The interplay of cytokines in bovine tropical theileriosis: a mini review.

Studying cytokine profiling in Theleria annulata infection enhances our understanding of how the immune response unfolds, the intricate interactions between the host and the parasite, the strategies employed by the parasite to evade the immune system, and potential avenues for developing treatments. The generation of pro-inflammatory cytokines plays a pivotal role in the immune response against T. annulata infection. Elevated concentrations of these cytokines potentially contribute to the manifestation of clinical symptoms associated with the disease, such as fever, anemia, exophthalmia, and weight loss. The production of anti-inflammatory cytokines potentially serves as a regulatory mechanism for the immune response, preventing the development of severe disease. Nevertheless, in animals afflicted by T. annulata infection, there is often a notable decrease in the levels of these cytokines, suggesting that they may not be as effective in mitigating the disease as they are in uninfected animals. This knowledge can be harnessed to develop improved diagnostic methods, treatments, and vaccines for tropical theileriosis. The objective of this current mini review is to achieve the same goal by consolidating the available knowledge of cytokine interactions in Bovine Tropical Theileriosis (BTT).

Expression of IL-10 and TGF-ß1 in horses experimentally infected with T. equi merozoites is associated with antibody production but not modulation of pro-inflammatory responses.

Theileria equi (T. equi) is an apicomplexan parasite that causes severe hemolytic anemia in equids. Presently, there is inadequate knowledge of the immune responses induced by T. equi in equid hosts impeding understanding of the host parasite relationship and development of potent vaccines for control of T. equi infections. The objective of this study was to evaluate the host-parasite dynamics between T. equi merozoites and infected horses by assessing cytokine expression during primary and secondary parasite exposure, and to determine whether the pattern of expression correlated with clinical indicators of disease. Our findings showed that the expression of pro-inflammatory cytokines was very low and inconsistent during both primary and secondary infection. There was also no correlation between the symptoms observed during primary infection and expression of the cytokines. This suggests that the symptoms might have occurred primarily due to hemolysis and likely not the undesirable effects of pro-inflammatory responses. However, IL-10 and TGF-ß1 were highly expressed in both phases of infection, and their expression was linked to antibody production but not moderation of pro-inflammatory cytokine responses.

Systematic Determination of TCR-Antigen and Peptide-MHC Binding Kinetics among Field Variants of a Theileria parva Polymorphic CTL Epitope.

CTLs are known to contribute to immunity toward Theileria parva, the causative agent of East Coast fever. The Tp967-75 CTL epitope from the Muguga strain of T. parva is polymorphic in other parasite strains. Identifying the amino acids important for MHC class I binding, as well as TCR recognition of epitopes, can allow the strategic selection of Ags to induce cellular immunity toward T. parva In this study, we characterized the amino acids important for MHC class I binding and TCR recognition in the Tp967-75 epitope using alanine scanning and a series of variant peptide sequences to probe these interactions. In a peptide-MHC class I binding assay, we found that the amino acids at positions 1, 2, and 3 were critical for binding to its restricting MHC class I molecule BoLA-1*023:01. With IFN-γ ELISPOT and peptide-MHC class I Tet staining assays on two parasite-specific bovine CTL lines, we showed that amino acids at positions 5-8 in the epitope were required for TCR recognition. Only two of eight naturally occurring polymorphic Tp9 epitopes were recognized by both CTLs. Finally, using a TCR avidity assay, we found that a higher TCR avidity was associated with a stronger functional response toward one of two variants recognized by the CTL. These data add to the growing knowledge on the cross-reactivity of epitope-specific CTLs and specificities that may be required in the selection of Ags in the design of a wide-spectrum vaccine for East Coast fever.

CD4 T Cell Responses to Theileria parva in Immune Cattle Recognize a Diverse Set of Parasite Antigens Presented on the Surface of Infected Lymphoblasts.

