title: Enumeration of Gut-Homing β7-Positive, Pathogen-Specific Antibody-Secreting Cells in Whole Blood from Enterotoxigenic Escherichia coli- and Vibrio cholerae-Infected Patients, Determined Using an Enzyme-Linked Immunosorbent Spot Assay Technique.
abstract: Vibrio cholerae and enterotoxigenic Escherichia coli (ETEC) are noninvasive mucosal pathogens that cause acute watery diarrhea in people in developing countries. Direct assessment of the mucosal immune responses to these pathogens is problematic. Surrogate markers of local mucosal responses in blood are increasingly being studied to determine the mucosal immune responses after infection. However, the volume of blood available in children and infants has limited this approach. We assessed whether an approach that first isolates β7-positive cells from a small volume of blood would allow measurement of the antigen-specific immune responses in patients with cholera and ETEC infection. β7 is a cell surface marker associated with mucosal homing. We isolated β7-expressing cells from blood on days 2, 7, and 30 and used an enzyme-linked immunosorbent spot (ELISPOT) assay to assess the gut-homing antibody-secreting cells (ASCs) specific to pathogen antigens. Patients with ETEC diarrhea showed a significant increase in toxin-specific gut-homing ASCs at day 7 compared to the levels at days 2 and 30 after onset of illness and to the levels in healthy controls. Similar elevations of responses to the ETEC colonization factors (CFs) CS6 and CFA/I were observed in patients infected with CS6- and CFA/I-positive ETEC strains. Antigen-specific gut-homing ASCs to the B subunit of cholera toxin and cholera-specific lipopolysaccharides (LPS) were also observed on day 7 after the onset of cholera using this approach. This study demonstrates that a simple ELISPOT assay can be used to study the mucosal immunity to specific antigens using a cell-sorting protocol to isolate mucosal homing cells, facilitating measurement of mucosal responses in children following infection or vaccination.
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh.
authors.forename: Taufiqur Rahman
authors.initials: TR
authors.lastname: Bhuiyan
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh.
authors.forename: Mohammad Rubel
authors.initials: MR
authors.lastname: Hoq
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh.
authors.forename: Naoshin Sharmin
authors.initials: NS
authors.lastname: Nishat
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh.
authors.forename: Deena
authors.initials: D
authors.lastname: Al Mahbuba
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh.
authors.forename: Rasheduzzaman
authors.initials: R
authors.lastname: Rashu
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh.
authors.forename: Kamrul
authors.initials: K
authors.lastname: Islam
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh.
authors.forename: Lazina
authors.initials: L
authors.lastname: Hossain
authors.affiliation: International Vaccine Institute, Seoul, South Korea.
authors.forename: Ayan
authors.initials: A
authors.lastname: Dey
authors.affiliation: Division of Infectious Diseases, Massachusetts General Hospital, Boston, Massachusetts, USA.
authors.forename: Jason B
authors.initials: JB
authors.lastname: Harris
authors.affiliation: Division of Infectious Diseases, Massachusetts General Hospital, Boston, Massachusetts, USA.
authors.forename: Edward T
authors.initials: ET
authors.lastname: Ryan
authors.affiliation: Division of Infectious Diseases, Massachusetts General Hospital, Boston, Massachusetts, USA.
authors.forename: Stephen B
authors.initials: SB
authors.lastname: Calderwood
authors.affiliation: Department of Microbiology and Immunology, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Ann-Mari
authors.initials: AM
authors.lastname: Svennerholm
authors.affiliation: Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh fqadri@icddrb.org.
authors.forename: Firdausi
authors.initials: F
authors.lastname: Qadri
mesh_terms: D000328:Adult; D000921:Antibody-Producing Cells / Q000276:immunology*; D002771:Cholera / Q000276:immunology* / Q000382:microbiology; D054307:Enterotoxigenic Escherichia coli / Q000276:immunology*; D058501:Enzyme-Linked Immunospot Assay / Q000379:methods; D004927:Escherichia coli Infections / Q000276:immunology* / Q000382:microbiology; D005260:Female; D006801:Humans; D018928:Immunity, Mucosal; D007070:Immunoglobulin A; D007074:Immunoglobulin G / Q000097:blood; D039641:Integrin beta Chains / Q000276:immunology*; D007422:Intestines / Q000276:immunology; D008297:Male; D014734:Vibrio cholerae / Q000276:immunology*; D055815:Young Adult
chemical_list: D007070:Immunoglobulin A; D007074:Immunoglobulin G; D039641:Integrin beta Chains; C091368:integrin beta7