New insights into sequence variation in the IGS region of 21 cyathostomin species and the implication for molecular identification.
Cyathostomins comprise a group of 50 species of parasitic nematodes that infect equids. Ribosomal DNA sequences, in particular the intergenic spacer (IGS) region, have been utilized via several methodologies to identify pre-parasitic stages of the commonest species that affect horses. These methods rely on the availability of accurate sequence information for each species, as well as detailed knowledge of the levels of intra- and inter-specific variation. Here, the IGS DNA region was amplified and sequenced from 10 cyathostomin species for which sequence was not previously available. Also, additional IGS DNA sequences were generated from individual worms of 8 species already studied. Comparative analysis of these sequences revealed a greater range of intra-specific variation than previously reported (up to 23%); whilst the level of inter-specific variation (3-62%) was similar to that identified in earlier studies. The reverse line blot (RLB) method has been used to exploit the cyathostomin IGS DNA region for species identification. Here, we report validation of novel and existing DNA probes for identification of cyathostomins using this method and highlight their application in differentiating life-cycle stages such as third-stage larvae that cannot be identified to species by morphological means.
Veterinary Parasitology, Department of Infection Biology, Institute of Infection and Global Health, University of Liverpool, Liverpool, UK. K.Cwiklinski@liverpool.ac.uk
K
K
Cwiklinski
F N J
FN
Kooyman
D C K
DC
Van Doorn
J B
JB
Matthews
J E
JE
Hodgkinson
D000818:Animals; D001483:Base Sequence; D015342:DNA Probes / Q000737:chemistry / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics*; D005260:Female; D014644:Genetic Variation; D006736:Horses / Q000469:parasitology*; D008018:Life Cycle Stages; D008297:Male; D008969:Molecular Sequence Data; D058889:Molecular Typing; D009348:Nematoda / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013045:Species Specificity
D015342:DNA Probes; D021903:DNA, Ribosomal Spacer