Parasite-specific CD8 T cell responses play a key role in mediating immunity against Theileria parva in cattle (Bos taurus), and there is evidence that efficient induction of these responses requires CD4 T cell responses. However, information on the antigenic specificity of the CD4 T cell response is lacking. The current study used a high-throughput system for Ag identification using CD4 T cells from immune animals to screen a library of ∼40,000 synthetic peptides representing 499 T. parva gene products. Use of CD4 T cells from 12 immune cattle, representing 12 MHC class II types, identified 26 Ags. Unlike CD8 T cell responses, which are focused on a few dominant Ags, multiple Ags were recognized by CD4 T cell responses of individual animals. The Ags had diverse properties, but included proteins encoded by two multimember gene families: five haloacid dehalogenases and five subtelomere-encoded variable secreted proteins. Most Ags had predicted signal peptides and/or were encoded by abundantly transcribed genes, but neither parameter on their own was reliable for predicting antigenicity. Mapping of the epitopes confirmed presentation by DR or DQ class II alleles and comparison of available T. parva genome sequences demonstrated that they included both conserved and polymorphic epitopes. Immunization of animals with vaccine vectors expressing two of the Ags demonstrated induction of CD4 T cell responses capable of recognizing parasitized cells. The results of this study provide detailed insight into the CD4 T cell responses induced by T. parva and identify Ags suitable for use in vaccine development.

Integral Use of Immunopeptidomics and Immunoinformatics for the Characterization of Antigen Presentation and Rational Identification of BoLA-DR-Presented Peptides and Epitopes.

MHC peptide binding and presentation is the most selective event defining the landscape of T cell epitopes. Consequently, understanding the diversity of MHC alleles in a given population and the parameters that define the set of ligands that can be bound and presented by each of these alleles (the immunopeptidome) has an enormous impact on our capacity to predict and manipulate the potential of protein Ags to elicit functional T cell responses. Liquid chromatography-mass spectrometry analysis of MHC-eluted ligand data has proven to be a powerful technique for identifying such peptidomes, and methods integrating such data for prediction of Ag presentation have reached a high level of accuracy for both MHC class I and class II. In this study, we demonstrate how these techniques and prediction methods can be readily extended to the bovine leukocyte Ag class II DR locus (BoLA-DR). BoLA-DR binding motifs were characterized by eluted ligand data derived from bovine cell lines expressing a range of DRB3 alleles prevalent in Holstein-Friesian populations. The model generated (NetBoLAIIpan, available as a Web server at www.cbs.dtu.dk/services/NetBoLAIIpan) was shown to have unprecedented predictive power to identify known BoLA-DR-restricted CD4 epitopes. In summary, the results demonstrate the power of an integrated approach combining advanced mass spectrometry peptidomics with immunoinformatics for characterization of the BoLA-DR Ag presentation system and provide a prediction tool that can be used to assist in rational evaluation and selection of bovine CD4 T cell epitopes.

Theileria equi claudin like apicomplexan microneme protein contains neutralization-sensitive epitopes and interacts with components of the equine erythrocyte membrane skeleton.

Theileria equi is a widely distributed apicomplexan parasite that causes severe hemolytic anemia in equid species. There is currently no effective vaccine for control of the parasite and understanding the mechanism that T. equi utilizes to invade host cells may be crucial for vaccine development. Unlike most apicomplexan species studied to date, the role of micronemes in T. equi invasion of host cells is unknown. We therefore assessed the role of the T. equi claudin-like apicomplexan microneme protein (CLAMP) in the invasion of equine erythrocytes as a first step towards understanding the role of this organelle in the parasite. Our findings show that CLAMP is expressed in the merozoite and intra-erythrocytic developmental stages of T. equi and in vitro neutralization experiments suggest that the protein is involved in erythrocyte invasion. Proteomic analyses indicate that CLAMP interacts with the equine erythrocyte α-and ß- spectrin chains in the initial stages of T. equi invasion and maintains these interactions while also associating with the anion-exchange protein, tropomyosin 3, band 4.1 and cytoplasmic actin 1 after invasion. Additionally, serological analyses show that T. equi-infected horses mount robust antibody responses against CLAMP indicating that the protein is immunogenic and therefore represents a potential vaccine candidate.

Interaction between transforming Theileria parasites and their host bovine leukocytes.

Theileria are tick-transmitted parasites that cause often fatal leuko-proliferative diseases in cattle called tropical theileriosis (T. annulata) and East Coast fever (T. parva). However, upon treatment with anti-theilerial drug-transformed leukocytes die of apoptosis indicating that Theileria-induced transformation is reversible making infected leukocytes a powerful example of how intracellular parasites interact with their hosts. Theileria-transformed leukocytes disseminate throughout infected cattle causing a cancer-like disease and here, we discuss how cytokines, noncoding RNAs and oncometabolites can contribute to the transformed phenotype and disease pathology.

