Abstract Body

We determined the occurrence and genetic diversity of Blastocystis sp. in wild ungulates in the Iberian Peninsula including nationwide sample collections from Portugal (n = 222) and Spain (n = 1,058). Faecal samples from three wild ungulate species in Portugal (genera Capreolus, n = 38; Cervus, n = 79; and Sus, n = 105) and eight wild ungulate species in Spain (genera Ammotragus, n = 20; Capra, n = 89; Capreolus, n = 93; Cervus, n = 329; Dama, n = 96; Ovis, n = 10; Rupicapra, n = 62; and Sus, n = 359) were retrospectively investigated by PCR. Next generation amplicon sequencing (NGS) data was used to assess presence of Blastocystis and subtype diversity within a sample. Blastocystis sp. was identified in 37.4% (95% CI: 31.0–43.8%) of the Portuguese samples, with minimum and maximum rates detected in roe deer (31.6%) and red deer (40.5%), respectively. In Spain, 8.7% (95% CI: 7.1–10.4%) of the samples were Blastocystis-positive. Minimum and maximum rates were identified in Iberian wild goats (3.4%) and roe deer (12.9%), respectively. Thirteen Blastocystis subtypes (ST2, ST5, ST10, ST13, ST14, ST15, ST21, ST23, ST24, ST25, ST26, ST30, and ST31) were identified by NGS circulating in the wild ungulate populations in the Iberian Peninsula. The presence of zoonotic Blastocystis ST2 and ST5 in wild ungulates suggests that they could be reservoirs for human infection with this parasite. Blastocystis mixed subtype infections were frequent (42.3%, 74/175), with some animals harbouring up to 11 subtypes. Marked differences in Blastocystis subtype distribution according to host species were observed between countries. NGS was a powerful tool to discern underrepresented or mixed subtype infections that were undetectable or unreadable by Sanger sequencing.