Characterization of Anaplasma phagocytophilum Major Surface Protein 5 and the Extent of Its Cross-Reactivity with A. marginale

Authors: STRIK, N - UNIVERSITY OF FLORIDA; ALLEMAN, A - UNIVERSITY OF FLORIDA; BARBET, A - UNIVERSITY OF FLORIDA; SORENSON, H - UNIVERSITY OF FLORIDA; WAMSLEY, H - UNIVERSITY OF FLORIDA; GASCHEN, F - UNIVERSITY OF BERN; LUCKSCHANDER, N - UNIVERSITY OF BERN; WONG, S - NY ST DEPT OF HEALTH; CHU, F - NY ST DEPT OF HEALTH; FOLEY, J - UNIVERSITY OF CAL DAVIS; BJOERSDORFF, A - KALMAR COUNTY HOSP, SWEDE; STUEN, S - NORWEGIAN SCHL OF VET SCI; Knowles Jr, Donald

Submitted to: Journal of Clinical Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/2/2007
Publication Date: 3/15/2007
Citation: Strik, N.I., Alleman, A.R., Barbet, A.F., Sorenson, H.L., Wamsley, H.L., Gaschen, F.P., Luckschander, N., Wong, S., Chu, F., Foley, J.E., Bjoersdorff, A., Stuen, S., Knowles Jr, D.P. 2007. Characterization of Anaplasma phagocytophilum Major Surface Protein 5 and the Extent of Its Cross-Reactivity with A. marginale. Journal of Clinical Microbiology. 14(3):262-268

Interpretive Summary: ARS and their WSU collaborators had previously developed, patented and transferred to industry a competitive inhibition ELISA (cELISA) for the detection of cattle persistently infected with A. marginale. Questions arose concerning the specificity of this test and whether it detected antibodies against other Anaplasma species, specifically A. phagocytophilium. This research showed that the commercially available cELISA for detecting antibody to A. marginale is specific for A. marginale and doesn't detect antibodies to A. phagocytophilium. However serologic cross-reactivity was identified when recombinant major surface protein (rMSP5) was used in an indirect ELISA format.

Technical Abstract: The major surface protein 5 (MSP5) of A. marginale is the antigen used in a commercially available competitive ELISA for serologic identification of infected cattle. MSP5 is highly conserved in the genus Anaplasma. The objectives of this investigation were to determine the degree of conservation of MSP5 among various geographical isolates of A. phagocytophilum and to identify the extent of serologic cross-reactivity of rMSP5 between A. marginale and A. phagocytophilum. The msp5 gene of A. phagocytophilum from various geographic isolates in several mammalian species was amplified, cloned and sequenced and the nucleotide sequences were compared. Recombinant MSP5 of A. phagocytophilum and A. marginale were used in an indirect ELISA to detect cross-reactivity in serum samples from humans and dogs infected with A. phagocytophilum and cattle infected with A. marginale. Serum samples were also tested by the competitive ELISA that uses monoclonal antibody ANAF16C1. There was 100% sequence identity in the msp5 genes among all the A. phagocytophilum isolates from the United States and a horse isolate from Sweden. The sheep isolates from Norway and the dog isolates from Sweden were 99% identical to each other and differed in 17 base pairs from the U.S. isolates and the horse isolate from Sweden. Serologic cross-reactivity was identified when serum samples from cattle infected with A. marginale were reacted with rMSP5 of A. phagocytophilum in an indirect ELISA and when serum samples from people and dogs infected with A. phagocytophilum were reacted with rMSP5 of A. marginale in an indirect ELISA format. Serum samples from dogs or humans infected with A. phagocytophilum did not cross-react with rMSP5 of A. marginale when tested with the commercially available cELISA with monoclonal antibody ANAF16C1. These results indicate that rMSP5 of A. phagocytophilum is highly conserved among isolates from the United States and Europe. Serologic distinction between infections with A. phagocytophilum 2 and A. marginale cannot be accomplished if rMSP5 from either organism is used in an indirect ELISA.