Improved Detection Techniques for Foodborne Pathogens: Separation Techniques

Investigator: Michael Ladisch (Department of Agricultural and Biological Engineering)

Project Report 2010 - 2011

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Project Rationale

Microbial testing plays an important role in assuring food safety. Whether applying tests at production, processing, or retail/consumer level, identification of pathogens at the point of contamination is critical for food producers to assure the safety of their products. There are numerous methods for detecting or identifying food pathogens once food extracts are processed into relatively clean solutions. However, the challenge for detection is to amplify the pathogens, if present, in extracts of food matrices so that they may be readily detected and identified. The extracts that must be probed for pathogens are chemically, biologically, and physically complex, and if pathogens are present, they are typically found in relatively low numbers.

Preparation of foodborne pathogen samples typically requires a two-step approach, with primary (nonselective) enrichment followed by secondary (selective) enrichment. However, this approach is less than ideal as injured cells are often not detected due to overgrowth by background flora during nonselective enrichment and/or inhibition of recovery of injured cells during selective enrichment. Selective agents in enrichment media also may inhibit repair of sublethally injured cells of L. monocytogenes or other pathogens.

Despite the development of rapid detection methods (such as ELISA and PCR), reduction and/or elimination of culture enrichment remains an essential goal to enable truly real-time detection methods. While detection may be measured in hours, the enrichment steps leading up to detection add a day or more. The separation techniques developed as part of this research will reduce the time necessary to separate and concentrate pathogenic microorganisms from food matrices and systems, thereby reducing the time required to detect pathogens from foods.

Project Objectives

  • Identify and characterize membranes that will concentrate and recover microorganisms from food samples with minimal adsorption and blockage.
  • Demonstrate adsorption of microorganisms from food matrix, and establish procedures for separating particles from food.
  • Combine fundamental separation science and technology to integrate with detection and quantification systems.

Project Highlights

We developed a unique combination of microfluidic control, first-alert detection, and processing of liquid samples from food matrices to obtain interrogation-ready samples. Our basic approach of forcing fluids through a microfiltration membrane was not new, as there are numerous membranes available for concentrating cells in water. However, existing bioseparation technology was not capable of directly isolating cells from extracts of foods because food components would foul membrane-based separation devices within minutes. Our work has overcome these challenges by combining existing membranes with novel microfluidic engineering to obtain an automatically-controlled system. This enables hands-off operation, with a sequence of automatically controlled steps that reduce fouling of the membrane. Specifically, the filtration system was improved by optimizing filter materials, hollow fiber membrane designs, enzyme treatments before pre-filtration of food-derived samples, and cleaning and sterilization procedures. Results indicate the membrane can be reused up to 20 times, thereby reducing both cost and manual operations associated with recovery of concentrated microbial samples. The filtration device was used successfully to separate and concentrate pathogens from complex foods (e.g., Salmonella from chicken).

""Separation and concentration technologies have been developed that lessen microbial detection times for E. coli, Salmonella, and Listeria by 20 hours.""

Annual Reports