ArticleDiarrheal diseases

Centre international de recherche sur les maladies diarrhéiques de Bangladesh (icddr,b)

Enumeration of gut-homing β7-positive, pathogen-specific antibody secreting cells in whole blood in enterotoxigenic Escherichia coli and Vibrio cholerae-infected patients using an ELISPOT technique

Bhuiyan TR, Hoq MR, Nishat NS, Al Mahbuba D, Rashu R, Islam K, Hossain L, Dey A, Harris JB, Ryan ET, Calderwood SB, Svennerholm AM, Qadri F.

Clin Vaccine Immunol. 2015 Oct 28. pii: CVI.00526-15


Vibrio cholerae and enterotoxigenic Escherichia coli (ETEC) are noninvasive mucosal pathogens that cause acute watery diarrhea in developing countries. Directly assessing mucosal immune responses to these pathogens is problematic. Surrogate markers of local mucosal responses using blood are increasingly being studied to determine mucosal immune responses after infection. However, volume of blood available in children and infants has limited this approach. We assessed whether an approach first that isolates β7-positive cells from a small volume of blood would allow measurement of 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 ELISPOT assay to assess 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 days 2 and 30 after onset of illness, and in comparison to healthy controls. A similar elevation of responses to the ETEC colonization factors 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 LPS were also observed on day 7 after onset of cholera using this approach. This study demonstrates that a simple ELISPOT can be used to study 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.

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