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Theileriosis, a tick-borne disease caused by apicomplexan parasites of the genus Theileria , poses a major constraint to livestock productivity worldwide. Existing chemotherapeutics are increasingly limited by host toxicity and emerging resistance, underscoring the need for parasite-specific interventions. In this study, we integrated comparative proteomics, subtractive genomics, and structure-based modeling to systematically identify conserved and novel druggable targets across four clinically important Theileria species ( T. parva , T. annulata , T. orientalis Fish Creek, and T. orientalis Goon Nure). Sequence clustering and orthologous analysis yielded 15,439 unique proteins, from which 2,753 were identified as evolutionarily conserved across all species. Host-homology filtering against the Bos taurus proteome excluded 1,688 shared sequences, and integration with essential gene datasets revealed 69 parasite-specific proteins crucial for survival. The top three enzymes, ubiquitin carboxyl-terminal hydrolase (UCH), phosphatidylinositol 3/4-kinase (PI3/PI4K), and DNA polymerase A, were prioritized based on evolutionary conservation, functional essentiality, and predicted druggability. Three-dimensional structural models of these targets were refined and validated for stereochemical accuracy and energetic reliability. Virtual screening of small-molecule libraries identified potent lead candidates (CIDs 163724724, 22409369, and 91149393) exhibiting strong binding affinities within the catalytic sites of the respective proteins. Molecular dynamics simulations using GROMACS over 100-ns further confirmed stable protein-ligand interactions, reflected by minimal RMSD fluctuations (~ 1.3–2.5Å), consistent compactness, and persistent hydrogen bonding. These findings demonstrate that the complexes are energetically stable and maintain their conformations, supporting the suitability of the identified targets and their lead inhibitors. Overall, this integrative approach offers a validated framework for developing anti- Theileria drugs and emphasizes the importance of evolutionarily conserved, host non-homologous proteins as promising initial targets for rational therapeutic development against Theileria infections.

Theileriosis is a tropical and sub-tropical disease that causes economic losses in livestock. Theileriosis in small ruminants, manifests through a range of clinical signs, including fever, mucoid nasal secretions, anemia, jaundice, lacrimation, enlarged superficial lymph nodes, anorexia, and accelerated weight reduction. In this study, the prevalence of spp. was determined in asymptomatic small ruminants of the Sistan region located in the southeast of Iran. The collected samples of apparently healthy sheep (n=48) and goats (n=52), obtained between February 2023 and February 2024, were tested by PCR for theileriosis detection. Two positive PCR products sequenced and assembled sequences deposited in GenBank with PQ227215 and PQ227216 accession numbers. Phylogenetic analysis conducted based on partial 18S rRNA gene amplification. The prevalence of spp. in asymptomatic small ruminants was estimated at 32% in the Sistan region in this study. Risk factors were also investigated. A significant relationship was identified between infection and tick infestation in goats; however, no significant associations were found with other parameters such as species, gender, age, or location. Theileriosis appears to be endemic among small ruminants in Sistan. Expanding research efforts in the area and investigating potential risk factors to detect various circulating species of could aid in managing this disease within the region. Conducting larger-scale studies would be beneficial for disease control efforts.

Background Ticks and tick-borne pathogens (TBPs) represent a significant economic burden to cattle farming in sub-Saharan Africa including Nigeria. However, in the northern part of this country, where the largest livestock population resides, little is known about the contemporary diversity of ticks and TBPs. This area is particularly vulnerable to climate change, undergoing marked transformation of habitat and associated flora and fauna that is also likely to include ticks. This study aimed to document the occurrence of tick species and Apicomplexan TBPs in cattle from north-western Nigeria. Methods In 2017, ticks were collected from cattle in Zamfara and Sokoto States and identified morphologically. Additionally, a subset of ticks was screened molecularly for the detection of apicomplexan DNA. Results A total of 494 adult ticks were collected from 80 cattle in Zamfara and 65 cattle in Sokoto State. Nine tick species were encountered, among which the presence of one, Hyalomma turanicum , had not previously been recorded in Nigeria. Hyalomma rufipes was the most prevalent tick infesting cattle in Zamfara State (76%), while Hyalomma dromedarii was the most prevalent in Sokoto State (44%), confirming the widespread transfer of this species from camels onto livestock and its adaptation to cattle in the region. Of 159 ticks screened, 2 out of 54 (3.7%) from Zamfara State and 29 out of 105 (27.6%) from Sokoto State harboured DNA of Theileria annulata , the agent of tropical theileriosis. Conclusions This study confirms the presence of a broad diversity of tick species in cattle from north-western Nigeria, providing the first locality records for Zamfara State. The occurrence of H. turanicum indicates a distribution of this tick beyond northern Africa. This study provides the first report for T. annulata in Nigerian ticks. Given its enormous burden on livestock farming in north Africa and across Asia, further investigations are needed to better understand its epidemiology, vector transmission and potential clinical significance in cattle from northern Nigeria and neighbouring Sahelian countries. Graphical abstract

