SummaryAnaplasma species are obligate intracellular rickettsial pathogens transmitted by ticks with an impact on human and animal health. Anaplasma ovis infects sheep and goats in many regions of the world, and it can be diagnosed by different methods like Giemsa staining, PCR or competitive ELISA. In this study, a PCR based on the gene coding for major surface protein 4 (MSP-4) was used to examine field samples collected from sheep in different countries. Altogether, 1161 blood samples from Turkey (n = 830), Iraq (n = 195), Sudan (n = 96) and Portugal (n = 40) were examined, of which 31.4%, 66.6% 41.6% and 82.5%, respectively, were positive. This indicates high prevalence of A. ovis in the countries under investigation, and it can be assumed that the situation in other areas of the world might be similar. Thus, A. ovis should be considered as an important constraint of livestock production, and further efforts are needed to better understand the epidemiology and to implement suitable control measures.

Intracellular leukoproliferative Theileria are unique as eukaryotic organisms that transform the immune cells of their ruminant host. Theileria utilize the uncontrolled proliferation for rapid multiplication and distribution into host daughter cells. The parasite distribution into the daughter cells is accompanied by a tight association with the host cell mitotic apparatus. Since the molecular basis for this interaction is largely unknown, we investigated the possible involvement of the immunodominant Theileria annulata surface protein, TaSP, in the attachment of the parasite to host cell microtubule network. Confocal microscopic analyses showed co-localization of the TaSP protein with alpha-tubulin and reciprocal immuno-co-precipitation experiments demonstrated an association of TaSP with alpha-tubulin in vivo. In addition, the partially expressed predicted extracellular domain of TaSP co-localized with the mitotic spindle of dividing cells and was co-immunoprecipitated with alpha-tubulin in transiently transfected Cos-7 cells devoid of other T. annulata expressed proteins. Pull-down studies showed that there is a direct interaction between TaSP and polymerized microtubules. Analysis of the interaction of TaSP and host microtubulin during host cell mitosis indicated that TaSP co-localizes and interacts with the spindle poles, the mitotic spindle apparatus and the mid-body. Moreover, TaSP was demonstrated to be localized to the microtubule organizing center and to physically interact with gamma-tubulin. These data support the notion that the TaSP-microtubule interaction may be playing a potential role in parasite distribution into daughter host cells and give rise to the speculation that TaSP may be involved in regulation of microtubule assembly in the host cell.

The function of the p53 protein as the central effector molecule of the p53 apoptotic pathway was investigated in a reversible model of epigenetic transformation. The infection of bovine leukocytes by the intracellular protozoan parasite Theileria annulata results in parasitedependent transformation and proliferation of the host cells. We found p53 to be largely localized in the host cell cytoplasm and associated with the parasite membrane of isolated schizonts. Curing infected cells of the parasite with the theilericidal drug buparvaquone resulted in a time-dependent translocation of p53 into the host cell nucleus and the upregulation of the proapoptotic Bax and Apaf-1 and the downregulation of the anti-apoptotic Bcl-2 proteins. Although buparvaquone treatment led to apoptosis of the host cell, inhibition of either p53 or Bax significantly reduced buparvaquone-induced apoptosis of the transformed cells. Thus, the p53 apoptotic pathway of host cells is not induced by infection and transformation with Theileria by a mechanism involving cytoplasmic sequestration of p53. The close association of host cell p53 with the parasite membrane implies that the parasite either interacts directly with p53 or mediates cytoplasmic sequestration of p53 by interacting with other host cell proteins regulating p53 localization.

Summary Infections of small ruminants with Anaplasma, Theileria and Babesia species are widely distributed in the old world and are of great economic impact. In Iraq, data on disease occurrence in sheep caused by above‐mentioned infectious agents are scarce. This study provides information on various haemoparasitic agents infecting sheep in the Kurdistan Region, Iraq, using molecular diagnostic tools. Altogether, 195 samples originating from three governorates in the Kurdistan Region, namely Duhok, Erbil and Sulaimaniya, were analysed. The following pathogens were identified: Anaplasma ovis (62.6%), Theileria ovis (14.35%), T. lestoquardi (7.7%), T. uilenbergi (5.6%) and Babesia ovis (1.5%). T. uilenbergi is detected for the first time in Iraq. Coinfection of sheep with different pathogens could be observed in this study, and it was found that 45 of 195 (23%) of the samples contained more than one pathogen. Even triple‐positive samples were identified in 3% of the investigated animals. In conclusion, we confirm the coinfection of sheep with various haemoparasitic pathogen species in the Kurdistan Region of Iraq. Further investigations are needed to reveal the epidemiology of the diseases, the respective tick vectors, and, in the case of coinfection, pathogens′ interaction and possible cross‐protection.

Intracellular leucoproliferative Theileria are unique as eukaryotic organisms that transform the immune cells of their ruminant host. Theileria utilize the uncontrolled proliferation for rapid multiplication and distribution into host daughter cells. The equal distribution of the schizont into the daughter cells is thought to be accomplished by a tight association with the host cell mitotic apparatus. In this study, we describe a highly conserved novel 37 kD Theileria annulata protein (TaSE). TaSE was found to be localized inside the parasite, the parasite membrane and within the host cell cytoplasm. Moreover, it co-localized at distinct points with host cell microtubules, which was especially apparent during mitosis, where co-localization was found with the centromere, the mitotic spindle and the midbody. Association of TaSE with the host cell tubulin network was corroborated by coimmunoprecipitation and transient transfection experiments. This is the first description of a theilerial protein co-localizing and potentially interacting with a host cell protein. The distribution of TaSE during mitosis makes it a protein to consider as playing a potential role for parasite distribution into daughter host cells.