Molecular Characterization of Piroplasms and Identification of Tick Vectors in Central Spain

Introduction and objectives
Piroplasmosis is an endemic tick-borne disease in Spain caused by protozoa of the genera Babesia and Theileria, transmitted by hard ticks (Ixodidae). These hemoprotozoa can induce severe diseases in both animals and humans. Since not all tick species or individuals are infected or competent vectors, identifying the tick fauna present in each area and the piroplasms they harbor is crucial for improving surveillance, prevention, and control strategies.

Methodology
Between 2023 and 2024, the Environmental Risk Surveillance Unit of the Community of Madrid collected 302 ticks, grouped into 148 pools, from 21 host species in central Spain. Hosts included ungulates, wild carnivores, small mammals, birds, and domestic animals. Ticks were morphologically identified to species level. DNA extraction was followed by nested PCR for Babesia and Theileria detection according to Jefferies et al. (2007).

Results and discussion

Wild boar (31.8%), rabbit (19.6%), fox (10.8%), deer (8.8%), and raccoon (6.8%) were the most frequent hosts. The spatial distribution of samples reflects an extensive surveillance effort encompassing both wildlife management areas and urban–periurban environments. The predominant tick species were Hyalomma lusitanicum (32.4%) and Rhipicephalus pusillus (29.7%). Five piroplasm species were identified: Babesia pecorum, Babesia ovis, Babesia sp. (tavsan2/occultans), Theileria ovis, and Theileria capreoli.

The overall infection rate was 6.08% (9/148). Positive samples included B. pecorum (2.7%) in wild boar (Dermacentor marginatus), goat (Rhipicephalus sanguineus), and rabbit (R. pusillus); B. ovis (0,68%) in wild boar (H. lusitanicum); Babesia sp. tavsan2/occultans (0.68%) in Iberian Ibex (Rhipicephalus bursa); T. ovis (1.35%) in rabbit (R. pusillus) and cat (R. sanguineus); and T. capreoli (0.68%) in mouflon (H. lusitanicum). These findings suggest multispecies circulation and possible cross-transmission between wildlife and domestic environments.

Conclusions
The ecological and geographic spread of piroplasms in local tick populations underscores the need for continued surveillance in wildlife and domestic animals in central Spain.