Microchip capillary electrophoresis-based genetic comparison of closely related cyathostomin nematode parasites of horses using randomly amplified polymorphic DNA polymerase chain reaction.

The microchip-based capillary electrophoresis technology represents a valuable recent development for the analysis of complex DNA banding patterns. We have used this technology for the differentiation of the closely related cyathostomin species Cylicocyclus elongatus and C. insigne from the horse. We found that the Agilent 2100 bioanalyser in combination with the DNA 7500 Lab Chip were suited to perform a phylogenetic DNA fingerprinting analysis of the parasite species studied. The analysis of the electrophoretic data was optimised and it was possible to resolve a phylogenetic tree where all 12 individual worms of the two Cylicocyclus species studied were assigned to their species as determined by microscopic identification based on morphological traits. Thus, our data indicated that the procedure described here provides an additional powerful tool that can be employed for species delineation of closely related strains or species, such as the two taxa of Cylicocyclus investigated in the present study. Furthermore, by determining the second internal transcribed spacer region of three and nine individual worms for C. elongatus and C. insigne, respectively, low intraspecific variations of only up to 0.3% were demonstrated.