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Molecular characterization and immunodiagnostics of Dicrocoelium dendriticum species isolated from sheep of north-west Himalayan region

Published online by Cambridge University Press:  20 July 2020

J.S. Dar Open the ORCID record for J.S. Dar [Opens in a new window] ,
U. Shabir ,
S.A. Dar  and
B.A. Ganai
J.S. Dar*
Affiliation:
Centre of Research for Development, University of Kashmir, Srinagar, Jammu and Kashmir190 006, India
U. Shabir
Affiliation:
Centre of Research for Development, University of Kashmir, Srinagar, Jammu and Kashmir190 006, India
S.A. Dar
Affiliation:
Centre of Research for Development, University of Kashmir, Srinagar, Jammu and Kashmir190 006, India
B.A. Ganai
Affiliation:
Centre of Research for Development, University of Kashmir, Srinagar, Jammu and Kashmir190 006, India
*
Author for correspondence: J.S. Dar, E-mail: [email protected]
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Abstract

Despite its extensive presence among grazing ruminants, dicrocoeliosis, also known as ‘small liver fluke’ disease, is poorly known and often underestimated by researchers and practitioners in many countries. The accurate identification and prepatent diagnosis of Dicrocoelium dendriticum infection is an essential prerequisite for its prevention and control. In the present study, the morphologically identified specimens isolated from the bile ducts of sheep (Ovis aries) were validated through molecular data. The sequence analysis of the second internal transcribed spacer (ITS-2) of our isolates showed a high degree of similarity with D. dendriticum using the BLAST function of the National Center for Biotechnology Information (NCBI). The phylogenetic analysis of our isolates showed a close relationship with previously described D. dendriticum isolates from different countries. The antigenic profiles of somatic and excretory/secretory (E/S) antigens of D. dendriticum were revealed by sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE) and immunoblotting using sera from sheep naturally infected with D. dendriticum. By SDS–PAGE, 16 distinct bands were revealed from crude somatic fraction. Immunoblotting analysis of these proteins with positive sera exhibited six seroreactive bands ranging from 27 to 130 kDa. Among these, the 84 and 130 kDa bands were quite specific, with high diagnostic specificity and sensitivity. The E/S fraction comprised nine distinct bands, as revealed by SDS–PAGE analysis. Immunoblotting analysis of these proteins with positive sera exhibited five antigenic bands ranging from 27 to 130 kDa. Among these, the 130 kDa band was found to be quite specific, with high diagnostic specificity and sensitivity. The present study concludes that the protein bands of 84 and 130 kDa in somatic fraction and 130 kDa in E/S fraction can be used for the immunodiagnostic purpose for this economically important parasite, which may also encourage further studies regarding their vaccine potential.

Keywords

Dicrocoelium dendriticumITS-2BLASTexcretory/secretory antigen (E/S)sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE)immunoblotting
Type
Research Paper
Information
Journal of Helminthology , Volume 94 , 2020 , e174
Copyright
Copyright © The Author(s), 2020. Published by Cambridge University Press

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