Synergistic Effect of Two Nanotechnologies Enhances the Protective Capacity of the Theileria parva Sporozoite p67C Antigen in Cattle.

East Coast fever (ECF), caused by Theileria parva, is the most important tick-borne disease of cattle in sub-Saharan Africa. Practical disadvantages associated with the currently used live-parasite vaccine could be overcome by subunit vaccines. An 80-aa polypeptide derived from the C-terminal portion of p67, a sporozoite surface Ag and target of neutralizing Abs, was the focus of the efforts on subunit vaccines against ECF and subjected to several vaccine trials with very promising results. However, the vaccination regimen was far from optimized, involving three inoculations of 450 µg of soluble p67C (s-p67C) Ag formulated in the Seppic adjuvant Montanide ISA 206 VG. Hence, an improved formulation of this polypeptide Ag is needed. In this study, we report on two nanotechnologies that enhance the bovine immune responses to p67C. Individually, HBcAg-p67C (chimeric hepatitis B core Ag virus-like particles displaying p67C) and silica vesicle (SV)-p67C (s-p67C adsorbed to SV-140-C18, octadecyl-modified SVs) adjuvanted with ISA 206 VG primed strong Ab and T cell responses to p67C in cattle, respectively. Coimmunization of cattle (Bos taurus) with HBcAg-p67C and SV-p67C resulted in stimulation of both high Ab titers and CD4 T cell response to p67C, leading to the highest subunit vaccine efficacy we have achieved to date with the p67C immunogen. These results offer the much-needed research depth on the innovative platforms for developing effective novel protein-based bovine vaccines to further the advancement.

Prevalence of tick-transmitted pathogens in cattle reveals that Theileria parva, Babesia bigemina and Anaplasma marginale are endemic in Burundi.

BACKGROUND: Tick-borne diseases (TBDs) constitute a major constraint for livestock development in sub-Saharan Africa, with East Coast fever (ECF) being the most devastating TBD of cattle. However, in Burundi, detailed information is lacking on the current prevalence of TBDs and on the associated economic losses from mortality and morbidity in cattle as well as the costs associated with TBD control and treatment. The aim of this study was, therefore, to assess the prevalence and spatial distribution of tick-borne pathogens (TBPs) in cattle across the major agro-ecological zones (AEZs) in Burundi. METHODS: In a cross-sectional study conducted in ten communes spanning the five main AEZs in Burundi, blood samples were taken from 828 cattle from 305 farms between October and December 2017. Evidence of Theileria parva infection was assessed by antibody level, measured using a polymorphic immunodominant molecule (PIM) antigen-based enzyme-linked immunosorbent assay (ELISA) and by a T. parva-specific p104 gene-based nested PCR. Antibodies against Theileria mutans infection were detected using the 32-kDa antigen-based indirect ELISA, while the 200-kDa antigen and the major surface protein 5 (MSP5)-based indirect ELISA were used to detect antibodies against Babesia bigemina and Anaplasma marginale, respectively. RESULTS: The prevalence of T. parva across the ten communes sampled ranged from 77.5 to 93.1% and from 67.8 to 90.0% based on the ELISA and PCR analysis, respectively. A statistically significant difference in infection was observed between calves and adult cattle; however, T. parva infection levels were not significantly associated with sex and breed. The seroprevalence indicating exposure to T. mutans, B. bigemina and A. marginale ranged from 30 to 92.1%, 33.7 to 90% and 50 to 96.2%, respectively. Mixed infections of TBPs were detected in 82.91% of cattle sampled, with 11 different combinations of pathogen species detected . CONCLUSIONS: The findings indicate that T. parva, A. marginale and B. bigemina infections are endemic in Burundi. Knowledge of the spatial distribution of TBPs will facilitate the design of effective targeted strategies to control these diseases. There is a need for further investigations of the distribution of tick vectors and the population structure of TBPs in order to identify the key epidemiological factors contributing to TBD outbreaks in Burundi.

Sequence diversity of cytotoxic T cell antigens and satellite marker analysis of Theileria parva informs the immunization against East Coast fever in Rwanda.

