Identification of Circulating Bacterial Antigens by
            <i>In Vivo</i>
            Microbial Antigen Discovery

Nuti, Dana E.; Crump, Reva B.; Handayani, Farida Dwi; Chantratita, Narisara; Peacock, Sharon J.; Bowen, Richard A.; Felgner, Philip L.; Davies, D. Huw; Wu, Terry; Lyons, C. Rick; Brett, Paul J.; Burtnick, Mary N.; Kozel, Thomas R.; AuCoin, David P.

doi:10.1128/mbio.00136-11

Cited by 35 publications

(61 citation statements)

References 24 publications

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“…In this study, we used a protein microarray displaying 1,741 different proteins from F. tularensis Schu S4 to profile the antibodies present in Spanish tularemia cases and healthy controls. This expands on our previous studies with this array, in which we used samples from U.S. donors only (25) or from mouse models (27,32).…”

Section: Resultsmentioning

confidence: 74%

Towards Development of Improved Serodiagnostics for Tularemia by Use of Francisella tularensis Proteome Microarrays

et al. 2016

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e Tularemia in humans is caused mainly by two subspecies of the Gram-negative facultative anaerobe Francisella tularensis: F. tularensis subsp. tularensis (type A) and F. tularensis subsp. holarctica (type B). The current serological test for tularemia is based on agglutination of whole organisms, and the reactive antigens are not well understood. Previously, we profiled the antibody responses in type A and B tularemia cases in the United States using a proteome microarray of 1,741 different proteins derived from the type A strain Schu S4. Fifteen dominant antigens able to detect antibodies to both types of infection were identified, although these were not validated in a different immunoassay format. Since type A and B subspecies are closely related, we hypothesized that Schu S4 antigens would also have utility for diagnosing type B tularemia caused by strains from other geographic locations. To test this, we probed the Schu S4 array with sera from 241 type B tularemia cases in Spain. Despite there being no type A strains in Spain, we confirmed the responses against some of the same potential serodiagnostic antigens reported previously, as well as determined the responses against additional potential serodiagnostic antigens. Five potential serodiagnostic antigens were evaluated on immunostrips, and two of these (FTT1696/GroEL and FTT0975/conserved hypothetical protein) discriminated between the Spanish tularemia cases and healthy controls. We conclude that antigens from the type A strain Schu S4 are suitable for detection of antibodies from patients with type B F. tularensis infections and that these can be used for the diagnosis of tularemia in a deployable format, such as the immunostrip.T ularemia is a zoonotic disease caused by the Gram-negative facultative anaerobe Francisella tularensis. The organism infects a multitude of different animals, including mammals, birds, fish, and insects. Transmission to humans can occur by inhalation, although the most common natural route is likely to be direct contact with infected animals or via ticks and other biting arthropods that feed on infected mammals. Important animal reservoirs appear to be rodents, hares, and rabbits. For example, the 1997 outbreak in Castilla y León, Spain (1, 2), was transmitted by the handling of infected hares. The following year, another outbreak which was attributed to crayfish handling occurred in Castilla-La Mancha, Spain (3). Human-tohuman transmission is rare.Four closely related subspecies have been defined: F. tularensis subsp. tularensis (type A), F. tularensis subsp. holarctica (type B), F. tularensis subsp. novicida, and F. tularensis subsp. mediasiatica. Each subspecies possesses a different pathogenic potential in humans, as well as a distinctive geographic distribution, host preference, and route of transmission. Type A is found mainly in North America and is associated with severe tularemia that may be fatal, whereas type B is found throughout the Northern Hemisphere and is less virulent in humans (4-6). F. tularensis subsp. ...

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Section: Resultsmentioning

confidence: 74%

Towards Development of Improved Serodiagnostics for Tularemia by Use of Francisella tularensis Proteome Microarrays

et al. 2016

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“…Our aim was to provide as immunogens those proteins that were in a cell-free state in the blood by virtue of being released, secreted, or, more likely for a Borrelia species, incorporated into membrane vesicles (40,41,47,48). Carbohydrates or glycolipids might have been present in the filtrate and elicited an antibody response in the mice, as was the case for Nuti et al with Burkholderia pseudomallei (15), but our subsequent protocol favored identification of proteins.…”

Section: Discussionmentioning

confidence: 99%

“…In addition, the antibiotic carbenicillin, at a final concentration of 50 g/ml, was incubated with the filtered sera for 30 min prior to injection of naive mice. Immunization of immunocompetent 4-to 6-week-old female wildtype BALB/cByJ mice (Jackson Laboratory, Bar Harbor, ME) was carried out essentially as described previously by Nuti et al (15). Mice in groups of 5 were each injected subcutaneously with a mixture of 50 l of the serum filtrate, 50 l of PBS, and 100 l of TiterMax gold adjuvant (TiterMax USA, Inc.).…”

