Molecular and serological detection of Theileria equi and Babesia caballi infection in horses and ixodid ticks in Iran.

Equine piroplasmosis is a hemoprotozoan tick-borne disease with worldwide distribution that is caused by Theileria equi and Babesia caballi. However, the geographical distribution of equine piroplasmosis in Iran is unknown. The aim of the current study was to determine the causative agents and vector ticks of equine piroplasmosis in horses in the North Khorasan Province. In the year 2011, 100 horses were randomly selected from 14 villages. Blood samples and ixodid ticks were collected and examined using microscopical, molecular, and serological methods. Theileria equi infection was microscopically detected in 5 (5%) of the blood smears with low parasitemia, while serum samples were tested by the indirect immunofluorescent antibody test (IFAT). Antibodies against T. equi, B. caballi, and a mixed infection were detected in 48 (48%), 2 (2%), and 3 (3%) of the serum samples, respectively. A multiplex PCR was used to detect T. equi and B. caballi DNA in blood samples. No B. caballi infections could be found, but Theileria equi DNA was detected in 45 (45%) of the blood samples, and a BLAST analysis of the sequenced samples indicated a 99% similarity with T. equi 18S rRNA gene sequences in GenBank. Both molecular and serological results did not identify any significant association between T. equi infection and risk factors. A comparision of the results of 3 diagnostic methods demonstrated a poor agreement between microscopical examination with IFAT and PCR and a moderate agreement between IFAT and PCR. Thirty-seven adult ticks (20 females and 17 males) were collected from 15 horses. The most common tick was Hyalomma marginatum marginatum (n=19), followed by Hyalomma anatolicum excavatum (n=10), Rhipicephalus bursa (n=4), Hyalomma marginatum turanicum (n=3), and Hyalomma anatolicum anatolicum (n=1). The salivary glands and ovaries were also examined using PCR. The genomic DNA samples of the salivary glands of 3 ticks, H. a. excavatum (n=2) and R. bursa (n=1), had a positive reaction for T. equi, but no tick contained B. caballi DNA. Thus, our results indicate that T. equi occurs more frequently than B. caballi in the investigated geographical region.