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PubMed2004

Are rhoptries in Apicomplexan parasites secretory granules or secretory lysosomal granules?

The club-shaped rhoptries in Apicomplexan parasites are one of the most unusual secretory organelles among the eukaryotes, containing unusual lipid and protein cargo that is specialized for intracellular parasitism. Rhoptries have traditionally been viewed strictly as regulated secretory granules. We discuss in this article recent data on the cargo, function and biogenesis of rhoptries in two parasitic model systems, Toxoplasma and Plasmodium. Current findings suggest that rhoptries receive products from both biosynthetic and endocytic pathways and, therefore, they are most analogous to secretory lysosomal granules found in mammalian cells.

Animals↗
PubMed1980

Bovine lymphocyte antigens (BoLA) of bovine lymphocytes and derived lymphoblastoid lines transformed by Theileria parva and Theileria annulata.

The BoLA phenotypes of lymphocytes from seven cattle have been compared with those of 19 lymphoblastoid cell lines derived from them by infection in vitro with either Theileria parva or T. annulata. Two long-established lines were also studied. In all cases except one, the BoLA phenotypes of the lymphoblastoid lines were identical with those of the original animal from whose lymphocytes they were derived. The one exception was a chimeric twin where a minor population appeared to have been transformed by T. parva. The antigens present on this line were present in the parents of the chimera, but not detectable in its own peripheral blood. The implications of these results as they relate to the use of these cell lins in immunizing cattle against East Coast fever and tropical theileriosis are discussed.

Animals↗
PubMed1984

Monoclonal antibody neutralizes the sporozoite stage of different Theileria parva stocks.

Monoclonal antibodies were raised against sporozoites of Theileria parva. One of these antibodies (MAbD1) neutralized the infectivity of sporozoites for lymphocytes in vitro and for cattle in vivo. Neutralization seemed to occur by blocking sporozoite entry into the cell. MAbD1 neutralized sporozoites of four unrelated stocks of T. parva, indicating the presence of a common antigenic determinant which may be important in initiating protective immunity.

Animals↗
PubMed1985

Inhibition of lymphocyte invasion by sporozoites and the transformation of trophozoite infected lymphocytes in vitro by serum from Theileria annulata immune cattle.

Serum from calves immune to Theileria annulata inhibited sporozoite-induced infection and transformation of peripheral blood leucocytes (PBL) into macroschizont infected lymphoblastoid cells in vitro. Invasion by sporozoites was suppressed by some sera, especially that obtained after challenge, but in many instances suppression of infection could be attributed only to retardation of development of PBL infected with intracellular trophozoites into proliferating macroschizont infected cells. Suppression occurred both in cultures of PBL from susceptible calves and in cultures of PBL from T. annulata immune calves. More uninfected than infected blast cells were found both in cultures of PBL from susceptible calves and in cultures of PBL from immune animals. Results suggested that proliferation of uninfected PBL from susceptible calves was initiated by the transformation of sporozoite infected cells, whilst proliferation of uninfected PBL from immune calves also incorporated a component which was a mitotic response of sensitised lymphocytes to sporozoites (i.e. parasite antigen). Thymidine incorporation by cultures was a measure of proliferation of both uninfected and infected blast cells. It could be used, therefore, to monitor parasite development only where proliferation of total blast cells was directly related to proliferation of infected blasts, as in cultures of PBL from susceptible calves. This study suggests that resistance of cattle to tick derived challenge with sporozoites is mediated initially by serum factors acting both on sporozoites and on the development of intracellular trophozoites to schizonts; that the proliferation of uninfected lymphocytes in naive cattle infected with tropical theileriosis is a direct response to the proliferation of Theileria infected cells and that the proliferation of uninfected lymphocytes in Theileria immune cattle is a combined response to the proliferation of Theileria infected cells and parasite antigen.

Animals↗
PubMed1986

Identification of infection-associated antigens in Theileria annulata transformed cells.

The protozoan parasite Theileria annulata causes a severe disease of cattle in tropical countries; one stage in the parasite life cycle involves the transformation of bovine lymphocytes leading to rapid lymphoproliferation. Immunity to this disease is largely cell mediated and directed against the infected lymphocyte. In this paper we report the identification of three classes of infection specific antigen (using monoclonal antibodies) one of which is found on the surface of the lymphocyte. Such antigens can be used in parasite strain typing, in providing an understanding of the molecules involved in immunity and in providing the basis for a vaccine.

