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United States Department of Agriculture

Agricultural Research Service

Title: The Use of Time-Resolved Fluoroimmunoassay to Simultaneously Detect Escherichia Coli 0157:h7, Salmonella Typhimurium and Salmonella Enteriditisin Foods

Authors
item Tu, Shu-I
item Golden, Marsha
item Andreotti, Peter - PERKIN ELMER LIFE SCIENCE
item Irwin, Peter

Submitted to: Journal of Rapid Methods and Automation in Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: April 1, 2002
Publication Date: N/A

Interpretive Summary: Contamination of pathogenic bacteria, e.g., E coli O157:H7 and Salmonella in foods may lead to serious public health concerns. To minimize possible outbreak of food poisoning by these bacteria, sensitive and rapid detection techniques are needed to call for proper treatments to intervene further distribution of contaminated foods. In current study, we have developed a anew approach to simultaneously detect 1 cell of bacteria E. coli O157:H7 and/or Salmonella spiked in hamburger, ground pork and ground turkey. After an enrichment for 4.5 hours, cells of E. coli O157:H7 and Salmonella were captured by specific antibodies coated on magnetic beads. Captured E. coli and Salmonella were further tagged by metal europium and samarium, respectively. From the unique fluorescence properties of the two metals, we were able to determine the presence/absence of one or both bacteria in meat samples. With this method, the presence of low levels of E. coli O157:H7 and Salmonella may be simultaneously determined within 8 hours. Furthermore, the approach can meet the high throughput requirement by the use of 96-well micro-plate format. The information is useful for researcher and/or engineers to design an automated process for detecting multiple pathogens in foods.

Technical Abstract: Immunomagnetic beads (IMB) were used to capture and concentrate E. coli O157:H7 Salmonella Typhimurium and Salmonella Enteriditis. The captured bacteria were allowed to form sandwiched complexes with europium (Eu) labeled anti-E. coli O157 antibodies and/or samarium (Sm) labeled anti-Salmonella antibodies. After washing the complexes utilizing a magnetic particle concentrator (MPC), the lanthanide labels were extracted out from the complexes to form strongly fluorescent chelates. The specific time-resolved fluorescence (TRF) associated with Eu and Sm were measured to estimate the quantity of E. coli and Salmonella, respectively. The sensitivity of the method was in the range of one to ten thousands CFU/mL for pure cultures of E. coli O157:H7 and Salmonella Typhimurium. The approach of applying sandwich complexes yielded a minimum cross-reactivity with non-targeted bacteria. After an enrichment for 4.5 h at 37 0C in EC media, the approach was able to detect 1 CFU/g of either E. coli or Salmonella or both spiked in hamburger, turkey or pork. The results demonstrated that low levels of E. coli O157:H7 and Salmonella Typhimurium in foods could be simultaneously detected by the TRF approach within 8 hours.

Last Modified: 4/20/2014
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