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Acta Parasitologica

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Volume 62, Issue 3 (Sep 2017)

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Differentiation of Toxocara canis and Toxocara cati based on PCR-RFLP analyses of rDNA-ITS and mitochondrial cox1 and nad1 regions

Fattaneh Mikaeili
  • Department of Medical Parasitology and Mycology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
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/ Alexander Mathis
  • Institute of Parasitology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland
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/ Peter Deplazes
  • Institute of Parasitology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland
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/ Hossein Mirhendi
  • Department of Medical Parasitology and Mycology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran
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/ Afshin Barazesh
  • Department of Medical Parasitology and Mycology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
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/ Sepideh Ebrahimi
  • Department of Medical Parasitology and Mycology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
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/ Eshrat Beigom Kia
  • Corresponding author
  • Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran
  • Email
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Published Online: 2017-07-05 | DOI: https://doi.org/10.1515/ap-2017-0066

Abstract

The definitive genetic identification of Toxocara species is currently based on PCR/sequencing. The objectives of the present study were to design and conduct an in silico polymerase chain reaction-restriction fragment length polymorphism method for identification of Toxocara species. In silico analyses using the DNASIS and NEBcutter softwares were performed with rDNA internal transcribed spacers, and mitochondrial cox1 and nad1 sequences obtained in our previous studies along with relevant sequences deposited in GenBank. Consequently, RFLP profiles were designed and all isolates of T. canis and T. cati collected from dogs and cats in different geographical areas of Iran were investigated with the RFLP method using some of the identified suitable enzymes. The findings of in silico analyses predicted that on the cox1 gene only the MboII enzyme is appropriate for PCR-RFLP to reliably distinguish the two species. No suitable enzyme for PCR-RFLP on the nad1 gene was identified that yields the same pattern for all isolates of a species. DNASIS software showed that there are 241 suitable restriction enzymes for the differentiation of T. canis from T. cati based on ITS sequences. RsaI, MvaI and SalI enzymes were selected to evaluate the reliability of the in silico PCR-RFLP. The sizes of restriction fragments obtained by PCR-RFLP of all samples consistently matched the expected RFLP patterns. The ITS sequences are usually conserved and the PCR-RFLP approach targeting the ITS sequence is recommended for the molecular differentiation of Toxocara species and can provide a reliable tool for identification purposes particularly at the larval and egg stages.

Keywords: PCR-RFLP; Toxocara canis; Toxocara cati; internal transcribed spacers (ITS); identification

References

  • Aryaeipour M., Rouhani S., Bandehpour M., Mirahmadi H., Kazemi B., Rokni M.B. 2014. Genotyping and Phylogenetic Analysis of Fasciola spp. Isolated from Sheep and Cattle Using PCR-RFLP in Ardabil Province, Northwestern Iran. Iranian Journal of Public Health, 43, 1364–1371Google Scholar

  • Borecka A. 2004. Differentiation of Toxocara spp. eggs isolated from the soil by the PCR-linked RFLP method. Helminthologia, 41, 185–187Google Scholar

  • Borecka A., Gawor J. 2008. Modification of gDNA extraction from soil for PCR designed for the routine examination of soil samples contaminated with Toxocara spp. eggs. Journal of Helminthology, 82, 119–122. CrossrefGoogle Scholar

  • Chen J., Zhou D.H., Nisbet A.J., Xu M.J., Huang S.Y., Li M.W., Wang C.R., Zhu X.Q. 2012. Advances in molecular identification, taxonomy, genetic variation and diagnosis of Toxocara spp. Infection, Genetics and Evolution, 12, 1344–1348. CrossrefGoogle Scholar

  • Deplazes P., van Knapen F., Schweiger A., Overgaauw P.A. 2011. Role of pet dogs and cats in the transmission of helminthic zoonoses in Europe, with a focus on echinococcosis and toxocarosis. Veterinary Parasitology, 182, 41–53. CrossrefWeb of ScienceGoogle Scholar

  • Despommier D. 2003. Toxocariasis: clinical aspects, epidemiology, medical ecology, and molecular aspects. Clinical Microbiology Reviews, 16, 265–272. CrossrefGoogle Scholar

