Neospora caninum is recognized as a major cause of infectious abortion in cattle. Very little is known about immunity to Neospora. Cell mediated responses have previously been shown to be important in the development of protective immunity to the closely related parasite Toxoplasma gondii, and may therefore be an important component in the immune response to Neospora. In this paper we report that a group of low molecular weight NCI strain tachyzoite antigens (< or = 30 kDa) separated by SDS PAGE and bound to nitrocellulose membrane stimulated proliferation in vitro of CD4+ T cells from calves experimentally infected with N. caninum. Proliferation was accompanied by production of high concentrations of IFN-gamma. Several of these antigens were also recognized by antibody produced in these animals. As the most effective vaccines require the stimulation of both humoral and cell mediated immune responses, these antigens may be important in the development of a vaccine against neosporosis.

Iodination of intact Pasteurella haemolytica serotype A2 cells labelled a sub-set of total cellular proteins. Comparison of the autoradiographic patterns obtained from iodinated cells grown on complete medium and on iron-depleted medium showed that expression of three proteins, of 100,70 and 35 kDa, respectively, was increased by growth under iron-depleted conditions. Of these proteins, that of 35 kDa had not been reported previously. Like the 100 and 70 kDa proteins, the 35 kDa protein was expressed in natural infections, since it was recognized by antiserum from sheep that had recovered from an experimental infection with P. haemolytica A2. The 35 kDa protein was partially purified by reverse-phase HPLC and was found to be antigenic in both sheep and mice. A monoclonal antibody that was specific for the 35 kDa protein was used to identify the cellular location of the protein by immunoblotting of cell fractions enriched for particular cellular components. This demonstrated that the 35 kDa protein was located mainly in the periplasm.

Sheep immunized with Toxoplasma gondii (Toxovax) prior to pregnancy were tested for their ability to withstand a challenge at 90 days gestation with 107 Neospora caninum (NC1) tachyzoites. The antibody responses in sheep following immunization with T. gondii were specific for T. gondii whereas peripheral blood mononuclear cells responded to both T. gondii and N. caninum antigen in vitro. This suggested that there was induction of crossreactive immune recognition in the sheep, at least at the cellular level. Following challenge of sheep at mid-gestation with N. caninum, no febrile responses were recorded in the group of sheep which had previously received Toxovax while significant febrile responses were recorded in the group of sheep which received N. caninum challenge alone. Antibody responses to N. caninum developed in all sheep following N. caninum challenge and antibody responses to T. gondii were boosted in the group of sheep which had previously been immunized with Toxovax. No antibodies to T. gondii were observed in the sheep which received the N. caninum challenge alone. Peripheral blood mononuclear cells from both groups of sheep responded to T. gondii and N. caninum antigen in vitro and interferon gamma was present in the cell-free supernatant from activated cells. However despite evidence of the induction of crossreactive immunity between T. gondii and N. caninum, this was not sufficient to prevent foetal death. The group of sheep which had received Toxovax prior to pregnancy and the group of sheep which only received the N. caninum challenge experienced 100% foetal death compared with 0% in the unchallenged control group. Vaccination prior to pregnancy with Toxovax did protect against foetal death following oral challenge at 90 days with 2000 T. gondii oocysts which caused 100% foetal death in a control challenge group.

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