Development of a species-specific real-time PCR assay for the detection of Eucoleus aerophilus – the first molecular survey in dogs in Poland

Abstract Dogs are hosts to numerous helminth species, including the respiratory nematode Eucoleus aerophilus (syn. Capillaria aerophila ), which infects domestic and wild carnivores and has zoonotic potential. E. aerophilus has been reported in dogs from 24 European countries, with an average prevalence of 3.5%. In Poland, however, molecular data confirming the occurrence of E. aerophilus in dogs have been lacking, and previous investigations relied primarily on coproscopic detection and morphological identification of Capillariidae eggs. The aim of this study was to develop a species-specific real-time quantitative PCR (qPCR) assay for the detection of E. aerophilus and to apply the assay to estimate the prevalence of this parasite in dogs from selected regions of Poland. A total of 198 faecal samples were examined. The newly developed qPCR demponstrated high sensitivity and species-specificity for E. aerophilus . The prevalence determined by qPCR was 7.6%., compared with 4.0% by conventional PCR. Sequence analysis identified four E. aerophilus haplotypes: EaPL1, EaPL1A, EaPL6, and EaPL10. Three of them had previously been reported from red foxes in Poland, whereas EaPL1A was detected for the first time in the country. Coproscopic examination revealed Trichuris vulpis eggs as the most frequently detected parasite (10.5%), followed by Capillariidae eggs (7.0%). Eggs of Taeniidae, Toxocara spp., and hookworms were also identified. This study provides the first molecular evidence of E. aerophilus infection in dogs in Poland and confirms its occurrence in the investigated dog population. The findings contribute to a better understanding of the epidemiology and genetic diversity of E. aerophilus and highlight the value of molecular diagnostics for accurate species identification.