  • El-Shazly A.M., Mohammed R. M., El-Beshbishi S. N., Azab M. S., El-Ghareeb A. S., Abdeltawab A. H., Zalook T.K. 2009. Soil-parasites particularly Toxocara eggs in Egypt. Journal of the Egyptian Society of Parasitology, 39, 151–162Google Scholar

  • Epe C., Meuwissen M., Stoye M., Schnieder T. 1999. Transmission trials, ITS2-PCR and RAPD-PCR show identity of Toxocara canis isolates from red fox and dog. Veterinary Parasitology, 84, 101–112Google Scholar

  • Fahrion A., Schnyder M., Wichert B., Deplazes P. 2011. Toxocara eggs shed by dogs and cats and their molecular and morphometric species-specific identification: is the finding of T. cati eggs shed by dogs of epidemiological relevance? Veterinary Parasitology, 177, 186–189.CrossrefGoogle Scholar

  • Fogt R. 2006. Molecular techniques applied in species identification of Toxocara. Wiadomosci Parazytologiczne, 52, 31–35Google Scholar

  • Fogt-Wyrwas R., Jarosz W., Mizgajska-Wiktor H. 2007. Utilizing a polymerase chain reaction method for the detection of Toxocara canis and T. cati eggs in soil. Journal of Helminthology, 81, 75–78.CrossrefGoogle Scholar

  • Gasser R.B., Zhu X.Q., Jacobs D.E., Hu M., Chilton N.B. 2006. Molecular genetic characterisation of members of the genus Toxocara – taxonomic, population genetic and epidemiological considerations. In: (Eds: C.V. Holland, H.V. Smith), Toxocara: The Enigmatic Parasite. Wallingford, United Kingdom, 18–31Google Scholar

  • Gawor J., Borecka A., Marczynska M., Dobosz S., Zarnowska-Prymek H. 2014. Risk of human toxocarosis in Poland due to Toxocara infection of dogs and cats. Acta Parasitologica, 60, 99–104. CrossrefGoogle Scholar

  • Jacobs D.E., Zhu X., Gasser R.B., Chilton N.B. 1997. PCR-based methods for identification of potentially zoonotic ascaridoid parasites of the dog fox and cat. Acta Tropica, 68, 191–200Google Scholar

  • Khademvatan S., Abdizadeh R., Tavalla M. 2014. Molecular characterization of Toxocara spp. From soil of public areas in Ahvaz southwestern Iran. Acta Tropica, 135, 50–54. CrossrefGoogle Scholar

  • Khazan H., Khazaei M., Tabaee S.S., Mehrabi A. 2012. Prevalence of Toxocara spp. eggs in Public Parks in Tehran City, Iran. Iranian Journal of Parasitology, 7, 38–42.Google Scholar

  • Lee C.C., Cheng N.A.B.Y., Bohari Y. 1993. Toxocara canis from domestic cats in Kuala Lumpur. Tropical Biomedicine, 10, 79–80Google Scholar

  • Li M.W., Zhu X.Q., Gasser R.B., Lin R.Q., Sani R.A., Lun Z.R., Jacobs D.E. 2006. The occurrence of Toxocara malaysiensis in cats in China, confirmed by sequence based analyses of ribosomal DNA. Parasitology Research, 99, 554–557. CrossrefGoogle Scholar

  • Mahami-Oskouei M., Dalimi A., Forouzandeh-Moghadam M., Rokni M.B. 2011. Molecular Identification and Differentiation of Fasciola Isolates Using PCR- RFLP Method Based on Internal Transcribed Spacer (ITS1, 5.8S rDNA, ITS2). Iranian Journal of Parasitology, 6, 35–42Google Scholar

  • Mikaeili F., Mirhendi H., Hosseini M., Asgari Q., Kia E.B. 2013. Toxocara nematodes in stray cats from Shiraz, southern Iran: intensity of infection and molecular identification of the isolates. Iranian Journal of Parasitology, 8, 593–600Google Scholar

  • Mikaeili F., Mirhendi H., Mohebali M., Hosseini M., Sharbatkhori M., Zarei Z., Kia E.B. 2015. Sequence variation in mitochondrial cox1 and nad1 genes of ascaridoid nematodes in cats and dogs from Iran. Journal of Helminthology, 89, 496–501. CrossrefGoogle Scholar

