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Detection of Dicrocoelium dendriticum larval stages in mollusc and ant intermediate hosts by PCR, using mitochondrial and ribosomal internal transcribed spacer (ITS-2) sequences

Published online by Cambridge University Press:  24 August 2011

A. M. MARTÍNEZ-IBEAS ,
M. MARTÍNEZ-VALLADARES ,
C. GONZÁLEZ-LANZA ,
B. MIÑAMBRES  and
M. Y. MANGA-GONZÁLEZ
A. M. MARTÍNEZ-IBEAS
Affiliation:
Departamento de Sanidad Animal, Consejo Superior de Investigaciones Científicas (CSIC), Instituto de Ganadería de Montaña (CSIC-ULE), 24346 Grulleros, León, Spain
M. MARTÍNEZ-VALLADARES
Affiliation:
Departamento de Sanidad Animal, Consejo Superior de Investigaciones Científicas (CSIC), Instituto de Ganadería de Montaña (CSIC-ULE), 24346 Grulleros, León, Spain
C. GONZÁLEZ-LANZA
Affiliation:
Departamento de Sanidad Animal, Consejo Superior de Investigaciones Científicas (CSIC), Instituto de Ganadería de Montaña (CSIC-ULE), 24346 Grulleros, León, Spain
B. MIÑAMBRES
Affiliation:
Departamento de Sanidad Animal, Consejo Superior de Investigaciones Científicas (CSIC), Instituto de Ganadería de Montaña (CSIC-ULE), 24346 Grulleros, León, Spain
M. Y. MANGA-GONZÁLEZ*
Affiliation:
Departamento de Sanidad Animal, Consejo Superior de Investigaciones Científicas (CSIC), Instituto de Ganadería de Montaña (CSIC-ULE), 24346 Grulleros, León, Spain
*
*Corresponding author: Tel: +34 987 317064/ 317156. Fax: +34 987 317161. E-mail: y.manga@eae.csic.es
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Summary

The aim of this study was to develop, perfect and validate the PCR (polymerase chain reaction) technique using mitochondrial (mt) and ribosomal (ITS-2) DNA for the accurate identification of Dicrocoelium dendriticum in molluscs and ants, the first and second intermediate hosts, and their early detection. The first primers that were designed amplified a 169 pb mtDNA fragment of D. dendriticum permitted the detection of a single D. dendriticum metacercaria from the Formica rufibarbis and Formica pratensis abdomen, as well as the detection of the brainworm in the head of the ants collected in tetania. Although these primers did not amplify Dicrocoelium chinensis DNA and permitted detected D. dendriticum in the molluscs, they did not discriminate Brachylaimidae metacercariae found in the same mollusc. The PCR that was designed to amplify a 93 bp fragment of the ITS-2 is D. dendriticum specific as it did not amplify D. chinensis, Brachylaimidae and other trematodes. This technique is very sensitive since it permitted the detection of D. dendriticum in the molluscs from the first day post-infection, the brainworm in the head of the ants and only 1 D. dendriticum metacercaria from the abdomen of the ants. Both techniques are important, mainly the latter.

Keywords

Dicrocoelium dendriticummollusc and ant intermediate hostsPCR diagnosismitochondrial and ITS-2 sequences
Type
Research Article
Information
Parasitology , Volume 138 , Issue 14 , December 2011 , pp. 1916 - 1923
Copyright
Copyright © Cambridge University Press 2011

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