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

ABSTRACT Major surface protein 5 (Msp5) of Anaplasma marginale is highly conserved in the genus Anaplasma and the antigen used in a commercially available competitive enzyme-linked immunosorbent assay (cELISA) for serologic identification of cattle with anaplasmosis. This study analyzes the degrees of conservation of Msp5 among various isolates of Anaplasma phagocytophilum and the extent of serologic cross-reactivity between recombinant Msp5 (rMsp5) of Anaplasma marginale and A. phagocytophilum. The msp5 genes from various isolates of A. phagocytophilum were sequenced and compared. rMsp5 proteins of A. phagocytophilum and A. marginale were used separately 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 with a commercially available competitive ELISA that uses monoclonal antibody ANAF16C1. There were 100% sequence identities in the msp5 genes among all of the A. phagocytophilum isolates from the United States and a horse isolate from Sweden. Sheep isolates from Norway and dog isolates from Sweden were 99% identical to one another but differed in 17 base pairs from the United States isolates and the horse isolate. Serologic cross-reactivity was identified when serum samples from cattle infected with A. marginale were reacted with rMsp5 of A. phagocytophilum and when serum samples from humans 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. These results suggest that rMsp5 of A. phagocytophilum is highly conserved among United States and European isolates and that serologic distinction between A. phagocytophilum and A. marginale infections cannot be accomplished if rMsp5 from either organism is used in an indirect ELISA.

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Infection of Tick Cells and Bovine Erythrocytes with One Genotype of the Intracellular Ehrlichia Anaplasma marginale Excludes Infection with Other Genotypes

Clinical and Vaccine Immunology ◽

10.1128/cdli.9.3.658-668.2002 ◽

2002 ◽

Vol 9 (3) ◽

pp. 658-668 ◽

Cited By ~ 11

Author(s):

José de la Fuente ◽

Jose C. Garcia-Garcia ◽

Edmour F. Blouin ◽

Jeremiah T. Saliki ◽

Katherine M. Kocan

Keyword(s):

United States ◽

Tandem Repeats ◽

Surface Protein ◽

Anaplasma Marginale ◽

The United States ◽

The Other ◽

Tick Cell ◽

Anaplasma Ovis ◽

Major Surface ◽

Bovine Erythrocytes

ABSTRACT Anaplasma marginale, a tick-borne rickettsial pathogen of cattle, is endemic in several areas of the United States. Many geographic isolates of A. marginale that occur in the United States are characterized by the major surface protein 1a, which varies in sequence and molecular weight due to different numbers of tandem repeats of 28 or 29 amino acids. Recent studies (G. H. Palmer, F. R. Rurangirwa, and T. F. McElwain, J. Clin. Microbiol. 39:631-635, 2001) of an A. marginale-infected herd of cattle in an area of endemicity demonstrated that multiple msp1α genotypes were present but that only one genotype was found per individual bovine. These findings suggested that infection of cattle with other genotypes was excluded. The present study was undertaken to confirm the phenomenon of infection exclusion of A. marginale genotypes in infected bovine erythrocytes and cultured tick cells. Two tick-transmissible isolates of A. marginale, one from Virginia and one from Oklahoma, were used for these studies. In two separate trials, cattle inoculated with equal doses of the two isolates developed infection with only one genotype. Tick cell cultures inoculated with equal doses of the two isolates became infected with only the Virginia isolate of A. marginale. When cultures were inoculated with different ratios of the Oklahoma and Virginia isolates of A. marginale, the isolate inoculated in the higher ratio became established and excluded infection with the other. When cultures with established infections of one isolate were subsequently infected with the other, only the established isolate was detected. We documented infection exclusion during initial infection in cell culture by labeling each isolate with a different fluorescent dye. After 2 days in culture, only a single isolate was detected per cell by fluorescence microscopy. Finally, when Anaplasma ovis infections were established in cultures that were subsequently inoculated with the Virginia or Oklahoma isolate of A. marginale, A. marginale infection was excluded. These studies confirm that infection exclusion occurs with A. marginale in bovine erythrocytes and tick cells, resulting in the establishment of only one genotype, and appears to be the first report of infection exclusion for Anaplasma and Ehrlichia species.

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Characterization of Anaplasma marginale Isolated from North American Bison

Applied and Environmental Microbiology ◽

10.1128/aem.69.8.5001-5005.2003 ◽

2003 ◽

Vol 69 (8) ◽

pp. 5001-5005 ◽

Cited By ~ 28

Author(s):

José de la Fuente ◽

Elizabeth J. Golsteyn Thomas ◽

Ronald A. Van Den Bussche ◽

Robert G. Hamilton ◽

Elaine E. Tanaka ◽

...