Animals↗
PubMed1986

Specific lysis of Theileria annulata-infected lymphoblastoid cells by a monoclonal antibody recognizing an infection-associated antigen.

A monoclonal antibody (4H5) recognizing a Theileria annulata infection-associated antigen was assayed to see if it could either suppress the proliferation of T. annulata-infected lymphoblastoid cells, as monitored by the incorporation of tritiated thymidine by proliferating host cells, or lyse T. annulata-infected lymphoblastoid cells, as assessed by counts of target cell numbers and examination of Giemsa stained smears. These assays showed that binding of the monoclonal antibody, in the presence of complement, both lysed and suppressed the proliferation of the T. annulata Hissar-infected cell line against which this monoclonal antibody was raised. This effect extended both to other (allogeneic) T. annulata Hissar-infected cell lines and to lymphoblastoid cell lines infected with other geographical stocks of T. annulata. An uninfected bovine lymphoid cell line was not affected by the antibody. The results obtained in these in vitro experiments are taken to mean that binding of the monoclonal antibody 4H5 to a T. annulata infection-associated antigen, in the presence of complement, will lyse and suppress specifically the proliferation of T. annulata-infected lymphoblastoid cells. These observations raise the possibility that immunization of cattle with the purified T. annulata infection-associated antigen recognized by this monoclonal antibody may provoke immune responses which are capable of suppressing the proliferation of T. annulata-infected lymphoblastoid cells in vivo and thus provide an effective method of immunoprophylaxis against tropical theileriosis.

Animals↗
PubMed1987

Bovine helper T-cell clones specific for lymphocytes infected with Theileria parva (Muguga).

T-cell clones specific for lymphocytes infected with Theileria parva were derived from animals immunized by infection with T. parva (Muguga). These clones were non-cytolytic and had the BoT4+ BoT8- surface phenotype, BoT4 and BoT8 being the bovine analogues of human CD4 and CD8 molecules. The clones proliferated in response to irradiated autologous lymphoblasts infected with T. parva (Muguga) but not to autologous uninfected lymphoblasts or monocytes. They were parasite strain-specific, in that they did not respond to autologous lymphoblasts infected with another parasite stock, T. parva (Marikebuni). The clones proliferated in the absence of exogenous T-cell growth factor (TCGF) and produced TCGF when stimulated with concanavalin A. Induction of proliferation of the cloned T-cells was genetically restricted, and evidence was obtained which indicated that they were restricted by determinants on class II major histocompatibility complex (MHC) molecules. These findings demonstrate that infections with T. parva stimulate antigen-specific MHC-restricted T-cells with the properties of T-helper cells. The results also provide further evidence for the expression of a parasite strain-specific antigen on the surface of T. parva-infected lymphocytes.

Animals↗
PubMed1987

Cytotoxic T-cells elicited in cattle challenged with Theileria parva (Muguga): evidence for restriction by class I MHC determinants and parasite strain specificity.

The MHC restriction and parasite strain specificity of cytotoxic cells elicited in a group of Theileria parva (Muguga)-immunized cattle following homologous challenge, were investigated. The cytotoxic cells were specific for parasitized target cells and in 9 of the 10 animals examined, they were clearly genetically restricted. Cytotoxicity could be inhibited by monoclonal antibodies (MoAb) to class I MHC molecules but not by MoAb to class II molecules, indicating that a large component of the response was restricted by class I MHC determinants. Low levels of inhibition of cytotoxicity were also obtained with a MoAb to the T-cell subset marker BoT8, suggesting that at least part of the response was mediated by BoT8+ lymphocytes. When cytotoxic cells from individual cattle were assayed on panels of parasitized target cells, there was a close correlation between susceptibility of the target cells to lysis and sharing of BoLA-A locus-encoded specificities with the effectors. This observation, taken together with the knowledge that within several of the sets of BoLA-A-matched targets the relevant BoLA-A specificities were on different MHC haplotypes, indicated that the responses were restricted predominantly by BoLA-A products. In individual cattle there was a striking bias in the restriction of the response to one or other BoLA-A specificity. Among the six specificities represented, responses restricted by w6, w8 and KN18 consistently predominated over responses restricted by w7, w10 and w11. In the three cattle tested for parasite strain specificity, two showed complete specificity and one partial specificity for cells infected with the parasite stock used for immunization, T. parva (Muguga).

