Isolates of the rumen fluke Calicophoron daubneyi (Digenea: Paramphistomidae) from various hosts and three locations in southern Italy were characterized genetically. The second internal transcribed spacer (ITS-2) of ribosomal DNA (rDNA) plus flanking 5.8S and 28S sequence (ITS-2+) was amplified from individual rumen flukes by PCR. PCR-linked restriction fragment length polymorphism (PCR-RFLP) analysis was performed using four different restriction endonucleases, and PCR products were sequenced. The PCR analyses from all the C daubneyi specimens produced identical fragments, and the PCR-RFLP analyses did not show, with respect to any of the four restriction endonucleases, any differences between the C. daubneyi specimens. The sequence analyses of the ITS-2+ from each of the C daubneyi specimens showed them all to be 428 bp, and composed of the entire ITS-2 sequence (282 bp) plus the two partial flanking conserved sequences, 5.8S (99 bp) and 28S (47 bp). No intra-specific variation was observed in the nucleotide composition of the 1TS-2+ (homology = 100%). There was, however, an observable interspecific variation between the ITS-2+ of C daubneyi and the ITS-2+ of both Calicophoron calicophorum (homology = 97.2%) and Calicophoron microbothrioides (homology = 97.4%), both previously deposited in the GenBank (TM). The finding of the present study shows that, as has already demonstrated for other parasitic helminths, ITS-2 can serve as an effective genetic marker for the molecular identification of pararaphistornes, and as a useful tool for developing molecular epidemiological techniques for the study of C daubneyi transmission patterns and prevalence in definitive and intermediate hosts. (c) 2005 Elsevier B.V. All rights reserved.

Characterization of the second internal transcribed spacer of ribosomal DNA of Calicophoron daubneyi from various hosts and locations in southern Italy

Musella V;
2005-01-01

Abstract

Isolates of the rumen fluke Calicophoron daubneyi (Digenea: Paramphistomidae) from various hosts and three locations in southern Italy were characterized genetically. The second internal transcribed spacer (ITS-2) of ribosomal DNA (rDNA) plus flanking 5.8S and 28S sequence (ITS-2+) was amplified from individual rumen flukes by PCR. PCR-linked restriction fragment length polymorphism (PCR-RFLP) analysis was performed using four different restriction endonucleases, and PCR products were sequenced. The PCR analyses from all the C daubneyi specimens produced identical fragments, and the PCR-RFLP analyses did not show, with respect to any of the four restriction endonucleases, any differences between the C. daubneyi specimens. The sequence analyses of the ITS-2+ from each of the C daubneyi specimens showed them all to be 428 bp, and composed of the entire ITS-2 sequence (282 bp) plus the two partial flanking conserved sequences, 5.8S (99 bp) and 28S (47 bp). No intra-specific variation was observed in the nucleotide composition of the 1TS-2+ (homology = 100%). There was, however, an observable interspecific variation between the ITS-2+ of C daubneyi and the ITS-2+ of both Calicophoron calicophorum (homology = 97.2%) and Calicophoron microbothrioides (homology = 97.4%), both previously deposited in the GenBank (TM). The finding of the present study shows that, as has already demonstrated for other parasitic helminths, ITS-2 can serve as an effective genetic marker for the molecular identification of pararaphistornes, and as a useful tool for developing molecular epidemiological techniques for the study of C daubneyi transmission patterns and prevalence in definitive and intermediate hosts. (c) 2005 Elsevier B.V. All rights reserved.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/20.500.12317/4416
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