Parasites of the genus are intracellular protozoa that infect the leukocytes and erythrocytes of animals, causing theileriosis. The aim of the study was to examine the presence of spp. in adult ticks and their offspring in the Lublin region of eastern Poland. Ticks were collected from vegetation and from wild and domestic animals in Lublin Province, Poland. Engorged females were left in laboratory conditions to lay eggs. Detection of spp. was performed by amplifying a fragment of the 18S rRNA gene. Overall, 1,000 ticks collected from vegetation, 50 engorged female ticks from animals, and eggs and larvae from 5 females were tested for spp. The parasite was not detected in adult ticks or samples isolated from eggs and larvae. The results did not confirm the prevalence of in ticks in eastern Poland. The distribution of in ticks and animals in the Lublin macroregion requires further monitoring.

Background There had been isolated reports of the presence of novel Theileria annulata genotypes based on the 18S rRNA gene sequence data from India, Pakistan and Saudi Arabia; but, these studies were restricted to limited field samples. Additionally, no comparative study has been conducted on all the isolates of this parasite from different countries whose sequences are available in the nucleotide databases. Therefore, we aimed to study the genetic diversity of T. annulata based on all available nearly complete 18S rRNA gene sequences in the GenBank™. Out of a total of 312 gene sequences of T. annulata available in the NCBI database, only 70 nearly complete sequences (> 1527 bp) were used for multiple sequence alignment. Results The maximum likelihood tree obtained using TN93 + G + I model manifested two major clades. All the valid host-cell transforming Theileria species clustered in one clade. The T. annulata designated sequences occupying this clade clustered together, excluding two isolates (DQ287944 and EU083799), and represented the true T. annulata sequences ( n  = 54). DQ287944 and EU083799 exhibited close association with Theileria lestoquardi . In addition, 14 Indian sequences formed a large monophyletic group with published Theileria orientalis sequences. The broad range of sequence identity (95.8–100%) of T. annulata designated sequences indicated the presence of different Theileria spp. A closer analysis revealed the presence of three Theileria spp., namely, T. annulata , T. orientalis, and two isolates (DQ287944 and EU083799) closely related to T. lestoquardi . The true T. annulata sequences manifested 98.8–100% nucleotide identity within them. EU083799 and 14 misidentified Indian T. annulata sequences exhibited the highest similarity with T. lestoquardi (98.6–98.8%) and T. orientalis (98.0-99.9%) in comparison with the other Theileria spp. of domestic and wild ruminants. Conclusion In the course of analyzing the genetic diversity of T. annulata , we identified the nearly complete 18S rRNA gene sequences of other Theileria spp. that have not only been misidentified as T. annulata in the GenBank™, but are also published as T. annulata . Moreover, a high level of sequence conservation was noticed in the 18S rRNA gene of true T. annulata and T. orientalis sequences.

Piroplasmosis, caused by Babesia spp. and Theileria spp., poses significant constraints for livestock production and upgradation in Bangladesh. Besides examining blood smears, few molecular reports are available from some selected areas in the country. Therefore, the actual scenario of piroplasmosis in Bangladesh is deficient. This study aimed to screen the piroplasms in different livestock species by molecular tools. A total of 276 blood samples were collected from cattle (Bos indicus), gayals (Bos frontalis) and goats (Capra hircus) in five geographies of Bangladesh. After that, screening was conducted through a polymerase chain reaction, and species were confirmed by sequencing. The prevalence of Babesia bigemina, B. bovis, B. naoakii, B. ovis, Theileria annulata and T. orientalis was 49.28%, 0.72%, 1.09%, 32.26%, 6.52% and 46.01%, respectively. The highest prevalence (79/109; 72.48%) of co-infections was observed with B. bigemina and T. orientalis. The phylogenetic analyses revealed that the sequences of B. bigemina (BbigRAP-1a), B. bovis (BboSBP-4), B. naoakii (AMA-1), B. ovis (ssu rRNA) and T. annulata (Tams-1) were included in one clade in the respective phylograms. In contrast, T. orientalis (MPSP) sequences were separated into two clades, corresponding to Types 5 and 7. To our knowledge, this is the first molecular report on piroplasms in gayals and goats in Bangladesh.

