Equine protozoal myeloencephalitis (EPM) due to Sarcocystis neurona infection is 1 of the most common neurologic diseases in horses in the United States. The mechanisms by which most horses resist disease, as well as the possible mechanisms by which the immune system may be suppressed in horses that develop EPM, are not known. Therefore, the objectives of this study were to determine whether horses experimentally infected with S. neurona developed suppressed immune responses. Thirteen horses that were negative for S. neurona antibodies in serum and cerebrospinal fluid (CSF) were randomly assigned to control (n = 5) or infected (n = 8) treatment groups. Neurologic exams and cerebrospinal fluid analyses were performed prior to, and following, S. neurona infection. Prior to, and at multiple time points following infection, immune parameters were determined. All 8 S. neurona –infected horses developed clinical signs consistent with EPM, and had S. neurona antibodies in the serum and CSF. Both infected and control horses had increased percentages ( P < 0.05) of B cells at 28 days postinfection. Infected horses had significantly decreased ( P < 0.05) proliferation responses as measured by thymidine incorporation to nonspecific mitogens phorbol myristate acetate (PMA) and ionomycin (I) as soon as 2 days postinfection.

Immunodeficient B-cell-deficient mice (mμMT) were infected with Sarcocystis neurona merozoites to determine the role of B cells and the humoral immune response in protective immunity. As expected, the mice did not seroconvert based on a direct agglutination test. Infected mice did not have significant changes in gross pathology at the time points examined. Histologic changes included mild perivascular and peribronchial infiltrate in the lungs; perivascular infiltrate and mild inflammatory sinusoidal foci in the liver; prominent high endothelial venules in the lymph nodes; and moderate cellular expansion of the periarteriolar sheaths (PALS) in the spleen. Changes resolved by day 60 postinfection. Mice developed significant CD4 and CD8 responses in lymphoid organs, including significant effector (CD45RB high ) and memory (CD44 high ) CD4 and CD8 responses. Flow cytometry confirmed the lack of B cells. Overall, these data suggest that B cells are not critical to the protective immune response to SN infection.

Immunocompetent C57BL/6 mice were infected with Sarcocystis neurona merozoites to assess the protective immune response to active infection. Using a direct agglutination test, all infected mice seroconverted to S. neurona merozoite antigens by day 14 postinfection (PI). Further, mice developed splenomegaly and bilateral symmetrical lymphadenopathy by day 14 PI, which appeared to be resolving by day 28 PI. Histologic analysis revealed a marked increase in germinal center formation in the spleen and lymph nodes by day 14 PI. Corresponding to gross and histopathological changes, the percentage of B-cells decreased significantly by day 14 PI but then increased significantly and persisted at day 28 PI in the blood, spleen, and multiple lymph nodes. There was a sharp nonspecific significant decrease in CD4 percentages by day 14 PI in the blood, spleen, and lymph nodes. Early-activation CD8 lymphocytes (CD62/CD8) were significantly down-regulated coinciding with a significant compensatory up-regulation of memory (CD44/CD8) lymphocytes in multiple organs. We propose that the protective cell-mediated immune response to S. neurona involves both CD4 and CD8 cells, with CD8 lymphocytes appearing to play a more critical role.

Interferon-gamma knockout (IFN-gamma KO) mice were infected with Sarcocystis neurona merozoites to characterize the immunopathology associated with infection. By day 14 postinfection (PI), mice developed splenomegaly and lymphadenopathy, characterized by marked lymphoid hyperplasia with increased numbers of germinal centers. Additional histopathologic changes included increased extramedullary hematopoiesis, multifocal mixed inflammatory infiltrates in the liver, perivascular infiltrate of the liver and lung, and interstitial pneumonia. The total number of B-cell splenocytes ( P < 0.05) and the percentage of B-cells increased on day 14 PI in the spleen and on day 28 PI in the lymph nodes ( P < 0.05). By day 28 PI, the number of B-cell splenocytes decreased significantly. A non–subset-specific decrease in percentages of CD4 lymphocytes throughout all lymphoid organs was observed on day 14 PI. However, total CD4 and CD44/CD4 splenocytes increased significantly by day 28 PI. Early-activation CD8 lymphocytes were reduced in the blood and spleen, whereas memory CD8 lymphocyte percentages and total numbers were significantly increased. On the basis of the results, we propose that S. neurona –infected IFN-gamma KO mice are immunocompromised and unable to clear the infection. Thus, they develop B-cell exhaustion and a delayed, but sustained, increased number of memory CD4 and CD8 lymphocytes due to chronic antigen stimulation.