Display of sequence variation in PCR-amplified mitochondrial DNA regions of Echinococcus by single-strand conformation polymorphism

Abstract

Echinococcosis, a disease caused by infection with the larval stage of a tapeworm parasite of the genus Echinococcus, is of major socio-economic importance, and studying genetic variability within and between Echinococcus populations has important implications for disease control and epidemiology. Various DNA approaches have been used to study Echinococcus genetics, but most methods do not allow the accurate display or definition of mutational/allelic variation. To overcome this limitation, we established a mutation scanning approach. Single-strand conformation polymorphism (SSCP) of two different enzymatically amplified mitochondrial (mt) DNA regions was evaluated using seven different genotypes of Echinococcus (defined as G1, G4, G6, G8, O, V and M2). The NADH dehydrogenase 1 gene ( ND1) or the cytochrome c oxidase subunit 1 ( CO1) were amplified by polymerase chain reaction from parasite DNA, denatured and directly subjected to electrophoresis in a non-denaturing gel matrix. Each of the seven genotypes examined could be delineated from one another based on characteristic and reproducible banding patterns. The results demonstrate the usefulness of SSCP for the direct visual display of sequence variation in mtDNA of Echinococcus without the need for DNA sequencing or restriction analyses, and indicate its potential for studying allelic variability in a range of other genes.