BACKGROUND: East Coast fever (ECF) caused by Theileria parva is endemic in Rwanda. In this study, the antigenic and genetic diversity of T. parva coupled with immunization and field challenge were undertaken to provide evidence for the introduction of ECF immunization in Rwanda. METHODS: Blood collected from cattle in the field was screened for T. parva using ELISA and PCR targeting the p104 gene. Tp1 and Tp2 gene sequences were generated from field samples and from Gikongoro and Nyakizu isolates. Furthermore, multilocus genotype data was generated using 5 satellite markers and an immunization challenge trial under field conditions using Muguga cocktail vaccine undertaken. RESULTS: Out of 120 samples, 44 and 20 were positive on ELISA and PCR, respectively. Antigenic diversity of the Tp1 and Tp2 gene sequences revealed an abundance of Muguga, Kiambu and Serengeti epitopes in the samples. A further three clusters were observed on both Tp1 and Tp2 phylogenetic trees; two clusters comprising of field samples and vaccine isolates and the third cluster comprising exclusively of Rwanda samples. Both antigens exhibited purifying selection with no positive selection sites. In addition, satellite marker analysis revealed that field samples possessed both shared alleles with Muguga cocktail on all loci and also a higher proportion of unique alleles. The Muguga cocktail (Muguga, Kiambu and Serengeti) genotype compared to other vaccine isolates, was the most represented in the field samples. Further low genetic sub-structuring (FST = 0.037) coupled with linkage disequilibrium between Muguga cocktail and the field samples was observed. Using the above data to guide a field immunization challenge trial comprising 41 immunized and 40 control animals resulted in 85% seroconversion in the immunized animals and an efficacy of vaccination of 81.7%, implying high protection against ECF. CONCLUSIONS: Antigenic and genetic diversity analysis of T. parva facilitated the use of Muguga cocktail vaccine in field conditions. A protection level of 81.7% was achieved, demonstrating the importance of combining molecular tools with field trials to establish the suitability of implementation of immunization campaigns. Based on the information in this study, Muguga cocktail immunization in Rwanda has a potential to produce desirable results.

Pathogenesis of liver lesions in Theileria orientalis-inoculated cattle and severe combined immunodeficiency mice with bovine erythrocyte transfusion.

Theileria orientalis (T. orientalis) is a bovine protozoal disease similar to malaria in humans. Although the common outcome of malaria in humans and T. orientalis infection in cattle is hepatic disorder, the mechanisms of its development remain unknown. In this study, we investigated hepatocyte injury characterized by accumulation of macrophages with ingested erythrocytes in sinusoid and extramedullary hematopoiesis in cattle and mice experimentally infected with T. orientalis (T. orientalis-infected cattle and T. orientalis-infected mice). Vacuolization of hepatic cells was frequently observed in the vicinity of the aggregated macrophages in the liver sinusoids of T. orientalis-infected mice. A significant percentage of the macrophages accumulated in the liver sinusoids of the severely infected cattle and mice (14.6% and 24.2 to 53.2%, respectively) reacted positively with interleukin-1, interleukin-6 and TNF-α antibodies. Increase in the production of these cytokines was confirmed in T. orientalis-infected cattle and mice by real-time RT-PCR. These findings strongly suggest that increased cytokine production by the macrophages that have phagocytosed T. orientalis-infected erythrocytes causes hepatic disorder in T. orientalis-infected animals.

Role of cytokines in the clinical manifestation of exophthalmia in newborn calves with tropical theileriosis.

The present study aimed to evaluate the pathology of the exophthalmia and the host-immune response in naturally Theileria annulata-infected calves. The newborn calves detected positive for theileriosis were grouped into calves with theileriosis and absence of exophthalmia (n = 30), and calves with theileriosis and the presence of exophthalmia (n = 13). Sixteen healthy calves, free from any haemoprotozoal infection, were kept as healthy controls. A significantly (P ≤ .001) higher circulating levels of tumour necrosis factor-α (TNF-α) and interleukin-10 (IL-10) were estimated in diseased calves with and without exophthalmia as compared to healthy controls. Contrarily, significantly (P ≤ .01) lower interferon-γ (IFN-γ) level was estimated in diseased calves. The diseased calves with exophthalmia revealed significantly higher levels of TNF-α (P ≤ .001) and IL-10 (P ≤ .006) as compared to the diseased calves without exophthalmia. The diseased calves were not found to have an elevated intraocular pressure; rather they had significantly (P ≤ .001) lower intraocular pressure compared to the healthy controls. An elevated systemic TNF-α level might be attributed to the exophthalmia in calves with tropical theileriosis. The elevated circulatory IL-10 and reduced IFN-γ levels could be one of the strategies of Theileria annulata to escape the host immunity.

