Very low intraspecific sequence variation in selected nuclear and mitochondrial Parascaris univalens genes.

Abstract

Equines were over decades considered to be infected by two morphologically virtually indistinguishable ascarid species, Parascaris univalens and Parascaris equorum. Reliable species discrimination is only possible using enzyme isoelectric focussing and karyotyping with P. univalens having one and P. equorum two chromosome pairs. However, presumably the complexity of both methods prevented their routine use in nearly all previous studies about prevalence and drug resistance of Parascaris spp. These have barely been performed on the species level although most studies stated presence of one or the other species. Recently, only P. univalens has been identified by karyotyping and the last published study identifying P. equorum dates back to 1989. In order to improve species-specific detection, molecular markers are required. Here, partial 12S rRNA, cytochrome oxidase I (COI) and complete internal transcribed spacer (ITS)-1 and-2 sequences were obtained from 24 karyotyped Parascaris specimens from Poland and 6 German specimens (not karyotyped) and used in phylogenetic analyses with orthologous sequences from GenBank. All karyotyped specimens were identified as P. univalens. In the phylogenetic analysis, they formed very homogenous clusters for all target genes and in a multi-locus analysis. Within this cluster, almost all sequences from GenBank were also included, no matter if they had been assigned to P. univalens or P. equorum. However, a small number of P. univalens ITS and COI sequences originating from donkeys from a single farm in China formed a highly supported sister cluster suggesting that they might represent another Parascaris genotype or species. Our data also strongly suggest that nearly all ITS and COI sequences previously deposited in GenBank and assigned to P. equorum actually represent P. univalens. The fact that significantly different sequences can be found in Parascaris spp. suggests that PCR-based species diagnosis will be possible once molecular markers have been identified for P. equorum from karyotyped specimens.