Section: Methodsmentioning

confidence: 99%

“…For a set of candidates of a manageable number and that might be enriched for outer membrane-associated proteins, we used an approach that was previously used to identify circulating antigens of the bacterial pathogens Burkholderia pseudomallei and Francisella tularensis in the serum of infected mice (15), namely, by immunizing immunocompetent mice with a cell-free filtrate of blood from immunodeficient mice bacteremic with B. hermsii. By this method, we identified a protein, BHA007, that is encoded by the large linear plasmid of this species and that has orthologs among other RF species but not Lyme disease Borrelia spp.…”

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Fibronectin-Binding Protein of Borrelia hermsii Expressed in the Blood of Mice with Relapsing Fever

et al. 2014

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To identify and characterize surface proteins expressed by the relapsing fever (RF) agent Borrelia hermsii in the blood of infected mice, we used a cell-free filtrate of their blood to immunize congenic naive mice. The resultant antiserum was used for Western blotting of cell lysates, and gel slices corresponding to reactive bands were subjected to liquid chromatography-tandem mass spectrometry, followed by a search of the proteome database with the peptides. One of the immunogens was identified as the BHA007 protein, which is encoded by a 174-kb linear plasmid. BHA007 had sequence features of lipoproteins, was surface exposed by the criteria of in situ protease susceptibility and agglutination of Vtp ؊ cells by anti-BHA007 antibodies, and was not essential for in vitro growth. BHA007 elicited antibodies during experimental infection of mice, but immunization with recombinant protein did not confer protection against needle-delivered infection. Open reading frames (ORFs) orthologous to BHA007 were found on large plasmids of other RF species, including the coding sequences for the CihC proteins of Borrelia duttonii and B. recurrentis, but not in Lyme disease Borrelia species. Recombinant BHA007 bound both human and bovine fibronectin with K d (dissociation constant) values of 22 and 33 nM, respectively, and bound to C4-binding protein with less affinity. The distant homology of BHA007 and its orthologs to BBK32 proteins of Lyme disease species, as well as to previously described BBK32-like proteins in relapsing fever species, indicates that BHA007 is a member of a large family of multifunctional proteins in Borrelia species that bind to fibronectin as well as other host proteins.

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“…This novel approach was used to construct protein microarrays containing pathogen proteomes, which were later used to analyze the antibody response in serum samples from infected mice and humans. These studies led to the identification of immunodominant antigens of diagnostic value (16,17 ) and also identified antibodies associated with protective immunity in response to vaccination (18).…”

Section: Whole Proteome Expression In Vitromentioning

confidence: 99%

Antigen Microarrays for the Study of Autoimmune Diseases

Yeste

Quintana

2013

Clinical Chemistry

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BACKGROUND The immune response involves the activation of heterogeneous populations of T cells and B cells that show different degrees of affinity and specificity for target antigens. Although several techniques have been developed to study the molecular pathways that control immunity, there is a need for high-throughput assays to monitor the specificity of the immune response. CONTENT Antigen microarrays provide a new tool to study the immune response. We reviewed the literature on antigen microarrays and their advantages and limitations, and we evaluated their use for the study of autoimmune diseases. Antigen arrays have been successfully used for several purposes in the investigation of autoimmune disorders: for disease diagnosis, to monitor disease progression and response to therapy, to discover mechanisms of pathogenesis, and to tailor antigen-specific therapies to the autoimmune response of individual patients. In this review we discuss the use of antigen microarrays for the study of 4 common autoimmune diseases and their animal models: type 1 diabetes, systemic lupus erythematosus, rheumatoid arthritis, and multiple sclerosis. CONCLUSIONS Antigen microarrays constitute a new tool for the investigation of the immune response in autoimmune disorders and also in other conditions such as tumors and allergies. Once current limitations are overcome, antigen microarrays have the potential to revolutionize the investigation and management of autoimmune diseases.

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Identification of Circulating Bacterial Antigens by In Vivo Microbial Antigen Discovery

Cited by 35 publications

References 24 publications

Towards Development of Improved Serodiagnostics for Tularemia by Use of Francisella tularensis Proteome Microarrays

Towards Development of Improved Serodiagnostics for Tularemia by Use of Francisella tularensis Proteome Microarrays

Fibronectin-Binding Protein of Borrelia hermsii Expressed in the Blood of Mice with Relapsing Fever

Antigen Microarrays for the Study of Autoimmune Diseases

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