Evaluation of cytokines and sialic acids contents in horses naturally infected with Theileria equi.

This study was undertaken to assess the effects of T. equi infection on serum concentrations of some important cytokines including interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and interleukin-1 beta (IL-1ß), IL-1α, IL-4, IL-6, IL-8, IL-10, IL-12α, IL-12ß, IL-18, as well as total, protein and lipid binding sialic acids (TSA, PBSA and LBSA). Furthermore, any probable relation among the parasitemia, cytokines and sialic acids (SAs) were calculated using Pearson correlation and simple linear regression. Almost 300 draft horses (Kurdish-breed) with age of 3-4 years old from north-west of Iran were examined and an infected group comprised of 28 mares, naturally infected with T. equi, was identified and divided into 3 subgroups according to their parasitemia rates (low <1 %, moderate 1-3 % and high 3-5 %). Twenty healthy horses were considered as a control. Characterization and differentiation of piroplasmosis were conducted using routine hematological procedures and specific PCR assay. The results revealed a significant increase (P < 0.05) in all of the cytokines and SAs in a parasitic burden-dependent fashion. Additionally, a strong and positive relation was detected among the parasitemia, cytokines and SAs. Conclusively, T. equi infection is associated with induction of severe inflammatory processes in horses and SA plays a pivotal role in pathophysiology of the disease as it is tightly correlated with the parasitemia rate.

Changes in the Molecular and Functional Phenotype of Bovine Monocytes during Theileria parva Infection.

Theileria parva is the causative agent of East Coast fever (ECF), a tick-borne disease that kills over a million cattle each year in sub-Saharan Africa. Immune protection against T. parva involves a CD8+ cytotoxic T cell response to parasite-infected cells. However, there is currently a paucity of knowledge regarding the role played by innate immune cells in ECF pathogenesis and T. parva control. Here, we demonstrate an increase in intermediate monocytes (CD14++ CD16+) with a concomitant decrease in the classical (CD14++ CD16-) and nonclassical (CD14+ CD16+) subsets at 12 days postinfection (dpi) during lethal infection but not during nonlethal T. parva infection. Ex vivo analyses of monocytes demonstrated upregulation of interleukin-1 beta (IL-1ß) and tumor necrosis factor alpha (TNF-α) mRNA and increased nitric oxide production during T. parva lethal infection compared to nonlethal infection at 10 dpi. Interestingly, no significant differences in peripheral blood parasite loads were observed between lethally and nonlethally infected animals at 12 dpi. In vitro stimulation with T. parva schizont-infected cells or Escherichia coli lipopolysaccharide (LPS) resulted in significant upregulation of IL-1ß production by monocytes from lethally infected cattle compared to those from nonlethally infected animals. Strikingly, monocytes from lethally infected animals produced significant amounts of IL-10 mRNA after stimulation with T. parva schizont-infected cells. In conclusion, we demonstrate that T. parva infection leads to alterations in the molecular and functional phenotypes of bovine monocytes. Importantly, since these changes primarily occur in lethal infection, they can serve as biomarkers for ECF progression and severity, thereby aiding in the standardization of protection assessment for T. parva candidate vaccines.

A Seroepidemiological Survey of Theileria equi and Babesia caballi in Horses in Mongolia.

Equine piroplasmosis caused by Theileria equi and Babesia caballi is an economically important disease with a worldwide distribution. The objective of the present study was to investigate the seroepidemiology of T. equi and B. caballi in horses reared in various Mongolian provinces. Serum samples prepared from blood collected from horses in 19 Mongolian provinces were screened for antibodies specific to T. equi and B. caballi using enzyme-linked immunosorbent assays based on recombinant forms of T. equi merozoite antigen-2 and the B. caballi 48-kDa merozoite rhoptry protein, respectively. Of 1,282 horses analyzed, 423 (33%) and 182 (14.2%) were sero-positive for T. equi and B. caballi, respectively. Additionally, 518 (40.4%) were positive for at least 1 parasite species, of which 87 (16.8%) were co-infected with both parasites. Both T. equi and B. caballi were detected in all surveyed provinces, and on a per province basis the positive rates ranged from 19.0 to 74.2% and 4.5 to 39.8%, respectively. Theileria equi- and B. caballi-positive rates were comparable between male horses (31.9 and 14.1%, respectively) and female horses (34.5 and 14.3%, respectively). However, the positive rates were higher in the >3-yr-old age group (37.7 and 15.6%, respectively) compared with the 1-3-yr-old age group (19.4 and 10.0%, respectively). These findings confirmed that T. equi and B. caballi infections are widespread among horses all over Mongolia, and that horse age is a risk factor for infection in this country. Our results will be useful for designing appropriate control measures to minimize T. equi and B. caballi infections among Mongolian horses.

