C. M., Torgerson, P. R., Thompson, R. C. and Deplazes, P. (2006). Reproductive potential of Echinococcus multilocularis in experimentally infected foxes, dogs, raccoon dogs and cats. International Journal for Parasitology 36, 79–86.CrossRefGoogle ScholarPubMed
Kocher, T. D., Thomas, W. K., Meyer, A., Edwards, S. V., Paabo, S., Villablanca, F. X. and Wilson, A. C. (1989). Dynamics of mitochondrial DNA evolution in animals: amplification and sequencing with conserved primers. Proceedings of the National Academy of Sciences, USA 86, 6196–6200.CrossRefGoogle ScholarPubMed
Liu, L. X., Blaxter, M. L. and Shi, A. (1996). The 5S ribosomal RNA intergenic region of parasitic nematodes: variation in size and presence of SL1 RNA. Molecular and Biochemical Parasitology 83, 235–239.CrossRefGoogle ScholarPubMed
Longo, M. C., Berninger, M. S. and Hartley, J. L. (1990). Use of uracil DNA glycosylase to control carry-over contamination in polymerase chain reactions. Gene 93, 125–128.CrossRefGoogle ScholarPubMed
Mathis, A., Deplazes, P. and Eckert, J. (1996). An improved test system for PCR-based specific detection of Echinococcus multilocularis eggs. Journal of Helminthology 70, 219–222.CrossRefGoogle ScholarPubMed
Mathis, A. and Deplazes, P. (2006). Copro-DNA tests for diagnosis of animal taeniid cestodes. Parasitology International 55, S87–90.CrossRefGoogle ScholarPubMed
McManus, D. P., Le, T. H. and Blair, D. (2004). Genomics of parasitic flatworms. International Journal for Parasitology 34, 153–158.CrossRefGoogle ScholarPubMed
McManus, D. P. and Bowles, J. (1996). Molecular genetic approaches to parasite identification: their value in diagnostic parasitology and systematics. International Journal for Parasitology 26, 687–704.CrossRefGoogle ScholarPubMed
Obwaller, A., Schneider, R., Walochnik, J., Gollackner, B., Deutz, A., Janitschke, K., Aspock, H. and Auer, H. (2004). Echinococcus granulosus strain differentiation based on sequence heterogeneity in mitochondrial genes of cytochrome c oxidase-1 and NADH dehydrogenase-1. Parasitology 128, 569–575.CrossRefGoogle ScholarPubMed
Schantz, P. M. (1997). Sources and uses of surveillance data for cystic echinococcosis. In Compendium on Cystic Echinococcosis in Africa and in Middle Eastern Countries with Special Reference to Morocco. (ed. Anderson, F. L., Ouhelli, H. and Kachani, M.), pp. 72–84. Brigham Young University Print Services, Provo, Utah, USA.Google Scholar
Scott, J. C. and McManus, D. P. (1994). The random amplification of polymorphic DNA can discriminate species and strains of Echinococcus. Tropical Medicine and Parasitology 45, 1–4.Google ScholarPubMed
Stefanic, S., Shaikenov, B. S., Deplazes, P., Dinkel, A., Torgerson, P. R. and Mathis, A. (2004). Polymerase chain reaction for detection of patent infections of Echinococcus granulosus (“sheep strain”) in naturally infected dogs. Parasitology Research 92, 347–351.CrossRefGoogle Scholar
Stieger, C., Hegglin, D., Schwarzenbach, G., Mathis, A. and Deplazes, P. (2002). Spatial and temporal aspects of urban transmission of Echinococcus multilocularis. Parasitology 124, 631–640.CrossRefGoogle ScholarPubMed
Thompson, R. C. and McManus, D. P. (2002). Towards a taxonomic revision of the genus Echinococcus. Trends in Parasitology 18, 452–457.CrossRefGoogle ScholarPubMed
Vincze, T., Posfai, J. and Roberts, R. J. (2003). NEBcutter: A program to cleave DNA with restriction enzymes. Nucleic Acids Research 31, 3688–3691.CrossRefGoogle ScholarPubMed
von Nickisch-Rosenegk, M., Lucius, R. and Loos-Frank, B. (1999 a). Contributions to the phylogeny of the Cyclophyllidea (Cestoda) inferred from mitochondrial 12S rDNA. Journal of Molecular Evolution 48, 586–596.CrossRefGoogle Scholar
von Nickisch-Rosenegk, M., Silva-Gonzalez, R. and Lucius, R. (1999 b). Modification of universal 12S rDNA primers for specific amplification of contaminated Taenia spp. (Cestoda) gDNA enabling phylogenetic studies. Parasitology Research 85, 819–825.CrossRefGoogle ScholarPubMed
Xiao, N., Qiu, J., Nakao, M., Li, T., Yang, W., Chen, X., Schantz, P. M., Craig, P. S. and Ito, A. (2005). Echinococcus shiquicus n. sp., a taeniid cestode from Tibetan fox and plateau pika in China. International Journal for Parasitology 35, 693–701.CrossRefGoogle Scholar
Yamasaki, H., Allan, J. C., Sato, M. O., Nakao, M., Sako, Y., Nakaya, K., Qiu, D., Mamuti, W., Craig, P. S. and Ito, A. (2004). DNA differential diagnosis of taeniasis and cysticercosis by multiplex PCR. Journal of Clinical Microbiology 42, 548–553.CrossRefGoogle ScholarPubMed