Introduction and objective Protozoa of the genus Theileria transmitted by ticks cause a severe disease in animals, mainly ruminants. The aim of the project was to investigate the occurrence of protozoa Theileria spp. in Ixodes ricinus and Dermacentor reticulatus species using molecular biology methods in the Lublin Province. Material and methods The research material was a group of 30 engorged female ticks removed from the host bodies. In laboratory conditions, the females laid eggs out of which some of the larvae emerged. Then, the isolated genetic material from females, eggs and larvae was analyzed by PCR using primers specific for the protozoa of the genus Theileria spp. Results In the study group of 30 female ticks, 15 females of the species Ixodes ricinus and 15 females of the species Dermacentor reticulatus were identified. In total, females laid 6,720 eggs, of which 2,440 larvae hatched (36.31%). Both eggs and larvae were tested in pools of 20. Theileria protozoa were detected neither in the tested DNA material isolated from females nor in egg and larvae isolates. Conclusions In Poland, there are few studies on Theileria spp. The tests for Theileria spp. showed negative results in the genetic material of females, eggs and larvae in both species of ticks. This made it impossible to draw conclusions about the potential role of transovarial transmission as a pathway for the spread of Theileria spp. in the environment. Summing up, the obtained results did not confirm the participation of the common ticks in the transmission of Theileria spp.

The northwestern region of China, known as the Qinghai–Tibet Plateau Area (QTPA), is characterized by unique climate conditions that support the breeding of various highly-adapted livestock species. Tick vectors play a significant role in transmitting Babesia and Theileria species, posing serious risks to animal health as well as the economy of animal husbandry in QTPA. A total of 366 blood samples were collected from Tibetan sheep (n = 51), goats (n = 67), yaks (n = 43), cattle (n = 49), Bactrian camels (n = 50), horses (n = 65), and donkeys (n = 40). These samples were examined using conventional and nested PCR techniques to detect Theileria and Babesia species. The overall infection rates were 0.3% (1/366) for Babesia spp. and 38.2% (140/366) for Theileria spp. Notably, neither Babesia nor Theileria species were detected in donkeys and yaks. The infection rates of Babesia and Theileria species among animals in different prefectures were significantly different (p < 0.05). Furthermore, Babesia bovis, B. bigemina, B. caballi, and B. ovis were not detected in the current study. To our knowledge, this is the first documented detection of Theileria luwenshuni infection in Bactrian camels and goats, as well as T. sinesis in cattle and T. equi in horses on the Qinghai plateau. These novel findings shed light on the distribution of Babesia and Theileria species among livestock species in QTPA.

Hemoparasites, such as spp., spp. and spp., can negatively affect the health of farm animals resulting in significant losses in production. These losses inherently affect the economics of the livestock industry. Since increases in the severity of vector-borne diseases in the southeast Asian region have been reported, investigations of parasitic epidemiology in Thailand will be necessary to improve the existing parasite control strategies for blood parasitic infections. This study aims to investigate incidences of bovine hemoparasites throughout central and northern Thailand by focusing on areas of high-density cattle populations. Blood parasitic infections among cattle were screened and identified by microscopic examination. Anemia status was then determined by evaluation of the packed cell volume (PCV) of each animal. Furthermore, blood parasites were detected and identified by genus and species-specific primers through the polymerase chain reaction method. Amplicons were subjected to DNA sequencing; thereafter, phylogenetic trees were constructed to determine the genetic diversity and relationships of the parasite in each area. A total of 1,066 blood samples were found to be positive for blood parasitic infections as follows: 13 (1.22%), 389 (36.50%), and 364 (34.15%) for , and , respectively. Furthermore, multiple hemoparasitic infections in the cattle were detected. The hematocrit results revealed 161 hemoparasitic infected samples from 965 blood samples, all of which exhibiting indications of anemia with no significant differences. Sequence analysis of the identified isolates in this study revealed that , four separate clades of , and exhibited considerable sequence similarity to homologous sequences from isolates obtained from other countries. Sequence similarity ranged between 98.57-100%, 83.96-100%, and 97.60-100% for , , and , respectively. In this study, the analyzed incidence data of cattle hemoparasitic infection in Thailand has provided valuable and basic information for the adaptation of blood-borne parasitic infections control strategies. Moreover, the data obtained from this study would be useful for future effective parasitic disease prevention and surveillance among cattle.

Equine piroplasmosis is a tick-borne disease caused by Babesia and Theileria species. Despite its presence in Europe, no laboratory testing is required for animal movement, even though some countries remain free of this disease. Differentiating between species and genotypes is crucial to determine the most effective treatment, as dosage, active compounds, and duration vary. However, diagnosis is often challenging due to genetic variability and the limited sensitivity of molecular methods. The aims of this study were to compare the performances of different molecular diagnostic tests to identify the most effective assay for piroplasm DNA detection and to genetically characterize the piroplasms identified in horses in Northern Italy from 2016 to 2022. Among 63 horses tested, 24 (38.1%) were positive in at least one of the tests used. Four horses tested positive for Babesia caballi with identical nucleotide sequences, while 22 horses tested positive for genetically different Theileria species, including Theileria equi, Theileria haneyi-like species, and Theileria sp. Africa. Two horses were coinfected by Babesia caballi and Theileria haneyi-like species. The best diagnostic approach to avoid false negative results was a combination of different assays. Further studies will be necessary to better assess the prevalence and genetic diversity of piroplasms involved in equine piroplasmosis.