Infection dynamics of Theileria equi and Theileria haneyi, a newly discovered apicomplexan of the horse.

Theileria equi infection, exotic to the United States has reemerged through intravenous (iatrogenic) and tick-borne transmission. Surveillance at the US-Mexico border identified a new species, Theileria haneyi, (T. haneyiEP) (EP = Eagle Pass, Texas) which warranted additional investigation due to inability to detect by PCR targeting of T. equi ema-1 and EMA-1-cELISA validated for T. equi. Infection dynamics of T. haneyiEP were evaluated, including ability to superinfect in the presence of T. equi-Texas (T. equiTX), the isolate responsible for the reemergence of T. equi in the U S. Experimental infection with T. equiTX or T. haneyiEP revealed minimal clinical disease however, T. equiTX infection led to significantly greater neutropenia. Comparison of time to antibody detection following inoculation revealed significantly greater time to detectable anti-T. haneyiEP antibody (26.67 days post-inoculation (DPI)) than T. equiTX (11.67 DPI). Regardless of initial infection with either T. equiTX or T. haneyiEP, superinfection was established. Comparative analysis of antibody responses from a splenectomized horse infected with T. haneyiEP to that of a spleen intact horse infected with T. equiFL revealed a different antibody binding profile to T. haneyiEP, T. equiTX and T. equiFL merozoite antigen and limited shared antigen/cross-reactive antibody(s). Affinity purified T. equi EMA-1 and EMA-2 from T. equiFL were shown as targets for horse antibodies against T. haneyi. Data presented here show (1) T. haneyiEP can superinfect in the presence of T. equiTX infection and co-persists for minimally 25 months, (2) intravenous challenge with T. haneyi is subclinical, and (3) limited cross-reactive antibody between T. haneyiEP and T. equi includes reactivity to EMA-1 and EMA-2.

Molecular and Antigenic Properties of Mammalian Cell-Expressed Theileria parva Antigen Tp9.

East Coast Fever (ECF), caused by the tick-borne apicomplexan parasite Theileria parva, is a leading cause of morbidity and mortality in cattle of sub-Saharan Africa. The infection and treatment method (ITM) is currently the only vaccine available to control T. parva. Although ITM elicits levels of protection, its widespread adoption is limited by costs, laborious production process, and antibiotic co-treatment requirement, necessitating the development of a more sustainable vaccine. To this end, efforts have been concentrated in the identification of new T. parva vaccine antigens and in the development of suitable platforms for antigen expression. In this study, we investigated the molecular and antigenic properties of T. parva antigen Tp9 expressed by mammalian cells. Data indicate that Tp9 contains a signal peptide that is weakly functional in mammalian cells. Thus, Tp9 secretion from mammalian cells increased 10-fold after the native signal peptide was replaced with the human tissue plasminogen activator signal peptide (tPA). Sera from all T. parva-immune cattle recognized this recombinant, secreted Tp9. Additionally, PBMC from ITM-immunized cattle produced significant (p < 0.05) amounts of IFNγ following ex vivo exposure to Tp9, but this response varied between cattle of different MHC class I and class II genotypes. In addition, depletion experiments demonstrated that IFNγ to Tp9 was primarily produced by CD4+ T cells. Molecular analysis demonstrated that Tp9 presents a signal peptide that is weakly functional in mammalian cells, suggesting that it remains within lymphocytes during infection. Tp9 secretion from mammalian cells was substantially increased when the tPA secretion signal sequence was substituted for the native secretion signal sequence. Using full-length, recombinant Tp9 secreted from mammalian cells, we demonstrated that T. parva-immune cattle develop both humoral and cellular immune responses to this antigen. Collectively, these results provide rationale for further evaluation of Tp9 as a component of a T. parva subunit vaccine.

