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Secretomes of bacterial foodborne pathogens and functional assessment during interaction with intestinal epithelium

Reference	BB/C508485/1
Principal Investigator / Supervisor	Professor David Smith
Co-Investigators / Co-Supervisors	Professor Willie Donachie, Dr Paul Everest, Professor Mark Roberts
Institution	University of Glasgow
Department	Veterinary Pathology
Funding type	Research
Value (£)	268,095
Status	Completed
Type	Research Grant
Start date	01/05/2005
End date	30/04/2008
Duration	36 months

Abstract

This investigation is targetted towards improving the understanding of the fundamental basis of the differential pathogenicity of the main foodborne pathogens in livestock and human hosts, i.e. Campylobacter, Salmonella and E. coli O157:H7. We will take a novel experimental approach to test the effect of biological signals relevant to the intestinal environment on secreted bacterial determinants (secretome). To achieve this, intestinal epithelial cells of bovine and human origins will be cultured in vitro to produce preconditioned medium. This will necessitate culture of primary intestinal epithelial cells to produce relevant materials - this is a unique resource not previously utilised in studies of protein expression by these (or other) organisms. We have gained considerable experience in primary culture procedures over the past 4 years. Bacterial strains will be cultured in preconditioned and control media under standardised conditions and secretomes will be examined by SELDI-MS and 2d electrophoresis in combination with MALDI-MS to characterise secreted proteins. DIGE will be employed on selected samples for direct analysis of differential protein complement reciprocal studies of the function of selected bacterial secreted determinants identified through these procedures will be carried out using global (proteomic) and directed (signal transduction) procedures. Protein phosphorylation is a central process in the control of cellular function hence this will form the main focus for functional studies. Phosphoproteome will be characterised using a combination of SELDI-MS, 2d electrophoresis and MALDI-MS. Signalling cascades will be further defined by directed approaches including immunochemical assays, kinase assays and EMSA. Certain phenotypic aspects may be pursued subject to progress in other aspects of study. This investigation will therefore combine aspects of proteomics and cell biology to define aspects of both bacterial and host biology and assess functional consequence of selected determinants on intestinal epithelial cell function.

Summary

unavailable

Committee	Closed Committee - Agri-food (AF)
Research Topics	Animal Health, Microbial Food Safety, Microbiology
Research Priority	X – Research Priority information not available
Research Initiative	Proteomics and Cell Function (PCF) [2003-2004]
Funding Scheme	X – not Funded via a specific Funding Scheme

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