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ARS Home » Pacific West Area » Albany, California » Western Regional Research Center » Produce Safety and Microbiology Research » Research » Publications at this Location » Publication #337484

Research Project: Ecology and Detection of Human Pathogens in the Produce Production Continuum

Location: Produce Safety and Microbiology Research

Title: Simultaneous direct detection of Shiga-toxin producing Escherichia coli (STEC) strains by gold nanoparticle optical sensing

item QUINTELA, IRWIN - University Of Maine
item DE LOS REYES, BENILDO - Texas Tech University
item LIN, CHIH-SHENG - National Chaio Tung University
item Wu, Vivian

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 1/23/2017
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: Shiga-toxin producing Escherichia coli (STEC) strains (“Big Six” – O26, O45, O103, O111, O121, O145, and O157) represent significant groups of pathogens responsible for foodborne diseases. The objective of this study was to develop a colorimetric optical sensing assay that can simultaneously detect various STEC strains. Pairs of single-stranded thiol-modified oligonucleotides (30-mer) were designed and immobilized onto gold nanoparticle (AuNPs) to target stx1 (119-bp) and/or stx2 (104-bp) genes of STEC big six and O157 serogroups. Oligonucleotide-functionalized AuNps were used as probes based on the sequence-specific hybridization properties of DNA and stable colorimetric properties of AuNPs. Ground beef and blueberries (50 samples each) were randomly inoculated with STEC strains at <1 log CFU/g. A sample pooling plan with brief enrichment procedures (modified Tryptic Soy Broth, at 42'1oC for 6 hours) was incorporated into the assay to ensure detection of viable STEC in food. When amplified target DNA from post-enriched food samples was hybridized with complementary AuNP-probes, reaction mixtures retained its initial red color following an increased salt concentration (2M). For non-target/non-STEC strains such as Salmonella Typhimurium, a change from red to gray-blue was induced due to aggregation of functionalized AuNPs, providing the basis of direct and simultaneous detection of targets. The detection limit is <1 log CFU/g in food. Twenty out of 50 ground beef and blueberries were tested positive for STEC, which is 100% accuracy. It requires less than one hour to complete after DNA sample preparation. Gel electrophoresis and spectrophotometric data confirmed successful DNA sandwich hybridization and AuNPs aggregation. The optical sensing assay provided a reliable means for direct and simultaneous detection of various STEC strains, utilizing AuNPs as inexpensive materials. The strategy was found to be superior over traditional approaches for detection of STEC in food samples even at low-level contamination.