Pathogenic, immunologic, and molecular differences between two Ehrlichia risticii strains

Ehrlichia risticii is the causative agent of Potomac horse fever (PHF), an acute infectious disease of horses. In the last few years, there have been several reports of PHF cases occurring even in vaccinated horses. We isolated a new strain of E. risticii (90-12 strain) from a vaccinated horse suffering from clinical PHF. The major pathogenic, immunologic, and molecular differences between the 90-12 strain and the 25-D stain, which was originally isolated during the outbreaks in 1984, were studied. The 90-12 strain was more pathogenic for mice and horses compared with the 25-D strain. In enzyme-linked immunosorbent assay and immunofluorescence assay with mouse and horse antisera of both the strains, two- to fourfold differences were observed between (immunoblot) with mouse and horse antisera and also with the recombinant clone-specific antibodies. Though several antigens were similar in both the strains, there were significant differences between them in the 110-, 85-, 70-, 51-, and 33-kDa antigens. The 85-kDa antigen was present only in the 90-12 strain but cross-reacted with a 50-kDa antigen of the 25-D strain. The 51-kDa antigens of both strains had different migration patterns, Southern blot hybridization of the genome from both the strains with DNA probes made from the 51-, 55-, and clones for both the strains, whereas the probe of the 85-kDa clone showed a completely different pattern. The 16S rRNA gene sequences from the two strains were identical. Neither strain replicated in gamma interferon-treated mouse peritoneal macrophages. In in vitro neutralization assay, sera from the 25-D strain-infected horse neutralized the homologous strain but did not neutralize the 90-12 strain, whereas sera from the 90-12 strain-infected horse neutralized both the strains. In mouse protection experiments, there was complete homologous protection. But in cross-protection, mice immunized with the 25-D strain were only partially protected against challenge with the 90-12 strain, whereas mice immunized with the 90-12 strain were completely protected against the 25-D strain challenge. These results clearly indicate that there are major differences between the 90-12 and 25-D strains which may have implications regarding the vaccine failure for PHF and the development of an efficient vaccine.