Sixty-eight dogs were examined for the presence of Encephalitozoon spp. antibodies. Twenty-one dogs (30.9%) were healthy without any clinical signs of diseases. Forty-seven animals (69.1%) developed clinical symptoms of diseases such as chronic otitis externa, conjunctivitis, upper respiratory tract inflammation, status epilepticus, pyodermatitis, skin hypersensitivity, demodicosis, flea allergy. Different detection methods of encephalitozoonosis including IFAT (Indirect Immunofluorescence Antibody Assay), in vitro cultivation, SDS PAGE electrophoresis, Western blot and PCR were applied. There were 33 (48.5%) positive reacting sera to E. cuniculi II (mouse type) antigen using IFAT, including 9 positive samples obtained from clinically healthy dogs. Sixteen samples with the antibody titer equal to 1 : 256 were then tested by Western Blot. Most of the samples reacted with E. cuniculi II and III antigens. The presence of E. intestinalis antibodies was lower and just a few samples reacted with E. hellem antigen. The electrophoretic analysis of the encephalitozoon strains used as antigens confirmed that they differ primarily in the molecular size. The strain of type II (mouse) expressed a double strip at 54 and 58 kDa level. The strain of type III (dog) expressed a wide strip at 59 kDa. E. cuniculi types II and III are more related in protein structure in comparison to the other analyzed strains. When PMP1/PMP2 primers were used in PCR, the size of the amplified product was 268 bp for E. cuniculi and 270 bp for E. intestinalis. A species-specific primer pair for E. cuniculi ECUNF/ECUNR gave a 549 bp fragment and V1/SI-500 primers specific for E. intestinalis gave a 370 bp fragment.
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