Keyword(s):

New World ◽

Surface Protein ◽

Anaplasma Marginale ◽

The United States ◽

Bison Bison ◽

Infected Cattle ◽

Major Surface Protein ◽

The World ◽

Major Surface

ABSTRACT Anaplasma marginale (Rickettsiales: Anaplasmataceae), a tick-borne pathogen of cattle, is endemic in tropical and subtropical regions of the world. Although serologic tests have identified American bison, Bison bison, as being infected with A. marginale, the present study was undertaken to confirm A. marginale infection and to characterize isolates obtained from naturally infected bison in the United States and Canada. Major surface protein (MSP1a and MSP4) sequences of bison isolates were characterized in comparison with New World cattle isolates. Blood from one U.S. bison was inoculated into a susceptible, splenectomized calf, which developed acute anaplasmosis, demonstrating infectivity of this A. marginale bison isolate for cattle. The results of this study showed that these A. marginale isolates obtained from bison were similar to ones from naturally infected cattle.

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Development and evaluation of a double-antigen sandwich ELISA to identify Anaplasma marginale–infected and A. centrale–vaccinated cattle

Journal of Veterinary Diagnostic Investigation ◽

10.1177/1040638719892953 ◽

2019 ◽

Vol 32 (1) ◽

pp. 70-76 ◽

Cited By ~ 1

Author(s):

Macarena Sarli ◽

Carolina S. Thompson ◽

María B. Novoa ◽

Beatriz S. Valentini ◽

Mariano Mastropaolo ◽

...

Keyword(s):

Sandwich Elisa ◽

Surface Protein ◽

Anaplasma Marginale ◽

Epidemiologic Studies ◽

Competitive Elisa ◽

Live Vaccine ◽

Serum Samples ◽

Agreement Rate ◽

Major Surface Protein ◽

Major Surface

Bovine anaplasmosis is a worldwide infectious disease caused by the intraerythrocytic bacterium Anaplasma marginale, which is transmitted by ticks and fomites. A. centrale is a less virulent subspecies used as a live vaccine in cohorts of 8- to 10-mo-old calves that did not naturally reach enzootic stability. We developed 3 variants of a double-antigen sandwich ELISA (dasELISA) using a recombinant major surface protein 5 (MSP5) from A. marginale (dasELISAm) or from A. centrale (dasELISAc) or using MSP5 from both organisms (dasELISAmc). Each dasELISA was tested for the detection of antibodies against A. marginale and A. centrale. The tests were validated using serum samples from cattle not infected with Anaplasma spp. ( n = 388), infected with A. marginale ( n = 436), and vaccinated with A. centrale ( n = 358), confirmed by nested PCR. A total of 462 samples were compared with a commercial competitive ELISA (cELISA). For dasELISAm, dasELISAc, and dasELISAmc, specificities were 98.7%, 98.7%, and 97.4%, and overall sensitivities were 92.6%, 85.7%, and 97.4%, respectively. For A. marginale–infected and A. centrale–vaccinated cattle, sensitivities were 97.7% and 86.3% for dasELISAm, and 77.7% and 95.5% for dasELISAc, respectively. Sensitivity of dasELISAmc was similar for both groups (>96%). The agreement rate between dasELISAmc and cELISA was 96.3% (κ = 0.92); the former test allowed earlier detection of seroconversion of vaccinated cattle than did cELISA. Based on these results, the test could be used to 1) determine the enzootic stability or instability of anaplasmosis in calves, 2) conduct epidemiologic studies, and 3) evaluate the immunogenicity of A. centrale live vaccine.

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Serologic Cross-Reactivity between Anaplasma marginale and Anaplasma phagocytophilum

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.12.10.1177-1183.2005 ◽

2005 ◽

Vol 12 (10) ◽

pp. 1177-1183 ◽

Cited By ~ 41

Author(s):

U. M. Dreher ◽

J. de la Fuente ◽

R. Hofmann-Lehmann ◽

M. L. Meli ◽

N. Pusterla ◽

...