Animals↗
PubMed1988

Evaluation of cytotoxic lymphocytes and their parasite strain specificity from African buffalo infected with Theileria parva.

Theileria parva-reactive cytotoxic lymphocytes and their precursors were examined in the blood of African buffalo infected with T. parva and uninfected African buffalo. Peripheral blood mononuclear cells (PBM) from eight of 11 infected buffalo were found to have potent cytotoxic activity after stimulation with autologous parasitized cells for 6 days in vitro, while PBM from uninfected buffalo or PBM from infected buffalo not stimulated in vitro had no cytotoxic activity. The cytotoxic activity was specific for parasitized cells and genetically restricted since there was no killing on uninfected autologous lymphoblasts and a lower percentage of killing on parasitized allogeneic lymphocytes than on targets of autologous parasitized cells. The cytotoxic cells tested for parasite strain specificity were shown to kill autologous cells transformed with different stocks of both cattle-derived (T. parva parva) and buffalo-derived (T. parva lawrencei) parasites.

Animals↗
PubMed1988

Bovine mononuclear cell lines transformed by Theileria parva or Theileria annulata express different subpopulation markers.

Bovine mononuclear cell lines infected with the protozoan parasites Theileria annulata and T. parva have been studied with a panel of monoclonal antibodies reacting with bovine lymphocyte subpopulation markers. All infected lines are MHC class II positive, though the amount of class II antigen expressed varied between lines, and within individual lines there was variation in the proportion of positive cells from 100% with many, to less than 10%. All lines were negative for a macrophage/monocyte marker and for surface IgM. The T. parva lines tested were all positive for BoT4 or BoT8 or both, whereas T. annulata lines were uniformly negative for both of these markers. These results suggest that the two parasites preferentially infect different lymphocyte subpopulations.

Animals↗
PubMed1988

Macrophage-mediated cytostasis and lymphocyte cytotoxicity in cattle immunized with Theileria annulata sporozoites or macroschizont-infected cell lines.

Immunization with either sporozoites or macroschizont-infected cell lines protected calves against challenge with lethal doses of sporozoites of Theileria annulata. Stocks from India, Turkey and Morocco all conferred protective immunity to each other, irrespective of the immunizing regime. Although heterogeneous clinical responses were induced by the two immunizing regimes, both stimulated similar patterns of macrophage cytostasis as expressed as an inhibition of proliferation of macroschizont-infected cell lines. Macrophage cytostasis was detected consistently after immunization and after challenge, arising at the same time as macroschizonts were detectable. Its expression was sustained and inhibited the proliferation of both autologous and allogeneic (BoLA-mismatched) cell lines. In contrast, these two immunizing regimes differed in their ability to stimulate the production of cytotoxic cells. Calves immunized with autologous cell lines or sporozoites developed very transient populations of cytotoxic cells expressing only a low level of specific lysis for autologous infected cells; agglutinating antibodies for immunizing or autologous cell lines were not detected in these calves. Calves immunized with allogeneic cell lines produced cytotoxic cells which were specific only for the immunizing cell lines; these calves also produced antibodies which agglutinated the immunizing cell lines.

Animals↗
PubMed1989

The effect of MHC compatibility between parasite-infected cell line and recipient in immunization against tropical theileriosis.

Lymphoblastoid cell lines, infected and transformed in vitro by a Moroccan stock of Theileria annulata, infected and immunized susceptible taurine cattle, at cell doses of 10(8), 10(6), 10(4) and 10(2), regardless of whether the recipients were BoLA matched or mismatched to the donor cell line. The MHC relationship between the cell line and recipient did affect the severity of the clinical response to cell line immunization which may reflect differences in the specific priming of the immune response. At the highest cell doses the BoLA-mismatched recipients reacted more severely than the BoLA-matched. This study shows that, unlike the closely related parasite T. parva, there is no histocompatibility barrier to immunization using T. annulata-infected cell lines which could be achieved with as few as 10(2) allogeneic infected cells. The role of MHC compatibility between cell line and recipient in the priming of a protective immune response is discussed.

Animals↗
PubMed1989

The development and specificity of cytotoxic cells in cattle immunized with autologous or allogeneic Theileria annulata-infected lymphoblastoid cell lines.

