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Richelle C Charles,
Alaullah Sheikh,
Bryan Krastins,
Jason B Harris, M Saruar Bhuiyan,
Regina C LaRocque,
Tanya Logvinenko,
David A Sarracino,
Indira T Kudva,
Jana Eisenstein,
Michael J Podolsky,
Anuj Kalsy,
W Abdullah Brooks,
Albrecht Ludwig,
Manohar John,
Stephen B Calderwood,
Firdausi Qadri,
Edward T Ryan
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ABSTRACT: Salmonella enterica serotype Typhi is the cause of typhoid fever and a human-restricted pathogen. Currently available typhoid vaccines provide 50 to 90% protection for 2 to 5 years, and available practical diagnostic assays to identify individuals with typhoid fever lack sensitivity and/or specificity. Identifying immunogenic S. Typhi antigens expressed during human infection could lead to improved diagnostic assays and vaccines. Here we describe a platform immunoaffinity proteomics-based technology (IPT) that involves the use of columns charged with IgG, IgM, or IgA antibody fractions recovered from humans bacteremic with S. Typhi to capture S. Typhi proteins that were subsequently identified by mass spectrometry. This screening tool identifies immunogenic proteins recognized by antibodies from infected hosts. Using this technology and the plasma of patients with S. Typhi bacteremia in Bangladesh, we identified 57 proteins of S. Typhi, including proteins known to be immunogenic (PagC, HlyE, OmpA, and GroEL) and a number of proteins present in the human-restricted serotypes S. Typhi and S. Paratyphi A but rarely found in broader-host-range Salmonella spp. (HlyE, CdtB, PltA, and STY1364). We categorized identified proteins into a number of major groupings, including those involved in energy metabolism, protein synthesis, iron homeostasis, and biosynthetic and metabolic functions and those predicted to localize to the outer membrane. We assessed systemic and mucosal anti-HlyE responses in S. Typhi-infected patients and detected anti-HlyE responses at the time of clinical presentation in patients but not in controls. These findings could assist in the development of improved diagnostic assays.
Clinical and vaccine immunology: CVI 08/2010; 17(8):1188-95. · 2.37 Impact Factor
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Alaullah Sheikh, M Saruar Bhuiyan,
Farhana Khanam,
Fahima Chowdhury,
Amit Saha,
Dilruba Ahmed,
K M A Jamil,
Regina C LaRocque,
Jason B Harris,
Mian Mashhud Ahmad,
Richelle Charles,
W Abdullah Brooks,
Stephen B Calderwood,
Alejandro Cravioto,
Edward T Ryan,
Firdausi Qadri
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ABSTRACT: Many currently available diagnostic tests for typhoid fever lack sensitivity and/or specificity, especially in areas of the world where the disease is endemic. In order to identify a diagnostic test that better correlates with typhoid fever, we evaluated immune responses to Salmonella enterica serovar Typhi (serovar Typhi) in individuals with suspected typhoid fever in Dhaka, Bangladesh. We enrolled 112 individuals with suspected typhoid fever, cultured day 0 blood for serovar Typhi organisms, and performed Widal assays on days 0, 5, and 20. We harvested peripheral blood lymphocytes and analyzed antibody levels in supernatants collected on days 0, 5, and 20 (using an antibody-in-lymphocyte-supernatant [ALS] assay), as well as in plasma on these days. We measured ALS reactivity to a serovar Typhi membrane preparation (MP), a formalin-inactivated whole-cell preparation, and serovar Typhi lipopolysaccharide. We measured responses in healthy Bangladeshi, as well as in Bangladeshi febrile patients with confirmed dengue fever or leptospirosis. We categorized suspected typhoid fever individuals into different groups (groups I to V) based on blood culture results, Widal titer, and clinical features. Responses to MP antigen in the immunoglobulin A isotype were detectable at the time of presentation in the plasma of 81% of patients. The ALS assay, however, tested positive in all patients with documented or highly suspicious typhoid, suggesting that such a response could be the basis of improved diagnostic point-of-care-assay for serovar Typhi infection. It can be important for use in epidemiological studies, as well as in difficult cases involving fevers of unknown origin.
