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Title:   Author  item
                                            Hill, Doloresitem
                                            SCOTT, ALAN - JOHNS HOPKINS, BALTO., MDitem
                                            DYER, ROBERT - UNIV MARYLAND, COLLEGE PKitem
                                            Dubey, Jitender
  Submitted to: International Journal for Parasitology
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 6/15/2001
                                    Publication  Date: N/A
                                    Citation: N/AInterpretive Summary: Neospora caninum is a protozoan parasite which has been shown to cause abortion in dairy and beef cattle, as well as neurological disorders in dogs and horses.  Vertical transmission has been documented in cattle, though the fidelity of transmission from dam to offspring is variable (Davison et al., 1999; French et  al., 1999; Schares et al., 1998; Dyer et al., 2000). The development of resistance to Neospora infection and transmission of N. caninum from dams to offspring may be affected by immunological recognition of specific N. caninum proteins by infected cattle.  Major efforts in the field of parasite proteomics (Banks et al., 2000; Chambers et al., 2000; Gavaghan, 2000) have been undertaken in an attempt to relate genomic information to protein products which may have important roles in  disease (Alaiya et al., 1999; Edgar et al., 2000), as well as development, transmission, and resistance to parasite pathogens of humans and livestock (Barrett et al., 2000; Gutierrez, 2000). In this study, we have used 2-dimensional gel electrophoresis, Western blotting, and N-terminal amino acid sequence information to identify specific proteins from N. caninum (NC-1) which are recognized  as immunodominant by sera from N. caninum infected cattle.
                                Technical Abstract: Neospora caninum tachyzoites were isolated from in vitro cell culture and proteins were extracted using a chaotropic detergent solution.  Two-dimensional electrophoresis was carried out on the solubilized proteins using immobilized pH gradient (3-10) strips in the first dimension and 4-12% gradient slab gels in the second  dimension. Separated proteins were blotted onto nylon membranes and subjected to Western blot analysis using polyclonal antiserum from mature cows (positive at 1:1200 for N. caninum by IFA ) from a N. caninum endemic farm.  Proteins identified in Western blots by antisera from infected cows were cut from identical blots run in parallel and were subjected to N-terminal amino acid sequence analysis for identification.  Twenty  immunodominant proteins were identified using  sera from N. caninum infected cattle.  Four of the  proteins were identified as surface associated proteins (SAG-1, SRS-2, Nc-p35, and Nc-p43); 3 were identified as dense granule-associated proteins (NTPase, Nc-GRA-7, and Nc-GRA- 2), 4 were identified as actin, myosin, or tubulin chains,  2 were identified as microneme associated proteins (Nc-p38 and Nc- MIC-10), and 3 were identified as cytoplasmic proteins (HGPRT, Nc-SUL-1, 14-3-3 antigen).  Two proteins were found which showed no homology with existing SwissPro entries, and may represent proteins which are unique to N. caninum


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