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Evaluation of peptide- and recombinant protein-based assays for detection of anti-Ehrlichia ewingii antibodies in experimentally and naturally infected dogs

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  • 1 Department of Immunoassay R&D, IDEXX Laboratories Incorporated, 1 Indexx Dr, Westbrook, ME 04092.
  • | 2 Department of Immunoassay R&D, IDEXX Laboratories Incorporated, 1 Indexx Dr, Westbrook, ME 04092.
  • | 3 Department of Immunoassay R&D, IDEXX Laboratories Incorporated, 1 Indexx Dr, Westbrook, ME 04092.
  • | 4 Department of Immunoassay R&D, IDEXX Laboratories Incorporated, 1 Indexx Dr, Westbrook, ME 04092.
  • | 5 Department of Immunoassay R&D, IDEXX Laboratories Incorporated, 1 Indexx Dr, Westbrook, ME 04092.
  • | 6 Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University Columbus, OH 43210.
  • | 7 Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University Columbus, OH 43210.
  • | 8 Southeastern Cooperative Wildlife Disease Study, Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, GA 30602.
  • | 9 Warnell School of Forestry and Natural Resources, University of Georgia, Athens, GA 30602.
  • | 10 Southeastern Cooperative Wildlife Disease Study, Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, GA 30602.
  • | 11 Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.
  • | 12 Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.
  • | 13 Clinical Pathology Laboratory School of Veterinary Medicine, Louisiana State University Baton Rouge, LA 70803.
  • | 14 Department of Immunoassay R&D, IDEXX Laboratories Incorporated, 1 Indexx Dr, Westbrook, ME 04092.

Abstract

Objective—To evaluate microtiter-plate format ELISAs constructed by use of different diagnostic targets derived from the Ehrlichia ewingii p28 outer membrane protein for detection of E ewingii antibodies in experimentally and naturally infected dogs.

Sample Population—Serum samples from 87 kenneled dogs, 9 dogs experimentally infected with anti-E ewingii, and 180 potentially naturally exposed dogs from Missouri.

Procedures—The capacities of the synthetic peptide and truncated recombinant protein to function as detection reagents in ELISAs were compared by use of PCR assay, western blot analysis, and a full-length recombinant protein ELISA. Diagnostic targets included an E ewingii synthetic peptide (EESP) and 2 recombinant proteins: a full-length E ewingii outer membrane protein (EEp28) and a truncated E ewingii outer membrane protein (EETp28)

Results—A subset of Ehrlichia canis-positive samples cross-reacted in the EEp28 ELISA; none were reactive in the EESP and EETp28 ELISAs. The EESP- and EETp28-based ELISAs detected E ewingii seroconversion at approximately the same time after infection as the EEp28 ELISAs. In afield population, each of the ELISAs identified the same 35 samples as reactive and 27 samples as nonreactive. Anaplasma and E can is peptides used in a commercially available ELISA platform did not detect anti-E ewingii antibodies in experimentally infected dogs.

Conclusions and Clinical Relevance—The EESP and EETp28 ELISAs were suitable for specifically detecting anti-E ewingii antibodies in experimentally and naturally infected dogs. [Am J Vet Res 2010;71:1195-1200)

Abstract

Objective—To evaluate microtiter-plate format ELISAs constructed by use of different diagnostic targets derived from the Ehrlichia ewingii p28 outer membrane protein for detection of E ewingii antibodies in experimentally and naturally infected dogs.

Sample Population—Serum samples from 87 kenneled dogs, 9 dogs experimentally infected with anti-E ewingii, and 180 potentially naturally exposed dogs from Missouri.

Procedures—The capacities of the synthetic peptide and truncated recombinant protein to function as detection reagents in ELISAs were compared by use of PCR assay, western blot analysis, and a full-length recombinant protein ELISA. Diagnostic targets included an E ewingii synthetic peptide (EESP) and 2 recombinant proteins: a full-length E ewingii outer membrane protein (EEp28) and a truncated E ewingii outer membrane protein (EETp28)

Results—A subset of Ehrlichia canis-positive samples cross-reacted in the EEp28 ELISA; none were reactive in the EESP and EETp28 ELISAs. The EESP- and EETp28-based ELISAs detected E ewingii seroconversion at approximately the same time after infection as the EEp28 ELISAs. In afield population, each of the ELISAs identified the same 35 samples as reactive and 27 samples as nonreactive. Anaplasma and E can is peptides used in a commercially available ELISA platform did not detect anti-E ewingii antibodies in experimentally infected dogs.

Conclusions and Clinical Relevance—The EESP and EETp28 ELISAs were suitable for specifically detecting anti-E ewingii antibodies in experimentally and naturally infected dogs. [Am J Vet Res 2010;71:1195-1200)

Contributor Notes

Address correspondence to Dr. O'Connor (tom-oconnor@idexx.com)

Presented in part at the annual meetings of the American College of Veterinary Internal Medicine, Seattle, June 2007, and Montreal, June 2009.