Haemaphysalis longicornis ticks are unable to transstadially transmit Theileria haneyi to horses

Authors: Poh, Karen; Oyen, Kennan; ONZERE, CYNTHIA - WASHINGTON STATE UNIVERSITY; KAPPMEYER, LOWELL; Bastos, Reginaldo

Submitted to: Frontiers in Veterinary Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/17/2025
Publication Date: 4/2/2025
Citation: Poh, K.C., Oyen, K.J., Onzere, C., Kappmeyer, L.S., Bastos, R.G. 2025. Haemaphysalis longicornis ticks are unable to transstadially transmit Theileria haneyi to horses. Frontiers in Veterinary Science. 12-2025. https://doi.org/10.3389/fvets.2025.1572944.
DOI: https://doi.org/10.3389/fvets.2025.1572944

Interpretive Summary: With the emergence of the Asian long horned tick (Haemaphysalis longicornis) and its rapid spread throughout the U.S., it is important to investigate the vector competence of exotic tick species for pathogens that affect both humans and animals. Theileria haneyi is a tick-borne pathogen that causes mild signs of equine piroplasmosis, a disease that is known to affect the health of horses and results in animal movement restrictions globally. The goal of this study was to determine whether H. longicornis can acquire and transstadially transmit Theileria haneyi to horses. Larval and nymphal H. longicornis ticks were added to a T. haneyi-infected horse for the acquisition phase and allowed to feed to repletion. Fully fed ticks were then moved to an incubator to allow the ticks to molt to nymphal and adult ticks, respectively. The nymphal and adult ticks were then added to two horses to evaluate transmission. Furthermore, red blood cells were collected from the acquisition horse and then inoculated in a third transmission horse. Overall, while the larval and nymphal ticks fed on the acquisition horse during a period of detectable parasitemia, the subsequent nymphal and adult ticks were ultimately unable to transstadially transmit T. haneyi to naive horses. The transmission animals remained healthy and were PCR negative for T. haneyi during the observation period. The transmission horse that was inoculated with RBCs was PCR positive for T. haneyi after inoculation, demonstrating that there were infectious parasites in the blood of the acquisition horse. In conclusion, here we demonstrate that H. longicornis ticks are unable to transstadially transmit T. haneyi to horses.

Technical Abstract: Introduction: The recent discovery of Theileria haneyi, a tick-borne hemoparasite that causes mild clinical signs of equine piroplasmosis, has added complexity to this disease that affects equids globally. Equine piroplasmosis is considered a reportable disease by the World Organization for Animal Health (former Office International des Epizooties - OIE) and poses severe restrictions on animal movement worldwide. The knowledge gap regarding potential competent tick vectors for transmitting T. haneyi, combined with the recent emergence of Haemophysalis longicornis in the US, prompted us to conduct this controlled experiment to investigate the ability of H. longicornis to transstadially transmit T. haneyi to horses. Material and Methods: A chronically T. haneyi-infected horse was splenectomized and used for parasite tick acquisition. H. longicornis larvae (0.5g) and nymphs (n=500) were applied on the infected horse for parasite acquisition. During tick feeding, parasitemia in the acquisition horse was monitored by nested PCR (nPCR) and blood smear. Acquisition ticks fed for repletion were transferred to an incubator for molting. Simultaneously, red blood cells (RBC) from the acquisition horse were collected, suspended in 1-volume of 20% PVP-40, and frozen until use for infection. Freshly molted nymphs (n=282) and adults (n=212), offsprings of the acquisition larvae and nymphs, respectively, were placed on two individual naïve spleen-intact horses to evaluate transstadial parasite transmission. One additional naïve horse was inoculated with 1 ml of RBC from the acquisition horse. After tick infestation or RBC inoculation, the transmission horses were monitored for 30 days by nPCR and for the presence of clinical signs of T. haneyi infection. The presence of parasite DNA in the horse blood and tick samples was examined by nPCR while the presence of amplifiable DNA in the samples was conducted by nPCR for the H. longicornis GAPDH and horse ß-actin genes. Results and Discussion: After splenectomy, the acquisition horse developed canonical signs of acute T. haneyi infection, including the presence of parasites in peripheral blood, drop in PCV, and decrease of the total number of RBC. These data reinforce previously published results, demonstrating that the spleen is a critical immunological organ involved in the control of equine Theileria parasites during chronic infection. The percentage of parasitized RBC in the acquisition horse varied between 2.2%-8.1% during tick feeding. Out of a subset of 10 engorged larvae that fed on the acquisition horse, all ticks were nPCR positive for T. haneyi. However, only 4 out of 10 engorged nymphs that fed on the acquisition horse were T. haneyi PCR positive. After evaluating transstadial parasite transmission, there was no evidence for the presence of parasite DNA in the transmission ticks and horse blood nor for T. haneyi clinical signs of infection in the transmission horses. In contrast, the horse inoculated with RBC from the acquisition horse tested nPCR positive for T. haneyi 15 days after inoculation. After that, parasites in blood smears were observed, and the animal developed canonical clinical signs of acute infection, demonstrating the presence of viable, infectious parasites in the blood of the acquisition horse. Conclusion: By performing this controlled-experiment study, we demonstrate that H. longicornis ticks are unable to transstadially transmit T. haneyi to horses.