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

Author:

Bhuiyan Taufiqur Rahman1,Hoq Mohammad Rubel1,Nishat Naoshin Sharmin1,Al Mahbuba Deena1,Rashu Rasheduzzaman1,Islam Kamrul1,Hossain Lazina1,Dey Ayan2,Harris Jason B.3,Ryan Edward T.3,Calderwood Stephen B.3,Svennerholm Ann-Mari4,Qadri Firdausi1

Affiliation:

1. Centre for Vaccine Sciences, International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh

2. International Vaccine Institute, Seoul, South Korea

3. Division of Infectious Diseases, Massachusetts General Hospital, Boston, Massachusetts, USA

4. Department of Microbiology and Immunology, University of Gothenburg, Gothenburg, Sweden

Abstract

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.

Funder

Commission of the European Communities grant to STOPENTERICS

Fogarty International Centre

Swedish Collaborative grant SIDA

Swedish SIDA

HHS | National Institutes of Health

Publisher

American Society for Microbiology

Subject

Microbiology (medical),Clinical Biochemistry,Immunology,Immunology and Allergy

Reference48 articles.

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2. The global burden of diarrhoeal disease, as estimated from studies published between 1992 and 2000;Kosek M;Bull World Health Organ,2003

3. Colonization factors of human enterotoxigenic Escherichia coli (ETEC)

4. Modern vaccines. Enteric infections;Levine MM;Lancet,1990

5. Oral vaccines against cholera and enterotoxigenic Escherichia coli diarrhea;Svennerholm AM;Adv Exp Med Biol,1995

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