A new 7-plex PCR assay for simultaneous detection of shiga toxin-producing Escherichia coli O157 and Salmonella Enteritidis in meat products

Abstract

A 7-plex PCR assay was developed to achieve an effective detection and identification of serotype Enteritidis of Salmonella spp. and shiga toxin-producing type of Escherichia coli O157 in meat products. Six DNA sequences in the invA and sdfI genes of Salmonella Enteritidis as well as rfbE, eae, stx1, and stx2 genes of E. coli O157:H7 were employed to design primers. The multiplex PCR assay could specifically and sensitively detect and identify target pathogens. Applying the assay to meat samples, the multiplex PCR assay was able to simultaneously detect and identify the two pathogens at a sensitivity of three CFU/10 g raw meats after simple 16 h enrichment in buffered peptone water. Further applying in 21 retail meat samples revealed that two samples were positive for non-shiga toxin producing E. coli O157, one sample was positive for Stx2 producing E. coli O157 and five samples were positive for Salmonella enterica Enteritidis. Taken together, the 7-plex PCR assay is a rapid and reliable method for effectively screening single or multiple pathogens occurrences in various meat products, and could also identify the Salmonella enterica Enteritidis from all Salmonella spp. and shiga toxin producing type from all E. coli strains. Considering as a non expensive screening tool, the multiplex PCR assay has a great potential in complement for food stuff analysis by conventional microbiological tests.

Evaluation of a multiplex selective enrichment broth SEL for simultaneous detection of injured Salmonella, Escherichia coli O157:H7 and Listeria monocytogenes

“…Additionally, selective enrichment is also necessary to suppress the natural background microorganisms, to improve detection efficiency and to avoid false results. However, previously widely used broths for simultaneous enrichment of multiple pathogens before detection should belong to the nonselective media, such as buffered peptone water (BPW) (Alarcon et al , 2004; Wang and Suo, 2011), No. 17 medium (Kawasaki et al , 2010), universal pre-enrichment broth (UPB) (Bhagwat, 2003), et al .…”

Simultaneous Detection of Four Main Foodborne Pathogens in Ready-to-Eat Food by Using a Simple and Rapid Multiplex PCR (mPCR) Assay

“…Moreover, NB contains beef extract and some of its components, such as fats or muscle proteins, are considered as inhibitors that can affect PCR at multiple steps [48]. Wang and Suo (2011) [49] obtained very good detection results after 16 h of growth in BPW, using meat artificially contaminated by E. coli O157 and Salmonella Enteritidis, without any problematic interference due to the presence of background microbiota. The same observation was made by Alarcon et al (2004) [50] for the simultaneous detection of Salmonella spp., S. aureus and L. monocytogenes from artificially inoculated samples.…”

Detection of Salmonella enteritidis and Salmonella typhimurium in foods using a rapid, multiplex real‐time recombinase polymerase amplification assay

“…In the present study, the gene sdfI was selected for the detection of S. enteritidis due to its extensive application in different molecular methods such as PCR, real‐time PCR, and LAMP (Cardona‐Castro, Sánchez‐Jiménez, Lavalett, Múñoz, & Moreno, 2009; Muñoz, Osorio, Jaime, & Cardona‐Castro, 2010; Wang & Suo, 2011). The target has good specificity, sensitivity and repeatability.…”