A Review on Equine Piroplasmosis: Epidemiology, Vector Ecology, Risk Factors, Host Immunity, Diagnosis and Control.

Equine Piroplasmosis (EP) is a tick-borne disease caused by apicomplexan protozoan parasites, Babesia caballi and Theileria equi. The disease is responsible for serious economic losses to the equine industry. It principally affects donkeys, horses, mules, and zebra but DNA of the parasites has also been detected in dogs and camels raising doubt about their host specificity. The disease is endemic in tropical and temperate regions of the world where the competent tick vectors are prevalent. Infected equids remain carrier for life with T. equi infection, whilst, infection with B. caballi is cleared within a few years. This review focuses on all aspects of the disease from the historical overview, biology of the parasite, epidemiology of the disease (specifically highlighting other non-equine hosts, such as dogs and camels), vector, clinical manifestations, risk factors, immunology, genetic diversity, diagnosis, treatment, and prevention.

The haematological, proinflammatory cytokines and IgG changes during an ovine experimental theileriosis.

Malignant ovine theileriosis is caused by Theileria lestoquardi, which is highly pathogenic in sheep. Theileriosis involves different organs in ruminants. Little is known about the role of proinflammatory cytokines in the pathogenesis of T. lestoquardi infection. The aim of this study was to measure concentration changes of proinflammatory cytokines and immunoglobulin G (IgG) during an ovine experimental theileriosis and correlate it with clinical and haematological parameters. During an experimental study, seven healthy Baluchi sheep (four females and three males) about 6-8 months old were infected with T. lestoquardi by feeding of infected unfed ticks on the sheep's ears. The infected sheep were clinically examined during the study and blood samples were collected on days 0, 2, 5, 7, 10, 12, 14, 17 and 21. The haematological parameters were analysed by an automatic veterinary haematology cell counter and the inflammatory cytokines interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α), interferon-γ (IFN-γ) and IgG were measured by enzyme-linked immunosorbent assay. All infected sheep had temperatures above 40 °C on days 3-4 post infection (PI). The maximum temperature was noted on day 7, and it remained high until day 21. The parasitaemia of T. lestoquardi infection increased from 0.01% (day 7 PI) to 3.3% (day 21 PI). The mean white blood cell (WBC), red blood cell (RBC), lymphocyte, neutrophil and platelet values slightly increased on day 2 PI and decreased by day 17 and day 21 PI. The percentage parasitaemia and fever had a negative correlation with the numbers of WBCs, RBCs, lymphocytes, neutrophils and platelets. The serum concentration of IL-6, TNF-α and IFN-γ cytokines increased and peaked on day 12 and thereafter decreased to levels lower than 0. Out of all tested cytokines, the concentration of IL-6 was significantly higher, as early as day 2 PI. No significant changes were observed for the IgG levels during the course of disease. A significant and strong correlation was observed between IL-6, TNF-α and IFN-γ values and a moderate correlation between IL-6 and the numbers of lymphocytes in the present study. A strong correlation was determined between the percentage parasitaemia and haematological parameters in T. lestoquardi-infected sheep. In addition, preliminary results indicate that the measurement of the serum concentrations of IL-6 in combination with haematological parameters could be considered a good marker to estimate the pathogenicity of T. lestoquardi strain.

Gene gun DNA immunization of cattle induces humoral and CD4 T-cell-mediated immune responses against the Theileria parva polymorphic immunodominant molecule.

Theileria parva kills over one million cattle annually in sub-Saharan Africa. Parasite genetic complexity, cellular response immunodominance, and bovine MHC diversity have precluded traditional vaccine development. One potential solution is gene gun (GG) immunization, which enables simultaneous administration of one or more DNA-encoded antigens. Although promising in murine, porcine, and human vaccination trials, bovine GG immunization studies are limited. We utilized the model T. parva antigen, polymorphic immunodominant molecule (PIM) to test bovine GG immunization. GG immunization using a mammalian codon optimized PIM sequence elicited significant anti-PIM antibody and cell-mediated responses in 7/8 steers, but there was no difference between immunized and control animals following T. parva challenge. The results suggest immunization with PIM, as delivered here, is insufficient to protect cattle from T. parva. Nonetheless, the robust immune responses elicited against this model antigen suggest GG immunization is a promising vaccine platform for T. parva and other bovine pathogens.