Occurrence of Hookworm and the First Molecular and Morphometric Identification of Uncinaria stenocephala in Dogs in Central Europe

Purpose Hookworms are hematophagous parasitic nematodes that occur in the intestinal tract of various mammals, including humans. The objective of this work was to develop a two-step morphology-molecular analysis-based strategy to identify the genus and the species of eggs and larvae of the Ancylostomatidae family in dogs, which were kept in various living conditions in Slovakia. Methods Faecal samples were collected from 270 dogs kept in two different shelters (160 samples) and in a marginalised Roma community (110 samples). Faecal samples were processed using the flotation method. Microscopically positive faecal samples with hookworm eggs were subjected to a coproculture and the hatched larvae were identified morphometrically, prior to molecular testing. The faecal samples with hookworm ' s eggs and individual larvae were identified by a molecular assay based on the amplification of the 18S ribosomal RNA gene fragment. Further, species-specific primer sets were designed for the internal transcribed spacer (ITS 1 region) and the mitochondrial cytochrome c oxidase subunit I (COX1) gene section. Results Hookworm eggs were microscopically detected in 9.6% (26/270) of the total number of faecal samples. The prevalence in the Roma settlement was higher, 14.5% (16/110), than in shelters, 6.3% (10/160). Using PCR and subsequent Sanger sequencing, we identified the canine hookworm species Uncinaria stenocephala in all positive samples. Conclusion Our results have provided new data on the molecular identification of the neglected species U. stenocephala affecting dogs in Slovakia and supplemented the missing information on the prevalence and incidence of hookworms in dogs in Europe.