Molecular karyotyping and ribosomal-RNA studies of Sarcocystis muris, phylum Apicomplexa

The histopathology of normal beef carcasses and those condemned for eosinophilic myositis (EM) were described. Ultrastructural and transmission studies indicated that Sarcocystis cruzi (Apicomplexa: Eimeriina) is involved in granulomatous lesions, and probably that cats do not play a major role in the transmission of Sarcocystis spp. to cattle in parts of the USA. Results of pepsin-HCl digestion assays as well as histology showed that normal carcasses generally contained more infective parasites than animals that were condemned for EM, and seem to have greater potential of transmitting the infection. Pulsed field gel electrophoresis was used to determine the molecular karyotype of S. muris. Nine distinct bands of chromosome-sized DNA molecules between 0.2 and approximately 8 megabase pairs (Mb) were separated, and most chromosomes were between 3 and 8 Mb. Three of the bands were probably doublets because of their increased widths and intense staining. It is interpreted that there are at least 12 chromosomes in S. muris and probably there is no polymorphism between developmental stages of the parasite. However, there were size differences between the chromosomes of S. muris and S. cruzi. The small subunit-rRNA (ss-rRNA) gene of S. muris bradyzoites was isolated, cloned and sequenced. The inferred nucleotide sequence of the ss-rRNA was compared pairwise with those of 23 other organisms, in order to determine phylogenetic relationships. The phylum Apicomplexa appears to have a close affinity to the ciliates, and S. muris is much more closely related to the dinoflagellates than to Plasmodium spp. Apparently, modern dinoflagellates and coccidia share the same ancestral stock. Two regions of the primary structures of the ss-rRNA of S. muris were used to design hybridization probes for the detection and identification of the parasite. Total cellular RNA of purified coccidia or parasite-infected tissues were assayed. One probe detected only S. muris and another successfully hybridized to several species of coccidia, including S. muris, Toxoplasma gondii and Eimeria nieschulzi. Both probes displayed exquisite sensitivity that was capable of detecting single organisms, and can assay low levels of infections not detectable by other methods.