Theileria secretes proteins to subvert its host leukocyte.

Theileria parasites are classified in the phylum Apicomplexa that includes several genera of medical and veterinary importance such as Plasmodium, Babesia, Toxoplasma and Cryptosporidium. These protozoans have evolved subtle ways to reshape their intracellular niche for their own benefit and Theileria is no exception. This tick transmitted microorganism is unique among all eukaryotes in that its intracellular schizont stage is able to transform its mammalian host leukocytes into an immortalised highly disseminating cell that phenocopies tumour cells. Here, we describe what is known about secreted Theileria-encoded host cell manipulators.

Comparative proteomic and bioinformatic analysis of Theileria luwenshuni and Theileria uilenbergi.

Theileria is an obligatory intraerythrocytic protozoan parasite that causes economic losses to the cattle, sheep and goats industry. However, very little information is available on the genomes, transcriptomes, and proteomes of the ovine parasites, Theileria luwenshuni and Theileria uilenbergi. Differences in protein expression between these species were investigated to better understand their biology. Parasites were digested with trypsin, and the resulting peptides labeled with isobaric tags for relative and absolute quantification, followed by LC-MS/MS. More than 670 proteins, classified into categories primarily related to cellular process (29.78%), metabolic process (28.80%), localization (5.22%) and biological regulation (5.00%), were identified. Seventy-one proteins were differentially expressed; T. luwenshuni had 39 proteins more highly expressed than in T. uilenbergi, whereas T. uilenbergi had 32 that were more highly expressed. Several proteins related to parasite virulence and invasion (cysteine proteinase, histone deacetylase, pyruvate kinase, small nuclear ribonucleoprotein and orotate phosphoribosyltransferase) were differentially expressed. Real-time quantitative PCR validated protein expression changes at the transcript level. This is the first report on protein expression for the two most economically important Theileria species in China, and our findings may provide novel opportunities for ovine and caprine theileriosis control.

Identification of a piroplasm protein of Theileria orientalis that binds to bovine erythrocyte band 3.

Theileria orientalis infects cattle and causes various disease symptoms, including anaemia and icterus. The erythrocytic stages are responsible for these symptoms but the molecular events involved in these stages have not yet been fully elucidated. In this study, we identified a T. orientalis cDNA that encodes a polypeptide related to identity to the microneme-rhoptry protein of Theileria parva. Analysis of its recombinant product (ToMRP) by indirect fluorescent-antibody test revealed that it is specifically expressed at the early erythrocytic stage after invasion. This expression disappears during the intermediate stages of intra-erythrocytic development. Its expression then reappears at the late stages after the parasite has divided by binary fission into diad or tetrad forms and before these forms are released from the host erythrocyte. In vitro erythrocyte binding assays showed that ToMRP associates with the Triton X-insoluble fraction of erythrocytes membrane but not with intact erythrocytes. Cosedimentation and Western blot analyses revealed that ToMRP binds to band 3, a membrane component of bovine erythrocytes. These observations suggest that ToMRP may be involved in the parasite's egress from and/or invasion into the host erythrocytes by interacting with a protein in the membrane skeleton of the erythrocyte and thereby modifying the structure and function of the cell.

Expression of a 32 kilodalton Theileria sergenti piroplasm surface protein by recombinant baculoviruses.

Previous studies detected a single amino acid substitution (Ala196 to Gly196) between cDNA clones encoding a 32 kDa antigen (p32) of Theileria sergenti (Chitose stock) obtained from a persistently infected calf. In this study, 2 different recombinant baculoviruses (pAc/p32-Ala196 and pAc/p32-Gly196) were constructed for the expression of p32. Molecular masses of the polypeptides produced in Spodoptera frugiperda cells infected with the recombinant baculoviruses were the same as that of authentic p32. pAc/p32-Ala196 produced additional polypeptides, with molecular masses higher than 32 kDa, which resulted from differential N-glycosylation as revealed by endo N-glycosidase treatment. The results indicate that a single amino acid substitution may lead to a conformational change in p32 which affected post-translational modification of recombinant products.

Selection of diversity at putative glycosylation sites in the immunodominant merozoite/piroplasm surface antigen of Theileria parasites.

The immunodominant merozoite/piroplasm surface antigen of Theileria parasites has potential as a diagnostic reagent and as a component of a sub-unit vaccine. This molecule is known to be antigenically diverse, and it is important to determine the nature and extent of this heterogeneity. In the present study nucleotide sequences, representing alleles of the gene (Tams1) encoding this molecule in Theileria annulata were compared to each other and to sequences of homologous genes in Theileria sergenti, Theileria buffeli and Theileria parva. This analysis revealed that a region of the polypeptide which contains putative N-linked glycosylation sites is particularly diverse and, in analogy to retroviral systems, may indicate selection of variable glycosylation sites or amino acid epitopes to evade the bovine immune response. This conclusion was also made from the results of a phylogenetic analysis which compared the variable region of the genes with a second region, which appeared to show no bias for diversity or functional constraint. The results indicated that the variable sequence encoding putative glycosylation sites has diverged, both within and between Theileria species, at a much faster rate than the rest of the molecule. Southern blot analysis of T. annulata populations from within a single geographical region detected six possible variant Tams1 alleles. However, a correlation between restriction-fragment-length polymorphism (RFLP) patterns detected by the Tams1-1 gene probe and geographical location could not be made. In addition, although a high prevalence of one particular RFLP was found, this is unlikely to be the result of a clonal population structure, as we present evidence for significant parasite genotypic variability within a single endemic region.