"Foodborne disease is one of the most common causes of morbidity and mortality in the world..."

Multi-pathogen screening and/or confirmation via microarray detection

Investigators: Arun K. Bhunia (Department of Food Science), Mark Morgan (Department of Food Science), B. K. Hahm (Department of Food Science), Viswaprakash Nanduri (Department of Food Science), Shu-I Tu (USDA)

Project Report 2005 - 2006

» Download Project Report 2005 - 2006

Project Rationale

Foodborne disease is one of the most common causes of morbidity and mortality in the world, and more than 200 known diseases are transmitted through food. In the United States there are about 76 million cases each year, of which 325,000 require hospitalization and 5,000 die. The foodborne pathogens of concern are E. coli O157:H7, Salmonella, Listeria monocytogenes, Toxoplasma and Campylobacter. Therefore, detection of these pathogenic bacteria during food processing and storage is crucial for the microbiological safety and prevention of possible outbreaks.

Antibody-based detection methods are regarded as rapid and efficient and are widely used in conventional ELISA and dipstick methods. In recent years, antibodies have been successfully used in biosensor tools for rapid detection. Therefore, specificity and avidity of a given antibody for the target bacteria is extremely important, specifically those originating from stressful environments of food. We have also learned that stress can affect antigen expression on bacterial cells thus affecting antibody-based detection.

The goals of this project were to: (a) develop specific antibodies for Listeria monocytogenes, Salmonella enterica and E. coli O157:H7, (b) analyze effect of environmental and physiological stresses on antigen expression and antibody based detection, and (c) to develop antibody-based microarray system for simultaneous detection of multi-pathogens.

Once completed, this would allow development of a detection kit for multiple pathogens, thus saving time and money for product testing and also helping regulatory agencies for evaluation of a product for key food pathogens.

Project Objectives

  • Develop antibody specific for L. monocytogenes, Salmonella enterica and E. coli O157:H7.
  • Determine the effect of environmental and physiological stresses on antigen expression.
  • Develop sandwich ELISA for each pathogen.
  • Determine the selective enrichment media affecting the antibody-based detection of stress-exposed Listeria monocytogenes.
  • Development of pathogen enrichment and detection device (PEDD).

Project Highlights

Two polyclonal antibodies, PAb Lm404 and LmC369, were demonstrated to be specific for Internalin B (InlB) and actin polymerization protein (ActA) of L. monocytogenes, respectively. These antibodies could be potentially used for specific detection of this bacterium. These antibodies showed differential expression of antigens in different enrichment broths: selective media (like BLEB, UVM and FB) suppressed PAb Lm 404 reactive InlB expression whereas only FB suppressed Lm C369 reactive ActA expression. PAb Lm404 could be used only when bacteria are cultured in non selective media while PAb-Lm C369 could be used in an immunoassay with bacteria directly taken from selective enrichment broth. Surface localization of these two epitopes was confirmed by immuno-electron microscopy.

Annual Report

Investigators

  • Arun Bhunia
  • B. Hahm
  • Mark Morgan
  • Viswaprakash Nanduri
  • Shu-I Tu