Clinical and vaccine immunology: CVI 10/2009; 16(11):1587-94. · 2.37 Impact Factor
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Aaron M Harris, M Saruar Bhuiyan,
Fahima Chowdhury,
Ashraful I Khan,
Azim Hossain,
Emily A Kendall,
Atiqur Rahman,
Regina C LaRocque,
Jens Wrammert,
Edward T Ryan,
Firdausi Qadri,
Stephen B Calderwood,
Jason B Harris
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ABSTRACT: Cholera, caused by Vibrio cholerae, is a noninvasive dehydrating enteric disease with a high mortality rate if untreated. Infection with V. cholerae elicits long-term protection against subsequent disease in countries where the disease is endemic. Although the mechanism of this protective immunity is unknown, it has been hypothesized that a protective mucosal response to V. cholerae infection may be mediated by anamnestic responses of memory B cells in the gut-associated lymphoid tissue. To characterize memory B-cell responses to cholera, we enrolled a cohort of 39 hospitalized patients with culture-confirmed cholera and evaluated their immunologic responses at frequent intervals over the subsequent 1 year. Memory B cells to cholera antigens, including lipopolysaccharide (LPS), and the protein antigens cholera toxin B subunit (CTB) and toxin-coregulated pilus major subunit A (TcpA) were enumerated using a method of polyclonal stimulation of peripheral blood mononuclear cells followed by a standard enzyme-linked immunospot procedure. All patients demonstrated CTB, TcpA, and LPS-specific immunoglobulin G (IgG)and IgA memory responses by day 90. In addition, these memory B-cell responses persisted up to 1 year, substantially longer than other traditional immunologic markers of infection with V. cholerae. While the magnitude of the LPS-specific IgG memory B-cell response waned at 1 year, CTB- and TcpA-specific IgG memory B cells remained significantly elevated at 1 year after infection, suggesting that T-cell help may result in a more durable memory B-cell response to V. cholerae protein antigens. Such memory B cells could mediate anamnestic responses on reexposure to V. cholerae.
Infection and immunity 07/2009; 77(9):3850-6. · 4.21 Impact Factor
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ABSTRACT: Colonization factor CS6 expressed by enterotoxigenic Escherichia coli (ETEC) is a nonfimbrial polymeric protein. A substantial proportion of ETEC strains isolated from patients in endemic settings and in people who travel to regions where ETEC is endemic are ETEC strains expressing CS6, either alone or in combination with fimbrial colonization factor CS5 or CS4. However, relatively little is known about the natural immune responses elicited against CS6 expressed by ETEC strains causing disease. We studied patients who were hospitalized with diarrhea (n = 46) caused by CS6-expressing ETEC (ETEC expressing CS6 or CS5 plus CS6) and had a disease spectrum ranging from severe dehydration (27%) to moderate or mild dehydration (73%). Using recombinant CS6 antigen, we found that more than 90% of the patients had mucosal immune responses to CS6 expressed as immunoglobulin (IgA) antibody-secreting cells (ASC) or antibody in lymphocyte supernatant (ALS) and that about 57% responded with CS6-specific IgA antibodies in feces. More than 80% of the patients showed IgA seroconversion to CS6. Significant increases in the levels of anti-CS6 antibodies of the IgG isotype were also observed in assays for ASC (75%), ALS (100%), and serum (70%). These studies demonstrated that patients hospitalized with the noninvasive enteric pathogen CS6-expressing ETEC responded with both mucosal and systemic antibodies against CS6. Studies are needed to determine if the anti-CS6 responses protect against reinfection and if protective levels of CS6 immunity are induced by vaccination.
Infection and Immunity 06/2007; 75(5):2269-74. · 4.16 Impact Factor
