Taeniid cestodes in Tibetan foxes (Vulpes Ferrilata) detected by copro-PCR: Applications and challenges

Abstract

Tibetan foxes (Vulpes ferrilata) have been confirmed as the main wild definitive hosts in echinococcosis transmission in the eastern Tibetan Plateau. However, little information is available about the epidemiology in wildlife from the perspective of the Taeniidae family, which is essential knowledge in understanding the epidemiology and phylogeography of cestode species in the Tibetan plateau. Therefore, in this study, we used copro-PCR techniques, by amplifying nad1 and cox1 gene fragments, to detect the taeniid species from Tibetan fox feces collected in Shiqu County, (Sichuan Province, China), eastern Tibetan Plateau. Phylogenetic relationships between amplified sequences and existed Taenia species genotypes were evaluated. Then, the maximum prevalence (positive PCR results from at least one primer pair) and the conservative prevalence (positive PCR results from at least two primer pairs) were calculated. Thirty-six Tibetan fox feces were analyzed. Echinococcus multilocularis (conservative prevalence ± 95% CI: 22.2% ± 13.6%; maximum prevalence ± 95% CI: 33.3% ± 15.4%) and E. shiquicus (2.8 ± 5.4%; 8.3 ± 9.0%) was detected. Meanwhile, DNA fragments of T. polyacantha were detected with high similarity to NCBI sequences (cox1, 94.0%) and to the larva sample DNA sequenced in this study (93.4%), and were supported by phylogenetic analysis. Thus, T. polyacantha might infect Tibetan foxes (5.6% ± 7.5%, 11.1% ± 10.3%). Our limited findings in the epidemiology of parasitic Taenia species suggest that sylvatic transmission cycles for a more species-rich Taeniid community must be established between wild canids and small mammals than just for the two Echinococcus species. Besides, discrepancies in different primer pairs in detecting the taeniid species were evaluated. The sensitivity of some widely used universal primer pairs was poor in detecting Taenia species from canid copro-DNA samples. It is still challenging to the development of effective taeniid species-specific molecular markers especially for non-zoonotic species.

Keywords: Echinococcosis; Taenia polyacantha; Taeniidae; Tibetan fox; Tibetan plateau.

Graphical abstract

Fig. 1

Phylogram of Taenia species, which were found in China, especially in Tibetan Plateau, using the maximum likelihood method generated from partial cox1 (a) and nad1 (b) gene haplotypes. Isolates from Shiqu County were marked with ▲. Sequences from the referential T. polyacantha larva sample collected in Xinyuan County, Xinjiang Autonomous Region, China, shared the same haplotypes with published sequences from Finland marked with ☆. To each sequence, its accession number and geographic region were presented following the taeniid species name and haplotype number. Scale bar was represented in substitutions per nucleotide, and bootstrap values (with 1000 replicates) were marked at the nodes of branches.

Fig. 1

Phylogram of Taenia species, which were found in China, especially in Tibetan Plateau, using the maximum likelihood method generated from partial cox1 (a) and nad1 (b) gene haplotypes. Isolates from Shiqu County were marked with ▲. Sequences from the referential T. polyacantha larva sample collected in Xinyuan County, Xinjiang Autonomous Region, China, shared the same haplotypes with published sequences from Finland marked with ☆. To each sequence, its accession number and geographic region were presented following the taeniid species name and haplotype number. Scale bar was represented in substitutions per nucleotide, and bootstrap values (with 1000 replicates) were marked at the nodes of branches.