Keyword(s):

Animal Species ◽

Surface Protein ◽

Anaplasma Marginale ◽

Cross Reactivity ◽

Enzyme Linked Immunosorbent Assay ◽

Antibody Assay ◽

Serological Tests ◽

Major Surface Protein ◽

New Findings ◽

Major Surface

ABSTRACT In the context of a serosurvey conducted on the Anaplasma marginale prevalence in Swiss cattle, we suspected that a serological cross-reactivity between A. marginale and A. phagocytophilum might exist. In the present study we demonstrate that cattle, sheep and horses experimentally infected with A. phagocytophilum not only develop antibodies to A. phagocytophilum (detected by immunofluorescent-antibody assay) but also to A. marginale (detected by a competitive enzyme-linked immunosorbent assay). Conversely, calves experimentally infected with A. marginale also developed antibodies to A. phagocytophilum using the same serological tests. The identity of 63% determined in silico within a 209-amino-acid sequence of major surface protein 5 of an isolate of A. marginale and one of A. phagocytophilum supported the observed immunological cross-reactivity. These observations have important consequences for the serotesting of both, A. marginale and A. phagocytophilum infection of several animal species. In view of these new findings, tests that have been considered specific for either infection must be interpreted carefully.

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Groupings of highly similar major surface protein (p44)-encoding paralogues: a potential index of genetic diversity amongst isolates of Anaplasma phagocytophilum

Microbiology ◽

10.1099/mic.0.26648-0 ◽

2004 ◽

Vol 150 (3) ◽

pp. 727-734 ◽

Cited By ~ 30

Author(s):

A. N. J. Casey ◽

R. J. Birtles ◽

A. D. Radford ◽

K. J. Bown ◽

N. P. French ◽

...

Keyword(s):

Genetic Diversity ◽

Anaplasma Phagocytophilum ◽

Surface Protein ◽

Major Surface Protein ◽

Potential Index ◽

Major Surface

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Anaplasma marginale Major Surface Protein 2 CD4+-T-Cell Epitopes Are Evenly Distributed in Conserved and Hypervariable Regions (HVR), Whereas Linear B-Cell Epitopes Are Predominantly Located in the HVR

Infection and Immunity ◽

10.1128/iai.72.12.7360-7366.2004 ◽

2004 ◽

Vol 72 (12) ◽

pp. 7360-7366 ◽

Cited By ~ 35

Author(s):

Jeffrey R. Abbott ◽

Guy H. Palmer ◽

Chris J. Howard ◽

Jayne C. Hope ◽

Wendy C. Brown

Keyword(s):

T Cell ◽

B Cell ◽

Surface Protein ◽

Anaplasma Marginale ◽

T Cell Epitopes ◽

B Cell Epitopes ◽

Major Surface Protein ◽

Hypervariable Regions ◽

Major Surface ◽

Linear B

ABSTRACT Organisms in the genus Anaplasma express an immunodominant major surface protein 2 (MSP2), composed of a central hypervariable region (HVR) flanked by highly conserved regions. Throughout Anaplasma marginale infection, recombination results in the sequential appearance of novel MSP2 variants and subsequent control of rickettsemia by the immune response, leading to persistent infection. To determine whether immune evasion and selection for variant organisms is associated with a predominant response against HVR epitopes, T-cell and linear B-cell epitopes were localized by measuring peripheral blood gamma interferon-secreting cells, proliferation, and antibody binding to 27 overlapping peptides spanning MSP2 in 16 cattle. Similar numbers of MSP2-specific CD4+ T-cell epitopes eliciting responses of similar magnitude were found in conserved and hypervariable regions. T-cell epitope clusters recognized by the majority of animals were identified in the HVR (amino acids [aa] 171 to 229) and conserved regions (aa 101 to 170 and 272 to 361). In contrast, linear B-cell epitopes were concentrated in the HVR, residing within hydrophilic sequences. The pattern of recognition of epitope clusters by T cells and of HVR epitopes by B cells is consistent with the influence of protein structure on epitope recognition.

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Draft Genome Sequences ofAnaplasma phagocytophilum,A. marginale, andA. ovisIsolates from Different Hosts

Genome Announcements ◽

10.1128/genomea.01503-17 ◽

2018 ◽

Vol 6 (5) ◽

Cited By ~ 3

Author(s):

Sandra Diaz-Sanchez ◽

Angélica Hernández-Jarguín ◽

Isabel G. Fernández de Mera ◽

Pilar Alberdi ◽

Erich Zweygarth ◽

...

Keyword(s):

United States ◽

Genome Sequence ◽

Anaplasma Phagocytophilum ◽

Draft Genome ◽

Anaplasma Marginale ◽

The United States ◽

Genome Sequences ◽

Anaplasma Ovis ◽

First Report

ABSTRACTHere, we report the draft genome sequences of isolates ofAnaplasma phagocytophilum,Anaplasma marginale, andAnaplasma ovis. The genomes ofA. phagocytophilum(human),A. marginale(cattle), andA. ovis(goat) isolates from the United States were sequenced and characterized. This is the first report of anA. ovisgenome sequence.

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