The genome of Ehrlichia risticii, the etiologic agent of Potomac horse fever, was cloned in the Xgtll expression vector. The efficiency of recombinant phage production with different restriction fragments of E. risticii DNA was generally between 20 and 95%. The antigen-positive frequency, detected by immunoscreening with E. risticii antibodies, was between 8 and 40 per 104 recombinants. Four (70, 55, 51, and 44 kDa) major antigens of E. risticii were identified from the recombinant phages by using recombinant antigen-selected monospecific antibodies. Characterization of three (70, 55, and 44 kDa) of these recombinant antigens indicated that the 70and 44-kDa polypeptides were ,I-galactosidase fusion products that were dependent on isopropylthiogalactoside induction for expression; they contained about 50 and 73%, respectively, of the native polypeptides. The 55-kDa antigen was a nonfusion protein expressed independently of isopropylthiogalactoside induction; it was a complete protein with a molecular weight identical to that of its native counterpart. The cloned E. risticii DNAs from of the recombinants expressing 70-, 55-, and 44-kDa proteins were 3.5, 3.9, and 4.8 kb, respectively, in size, and they were unique. The insert DNAs hybridized to multiple restriction fragments of the genomic DNA, the sum of the sizes of which was much greater than that of the corresponding insert. Mice immunized with the affinity-purified 55-kDa recombinant antigen produced a high titer of antibody in serum as measured by an enzyme-linked immunosorbent assay and gave a monospecific reaction by Western immunoblotting. Challenge infection of these immunized mice showed low protection from clinical infection.
Antigenic and genomic relatedness among Ehrlichia risticii, E . sennetsu, and E . canis was analyzed by enzyme-linked immunosorbent assay, Western blotting (immunoblotting) and DNA-DNA hybridization. E . risticii and E . sennetsu were serologically related, and their Western blot antigen profiles were nearly identical. Two antigens of E . sennetsu corresponding to the 28-and 51-kDa antigens of E . risticii were apparently larger than the E . risticii antigens, and the 55-kDa antigen of E . risticii appeared to be unique to this species. The 110-, 70-, and 44-kDa antigens of these two species were identical, as determined by the use of monospecific antibodies. DNA homology between these two species was high. On the other hand, E . canis was antigenically least reactive with the antisera to E . risticii and E . sennetsu. However, a dog convalescent-stage E . canis antiserum recognized antigens in the other two species which were different from those recognized by their homologous antisera. Similarly, homology between the DNA of E . canis and the DNAs of the other two species was very minimal. These results indicate that E . risticii and E . sennetsu are closely related both at the genomic and antigenic levels and that the relationship of these two species with E . canis is minimal.Members of the family Ehrlichieae belong to a group of obligate intracellular parasites with complex mechanisms of growth-and multiplication which are poorly understood. Ehrlichia risticii was identified in 1985 as the causative agent of Potomac horse fever (3, 8, 16). It has recently been reported that antibodies to E. risticii are common in the United States and Europe (7a, 18). E. sennetsu is the causative agent of human sennetsu fever, found mainly in western Japan, while E. canis causes canine ehrlichiosis (tropical canine pancytopenia) and occurs worldwide (17, 20). One common feature among these ehrlichieae is their predilection for the monocytic series of cells in their respective hosts (17).The renewed interest in the study of Ehrlichieae is due to the recent identification of E. risticii, serologic evidence of an association of human ehrlichiosis with E. canis (7,11,14), and recent isolation of the etiological agent from humans with ehrlichiosis (la). -The ability of E. risticii and E. sennetsu to experimentally infect nonhuman primates (22a) and the economic considerations of Potomac horse fever and canine ehrlichiosis are the other reasons for this interest.Present knowledge of the relatedness among Ehrlichieae is based on similarity in morphologic and biologic characteristics and cross-reactivities of antisera (8, 15, 19). The use of modern approaches in the identification of cross-reacting antigens and quantitation of serum cross-reactivity allows a reevaluation of the relatedness among these species in greater detail. Because the analysis of phenotypic expression involves less than 20% of the total genomes of the bacteria (l), it is necessary and now possible to evaluate these relationships at the genomic level. Toward the...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.