Characterization of the Major Antigenic Protein 2 of Ehrlichia canis and Ehrlichia chaffeensis and Its Application for Serodiagnosis of Ehrlichiosis

Abstract

Canine monocytic ehrlichiosis, caused by Ehrlichia canis or Ehrlichia chaffeensis, can result in clinical disease in naturally infected animals. Coinfections with these agents may be common in certain areas of endemicity. Currently, a species-specific method for serological diagnosis of monocytic ehrlichiosis is not available. Previously, we developed two indirect enzyme-linked immunosorbent assays (ELISAs) using the major antigenic protein 2 (MAP2) of E. chaffeensis and E. canis. In this study, we further characterized the conservation of MAP2 among various geographic isolates of each organism and determined if the recombinant MAP2 (rMAP2) of E. chaffeensis would cross-react with E. canis-infected dog sera. Genomic Southern blot analysis using digoxigenin-labeled species-specific probes suggested that map2 is a single-copy gene in both Ehrlichia species. Sequences of the single map2 genes of seven geographically different isolates of E. chaffeensis and five isolates of E. canis are highly conserved among the various isolates of each respective ehrlichial species. ELISA and Western blot analysis confirmed that the E. chaffeensis rMAP2 failed to serologically differentiate between E. canis and E. chaffeensis infections.

Identification of a Glycosylated Ehrlichia canis 19-Kilodalton Major Immunoreactive Protein with a Species-Specific Serine-Rich Glycopeptide Epitope

“…Unlike gp19, MAP2 appears to have a mass consistent with that predicted by its amino acid sequence and does not have any serine-rich domains. There is substantial homology among MAP2 orthologs in Ehrlichia spp., and cross-reactions among heterologous MAP2 proteins have been reported (14,17). In contrast, antibodies generated to E. canis gp19 were not crossreactive with E. chaffeensis VLPT, and therefore, these proteins are species-specific orthologs.…”

“…In contrast, antibodies generated to E. canis gp19 were not crossreactive with E. chaffeensis VLPT, and therefore, these proteins are species-specific orthologs. Other notable differences between MAP2 and gp19 include a major serine-rich linear epitope of gp19 that is strongly recognized by antibodies by Western immunoblotting, while antibodies to MAP2 of E. canis and E. chaffeensis appear to be directed primarily at a conformational epitope (1,2,14). In a previous study, we suggested that the 19-kDa major immunoreactive protein that we identified may be MAP2 (19); however, the data presented in this study support our conclusion that this protein is not MAP2 but rather gp19.…”

Progress and obstacles in vaccine development for the ehrlichioses

“…Fewer of these orthologs have been molecularly identified and characterized in E. ruminantium (MAP1 family [16 genes], MAP2 and mucin-like protein [clone hw26; TRP36/47 ortholog]) [92-94], but additional immunoreactive proteins have been identified in E. ruminantium that have not been described in E. chaffeensis or E. canis [59]. Immunoreactivity of the E. chaffeensis and E. canis MAP2 is primarily dependent on a major conformational epitope that does not react by western immunoblot [95,96], while E. ruminantium MAP2 appears to have a linear B-cell epitope [92]. …”

Characterization of Anaplasma phagocytophilum Major Surface Protein 5 and the Extent of Its Cross-Reactivity with A. marginale

“…The msp5 gene of Anaplasma spp. and its ortholog, map2 of Ehrlichia spp., are both known to be highly conserved within the genera Anaplasma and Ehrlichia (2,5,18,20,38). Additionally, based on the amino acid identity of 40.2% among all known orthologs of Msp5/Map2 of Anaplasma and Ehrlichia (3), cross-reactivity among organisms within the genera appears likely when Msp5 of Anaplasma phagocytophilum is used as a test antigen in serologic assays.…”