Molecular cloning and analysis of recombinant major antigens of Ehrlichia risticii

Dutta, S.; Shankarappa, Basavaraju; Mattingly-Napier, Bonnie L.

doi:10.1128/iai.59.3.1162-1169.1991

Cited by 14 publications

(17 citation statements)

References 21 publications

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“…Variation in antigenic proteins is a widespread and common characteristic of many bacteria, including E. risticii (8,12,13,41). For example, there are strains of E. risticii which have been described that differ in their antigenic components but exhibit identity at the 16S rRNA gene level (41).…”

Section: Discussionmentioning

confidence: 99%

“…Segments of two additional ehrlichia genes were amplified by PCR. Sets of nested primers were designed to detect portions of the E. risticii homolog of the Escherichia coli groESL heat shock operon (39) and the E. risticii 51-kDa major antigen gene (12,13,41). The following primer sequences were used to amplify the heat shock operon: 5Ј-ACCAGGCTACCTCACAGGC-3Ј and 5Ј-TTGACC CTCGCATCAATG-3Ј (outer primers); and 5Ј-CACAAGTTGGTTCAATTTC TGC-3Ј and 5Ј-CCGAGATCTTCAACAGTAAGGC-3Ј (inner primers).…”

Section: Snail Collectionmentioning

confidence: 99%

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Detection ofEhrlichia risticii, the Agent of Potomac Horse Fever, in Freshwater Stream Snails (Pleuroceridae:Jugaspp.) from Northern California

Barlough¹,

Reubel²,

Madigan³

et al. 1998

Appl Environ Microbiol

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Ehrlichia DNA was identified by nested PCR in operculate snails (Pleuroceridae: Juga spp.) collected from stream water in a northern California pasture in which Potomac horse fever (PHF) is enzootic. Sequencing of PCR-amplified DNA from a suite of genes (the 16S rRNA, groESL heat shock operon, 51-kDa major antigen genes) indicated that the source organism closely resembledEhrlichia risticii, the causative agent of PHF. The minimum percentage of Juga spp. harboring the organism in the population studied was 3.5% (2 of 57 snails). No ehrlichia DNA was found in tissues of 123 lymnaeid, physid, and planorbid snails collected at the same site. These data suggest that pleurocerid stream snails may play a role in the life cycle of E. risticii in northern California.

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Section: Discussionmentioning

confidence: 99%

Section: Snail Collectionmentioning

confidence: 99%

Detection ofEhrlichia risticii, the Agent of Potomac Horse Fever, in Freshwater Stream Snails (Pleuroceridae:Jugaspp.) from Northern California

Barlough¹,

Reubel²,

Madigan³

et al. 1998

Appl Environ Microbiol

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“…Western blotting was performed according to the procedure described previously (25) with mouse and horse sera collected at 3 and 6 weeks p.i., respectively. Western blotting was also performed with the specific recombinant clone-specific antibodies prepared by eluting the absorbed rabbit anti-ER 90-12 antibodies from a nitrocellulose membrane containing the recombinant ZAP phage expressing the E. risticii antigens according to the previously described procedure (11).…”

mentioning

confidence: 99%

“…Southern blotting. Southern blotting was done with the genomic DNA of both the strains and the probes made from the inserts of ZAP clones expressing the 85-, 55-, and 51-kDa antigens of the 90-12 strain by the previously described procedure (11). The DNA of each strain was extracted from the Renografinpurified organisms as described earlier (12).…”

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confidence: 99%

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

1995

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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 strain, 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 homologous and heterologous antigens. The differences in antigen profiles were demonstrated by Western blot (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 85-kDa recombinant clones of the 90-12 strain showed a similar pattern with probes of the 51-and 55-kDa 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 interferontreated 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.

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“…Additionally, vaccine efficacy has been reported to be marginal, likely due to deficient immunogenic responses [13,20,22]. Studies have suggested that strain variants having pathogenic, immunologic and molecular differences may all contribute to vaccine failure [13,23]. Large controlled field studies evaluating the antibody response of horses to PHF vaccination have not been reported.…”

Section: Discussionmentioning

confidence: 99%

Immunogenicity of Potomac horse fever vaccine when simultaneously co‐administered with rabies vaccine in a multivalent vaccine or as two monovalent vaccines at separate sites

McKenzie

Funk

Trager

et al. 2019

Equine Veterinary Journal

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SummaryBackgroundPotomac horse fever (PHF) is a potentially fatal enterocolitis of horses caused by Neorickettsia risticii. The disease was originally recognised almost 40 years ago in the state of Maryland in the US. It is now known to occur in many areas of North America, as well as having been described in South America and Europe. Monocomponent PHF vaccines are available, but clinical protection with vaccination has been reported to be inconsistent.ObjectivesThis study was designed to assess the immunogenicity of a commercially available Potomac Horse Fever (PHF) vaccine when administered as either a monovalent PHF vaccine simultaneously co‐administered with a separate monovalent Rabies vaccine or as a multivalent PHF/Rabies vaccine in horses.Study designRandomised parallel group trial.MethodsNinety‐one client or University owned horses participated in this open‐label randomised study, with 45 horses receiving the monovalent vaccines at separate sites and 46 receiving the multivalent vaccine at a single site. Serum PHF IFA titres were determined twice prior to vaccination and at 1, 2 and 3 months after vaccination.ResultsBoth vaccination protocols exhibited poor immunogenicity, with only one‐third of all the animals demonstrating seroconversion, defined as an increase in titre of greater than 400 over baseline, at any time point after vaccination. The monovalent PHF vaccine exhibited significantly greater immunogenicity in terms of the number of horses exhibiting seroconversion, as compared to the multivalent vaccine, at one (20 vs. 11, P = 0.03) and two (18 vs. 9, p = 0.02) months post vaccination. The monovalent PHF vaccine also exhibited significantly greater immunogenicity in terms of the median (interquartile range) IFA titres, as compared to the multivalent vaccine, at one (800 [200–1600] vs. 400 [200–800], P = 0.009) and 2 months (400 [200–1600] vs. 400 [100–800], P = 0.02) post vaccination. There was no significant difference between groups at 3 months in either seroconversion rate or median IFA titers.Main limitationsThis study did not assess the actual protective effects of PHF vaccination but rather used the serologic response to vaccination as a surrogate biomarker of immunity.ConclusionsThe multivalent PHF/Rabies vaccine exhibited lower immunogenicity as compared to the monovalent PHF vaccine co‐administered with a separate Rabies vaccine.

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Molecular cloning and analysis of recombinant major antigens of Ehrlichia risticii

Cited by 14 publications

References 21 publications

Detection ofEhrlichia risticii, the Agent of Potomac Horse Fever, in Freshwater Stream Snails (Pleuroceridae:Jugaspp.) from Northern California

Detection ofEhrlichia risticii, the Agent of Potomac Horse Fever, in Freshwater Stream Snails (Pleuroceridae:Jugaspp.) from Northern California

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

Immunogenicity of Potomac horse fever vaccine when simultaneously co‐administered with rabies vaccine in a multivalent vaccine or as two monovalent vaccines at separate sites

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