  • Nichols R.L. 1956. The etiology of visceral larva migrans. I. Diagnostic morphology of infective second-stage Toxocara larvae. Journal of Parasitology, 42, 349–362CrossrefGoogle Scholar

  • Nuchprayoon S., Junpee A., Nithiuthai S., Chungpivat S., Suvannadabba S., Poovorawan Y. 2006. Detection of filarial parasites in domestic cats by PCR-RFLP of ITS1. Veterinary Parasitology, 140, 366–372. CrossrefGoogle Scholar

  • Pawar R.M., Lakshmikantan U., Hasan S., Poornachandar A., Shivaji S. 2012. Detection and molecular characterization of ascarid nematode infection (Toxascaris leonina and Toxocara cati) in captive Asiatic lions (Panthera leo persica). Acta Parasitologica, 57, 67–73. CrossrefWeb of ScienceGoogle Scholar

  • Shafiei R., Sarkari B., Moshfe A. 2013. A Consistent PCR-RFLP Assay Based on ITS-2 Ribosomal DNA for Differentiation of Fasciola Species. Iranian Journal of Basic Medical Sciences, 16, 1266–1269Google Scholar

  • Sheng Z.H., Chang Q.C., Tian S.Q., Lou Y., Zheng X., Zhao Q., Wang C.R. 2012. Characterization of Toxascaris leonina and Tococara canis from cougar (Panthera leo) and common wolf (Canis lupus) by nuclear ribosomal DNA sequences of internal transcribed spacers. African Journal of Microbiology Research, 14, 3545–3549. CrossrefGoogle Scholar

  • Soulsby E.J.L. (Ed.) 1982. Helminths, Arthropods and Protozoa of Domesticated Animals. Bailliere Tindall London, UKGoogle Scholar

  • Spotin A., Gholami S., Nasab A.N., Fallah E., Oskouei M.M., Semnani V., et al. 2015. Designing and conducting in silico analysis for identifying of Echinococcus spp. with discrimination of novel haplotypes: an approach to better understanding of parasite taxonomic. Parasitology Research, 114, 1503–9. CrossrefGoogle Scholar

  • Tavares R.G., Staggemeier R., Borges A.L.P., Rodrigues M.T., Castelan L.A., Vasconcelos J., et al. 2011. Molecular techniques for the study and diagnosis of parasite infection. The Journal of Venomous Animals and Toxins including Tropical Diseases, 17, 239–248CrossrefGoogle Scholar

  • Yamaguti S. (Ed.) 1961. Systema Helminthum: Nematodes of vertebrates. Inter Science publisher Inc., New York, USAGoogle Scholar

  • Zhu X.Q., Gasser R.B. 1998. Single-strand conformation polymorphism (SSCP)-based mutation scanning approaches to fingerprint sequence variation in ribosomal DNA of ascaridoid nematodes. Electrophoresis, 19, 1366–1373. CrossrefGoogle Scholar

  • Zhu X.Q., Jacobs D.E., Chilton N.B., Sani R.A., Cheng N.A., Gasser R.B. 1998. Molecular characterization of a Toxocara variant from cats in Kuala Lumpur, Malaysia. Parasitology, 117, 155–164Google Scholar

  • Zhu X.Q., Gasser R.B., Chilton N.B., Jacobs D.E. 2001. Molecular approaches for studying ascaridoid nematodes with zoonotic potential, with an emphasis on Toxocara species. Journal of Helminthology, 75, 101–108Google Scholar

  • Zibaei M., Abdollahpour F., Birjandi M., Firoozeh F. 2010. Soil contamination with Toxocara spp. eggs in the public parks from three areas of Khorram Abad, Iran. Nepal Medical College journal, 12, 63–65Google Scholar

About the article

Received: 2016-07-26

Revised: 2017-03-02

Accepted: 2017-04-07

Published Online: 2017-07-05

Published in Print: 2017-09-26


Citation Information: Acta Parasitologica, ISSN (Online) 1896-1851, ISSN (Print) 1230-2821, DOI: https://doi.org/10.1515/ap-2017-0066.

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© 2017 W. Stefański Institute of Parasitology, PAS. Copyright Clearance Center

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