Rapid Detection and Differentiation of Staphylococcus Colonies Using Optical Scattering Technology
Abstract
Staphylococcus species are major human and veterinary pathogens, which are also responsible for foodborne illnesses and nosocomial infections. Staphylococcus aureus is a significant cause of foodborne illnesses, causing an estimated 241,000 illnesses per year in the United States. S. aureus produces enterotoxins, which are heat stable and considered superantigens, and can cause foodborne diseases as soon as it is being ingested. Therefore, advanced technologies and methods are needed to rapidly detect the presence of Staphylococcus species including S. aureus in food samples to minimize the risk of contamination as well as the number of foodborne outbreaks and illnesses. We investigated if the laser optical sensor, BARDOT (bacterial rapid detection using optical scattering technology) could be used for rapid screening and detection of Staphylococcus at the genus level on agar plate and differentiate them from non- Staphylococcus spp. The data presented in this study demonstrate that, BARDOT could be used for real-time detection and identification of Staphylococcus colonies on agar plate. Among the different media, Phenol Red Mannitol Agar (PRMA), Mannitol Salt Agar (MSA), modified Tellurite Glycine Agar (mTGA) and modified Vogel and Johnson Agar (mVGA) media were tested to capture scatter patterns, PRMA revealed very high classification accuracy (87-100% accuracy) for identification of five different Staphylococcus species. Staphylococcus and non- Staphylococcus scatter patterns were profoundly different on the PRM agar. The Staphylococcus scatter image library was successfully used for the screening and detection of natural Staphylococcus isolates from chicken salad and bovine raw milk samples. A positive predictive value of 80% or higher was used as the threshold value to consider an isolate-scatter pattern to be positive. Three chicken salad samples and a bovine raw milk sample were processed to screen for Staphylococcus isolates. One of three chicken salad sample and the raw milk sample were positive for Staphylococcus species. A total of 22 natural isolates were further confirmed by tuf gene-specific PCR, 16S rRNA gene sequencing (1245-1409 bp) and analytical profile index (Staph-API™)-based biochemical tests. When comparing isolates with 16S rRNA gene sequencing, BARDOT yielded 4.5% false-negative and 13.6% false –positive identification rate. This novel, label-free, non-invasive on-plate colony screening technology has the potential for adaptation by the food industry, biotechnology companies and public health laboratories in the future for screening of Staphylococcus from various samples
Degree
M.S.
Advisors
Bhunia, Purdue University.
Subject Area
Food Science
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