No abstract

Ehrlichia risticii has a close antigenic relationship to E. sennetsu. Sera of ponies experimentally infected with E. risticii, the etiologic agent of Potomac horse fever, consistently reacted with E. sennetsu, a human pathogen, in indirect fluorescent-antibody (IFA) testing, while human E. sennetsu convalescent serum reacted with E. risticii by IFA testing and immunoferritin labeling of cells infected in vitro. Two ponies injected intravenously with live E. sennetsu did not develop clinical illness. Subsequent injection with live E. risticii also did not induce any disease, in contrast to two control ponies given E. risticii without prior exposure to E. sennetsu. Both controls developed fever, anorexia, depression, dehydration, and diarrhea, which are typical clinical signs of Potomac horse fever, and had characteristic lesions of enteritis and lymph node histiocytosis at postmortem examination. E. sennetsu-exposed ponies had normal gastrointestinal morphologies and lymph node hyperplasia. Ponies primed with E. sennetsu before E. risticii challenge developedhigh titers of immunoglobulin G antibody which reacted against both E. sennetsu and E. risticii antigens by IFA testing. The most prominent antigenic polypeptide in Western (immuno-) blot analysis of sera collected from ponies primed with E. sennetsu before subsequent challenge with E. risticii was present in lysates of both Ehrlichia species and had an apparent molecular mass of 44 kilodaltons. This band was not prominent in Western blots performed with sera of ponies injected with E. risticii alone. Thus, injection of E. sennetsu protects ponies from clinical and pathological manifestations of the disease induced by injection with E. risticii. Immunologic cross-reactivity of the two organisms with IFA testing and strong immunologic recognition by ponies of the 44-kilodalton antigen common to the two organisms may be related to the development of protective immunity against E. risticii.

An enzyme-linked immunosorbent assay (ELISA) for immunoglobulin G (IgG) and IgM in natural and experimental infections of equids with Ehrlichia risticii was developed. Ehrlichial organisms purified from an infected mouse macrophage cell line were used as the antigen. IgM was separated from serum IgG by the expedient of spun-column chromatography, allowing the use of an indirect ELISA for quantitation of both IgG and IgM in the test sera. Among 16 paired sera from horses exhibiting clinical signs of Potomac horse fever, 8 were positive by the indirect fluorescent-antibody test (IFA), 11 were positive by the IgG ELISA, and 8 were positive by the IgM ELISA. All IFA-positive specimens were positive by the IgG ELISA, which appeared to be more sensitive than the IFA. In all cases, the IgG ELISA alone would have sufficed for diagnosis when acuteand convalescent-phase sera were available. When 26 single acuteor convalescent-phase serum samples were tested, the IFA detected 8, the IgG ELISA detected 10, and the IgM ELISA detected 6 positive serum specimens. The kinetics of IgG and IgM responses as determined by ELISA in two experimentally infected ponies which survived infection and challenges revealed that specific IgM was short-lived, falling to undetectable levels by day 60 postinoculation, whereas specific IgG persisted for more than 1 year. IgM and IgG were detected as early as days 1 and 10, respectively, postinoculation. The results suggest that the ELISA is more sensitive than the IFA and that the IgM ELISA may provide a means for early diagnosis of Potomac horse fever at or before the onset of clinical signs.

A five-year survey of ticks and associated rickettsia in Ohio shows a partitioning of spotted fever group rickettsia (namely, Rickettsia montana and R. rickettsii) into three areas of the state where the majority of human spotted fever cases occur. The percentage of ticks infected, even in areas with a high incidence of disease, is remarkably low, less than 1%. The statewide infection rate for R. bellii approaches 3%, and this organism appears to be more evenly distributed throughout the state. Dermacentor variabilis, which accounts for 97% of Ohio ticks received for testing, is the only species which has been found to harbor rickettsia. Ohio's tick testing program is not an effective surveillance tool and is of no use in diagnosing tick-borne disease. However, the program does function as an excellent means of increasing public awareness of tick-borne disease.

Neorickettsia helminthoeca (tribe Ehrlichieae, family Rickettsiaceae) is the agent of salmon poisoning disease, which affects members of the family Canidae. This bacterium is unusual in that it is the only known obligately intracellular bacterium that is transmitted via a helminth vector. The nucleotide sequence of the N. helminthoeca 16s rRNA gene was determined and compared with the sequences of intracellular bacteria belonging to the alpha subgroup of the Proteobacteria. The N. helminthoeca sequence was most similar to the sequences of two Ehrlichia species, Ehrlichia risticii and Ehrlichia sennetsu (levels of sequence similarity, > 95%). All other members of the tribe Ehrlichieae, including members of the other Ehrlichia species, and the related species Cowdria ruminantium and Anaphsma ma~'naZe, were only distantly related phylogenetically (levels of sequence similarity, 84 to 86%). Our results corroborate the results of previous ultrastructural and Western blot (immunoblot) comparisons of N. helminthoeca with other ehrlichial species. The genus Ehrlichia is phylogenetically incoherent and can be separated into three identifiable clusters of species. Each cluster is closely associated with a species classified in another non-Ehrlichia bacterial genus. The close relationships among N. helminthoeca, E. risticii, and E. sennetsu and the striking differences between these organisms and other members of the tribe Ehrlichieae suggest that in the future, these organisms should be treated as members of a new bacterial genus separate from the genus Ehrlichia.