Sporogonic development of Hepatozoon felis in naturally infected Rhipicephalus turanicus and Rhipicephalus rutilus ticks and induction of sporocyst excystation by duodenal fluid

Hepatozoon felis is a protozoan parasite reported in Europe, Africa, Asia and the Americas with infection rates ranging from 4 % to 30 % in domestic cats in Mediterranean countries. Although the parasite was described in 1908, its arthropod vector has not been reported to date. Ticks removed from cats were dissected to prepare hemocoel smears with the aim of identifying the sporogonic life stages of H. felis. Light and live video microscopy were used to visualize these stages and PCR followed by DNA sequencing served to verify the Hepatozoon sp. found and to morphologically and molecularly define the tick species examined. Of 42 adult ticks, 30 were Rhipicephalus turanicus, eight Haemaphysalis adleri, three Rhipicephalus rutilus and one was Rhipicephalus secundus. Sporocysts and oocysts of H. felis verified by PCR were present in eleven R. turanicus and two R. rutilus ticks (31 %). The sporocysts of H. felis were round and each sporocyst harbored 9–11 sporozoites. Developing and mature H. felis oocysts, sporocysts and sporozoites were measured and described morphologically. Experiments to study the excystation of sporocysts and release of infectious sporozoites indicated that combined body temperature of 38.5 ℃ and contact with duodenal fluids are required for activation of sporozoites and excystation as viewed by live imaging. Sporocysts excysted within 250 s of exposure to duodenal fluid and excystation occurred significantly more frequently in the sporocysts that were incubated at 38.5 °C and exposed to duodenal fluid compared to those only incubated at 38.5 °C (P = 0.0001). In conclusion, sporogony of H. felis was described in R. turanicus and R. rutilus filling a gap in knowledge on the life cycle of this parasite whose arthropod vectors were unknown. Induction of H. felis sporocyst excystation required both body temperature and contact with duodenal fluid, simulating conditions met when the parasite reaches the vertebrate host's small intestine.