Two groups of animals were immunized with either 10(6) autologous or 10(6) allogeneic Theileria annulata-infected lymphoblastoid cells cultured in vitro. The development and specificity of cytotoxic cells generated in vivo were measured throughout immunization and challenge using a panel of target cells that were either Theileria-infected or uninfected blast cells of known bovine lymphocyte antigen (BoLA) specificities. After inoculation of the cell lines the two groups showed distinct differences in both their clinical responses and the target specificity of the cytotoxic cells detected. The allogeneic T. annulata cell line recipients showed a very mild clinical response, and on day 9 after inoculation a strong cytotoxic response was detected. The response appeared to be directed against the allogeneic major histocompatibility complex (MHC) antigens of the inoculated cell line in some form of graft rejection response. By day 23 the predominant cytotoxic response was directed against the recipient animals' own cells infected with the parasite. In contrast, the autologous T. annulata cell line recipients showed very severe clinical reactions, and low levels of cytotoxicity were detected. The cytotoxicity was directed against parasite-infected targets but did not appear to be MHC restricted until day 20. Both groups were immune to a heterologous sporozoite challenge that proved lethal to two susceptible control animals, and on day 10 after challenge a peak of cytotoxicity was detected which was directed against the autologous infected target cell. This would suggest that this cytotoxic response was MHC restricted and was also cross-reactive between the heterologous parasite stocks used.

Animals↗
PubMed1989

Differential response of bovine T-cell lines to membrane and soluble antigens of Theileria parva schizont-infected cells.

T-cell lines generated from Theileria parva-immune cattle were used to identify antigens associated with schizont-infected lymphoblastoid cells. Homogenates prepared from T. parva-infected cells were fractionated by differential centrifugation, and antigenically distinct soluble and membrane-bound antigens were detected by the differential stimulation of cell lines derived from two animals. Activity in the soluble fraction was not attributable to either a mitogen or interleukin 2. Activity in the membrane fraction was associated with schizont membranes as indicated by the presence in this fraction of a parasite protein detected by immunoblot analysis using a schizont-specific monoclonal antibody. Elimination of intracellular schizonts over time, using the anti-theilerial drug, parvaquone, resulted in a concomitant loss of antigenicity in infected cells and in subcellular fractions prepared from drug-treated cells, demonstrating that stimulation of Theileria-specific helper and cytotoxic T-cell responses is associated with the presence of the parasite.

Animals↗
PubMed1990

Establishment of Theileria-infected bovine cell lines in scid mice.

Bovine cells transformed by infection with the protozoan parasite Theileria annulata were inoculated subcutaneously or intraperitoneally into C.B.-17 scid mice. Mice injected subcutaneously developed solid tumours at the injection site, whilst those injected intraperitoneally developed ascites. Schizont-infected cells were found in other tissues: infected cells spread much more easily from the intraperitoneal site. Karyotyping of cells isolated from tumours showed no evidence of transfer of parasites to murine cells. These results show that the scid mouse can be used as a host for Theileria-infected bovine cells.

Animals↗
PubMed1990

Identification of a Theileria mutans-specific antigen for use in an antibody and antigen detection ELISA.

Purified piroplasms of Theileria mutans were used to immunize BALB/c mice to generate monoclonal antibodies (MoAbs). The MoAbs recognized an antigen of a relative molecular mass of 32 kDa in Western blots. This antigen was also recognized by sera from cattle which had recovered naturally from experimental tick-transmission or infections induced by the blood stages of T. mutans. The MoAbs did not react, in indirect immunofluorescence or enzyme-linked immunosorbent assays (ELISA), with the common haemoparasites of cattle, namely, T. parva, T. annulata, Babesia bigemina, B. bovis, Anaplasma marginale, Trypanosoma congolense, T. vivax or T. brucei. An antigen capture ELISA was established with two of the MoAbs which recognized different epitopes on the 32 kDa molecule. Using this test it was possible to detect circulating antigens or immune complexes in sera collected from cattle during the acute or chronic phases of infection. When the purified 32 kDa protein was used as antigen in a micro-ELISA to detect circulating antibodies in both experimental and field cattle sera, it was found that the titres of antibodies ranged between 1:20 and 1:10,240. Results of this study indicate that the antigen and immune complex capture assays and the antibody detection ELISA can be complementary in the immunodiagnosis of acute and chronic T. mutans infections. Moreover, the tests are useful in the differential diagnosis of the disease and for epidemiological studies.

Animals↗