Comparison of Phage-Based Magnetoelastic Biosensors with Taqman- Based Quantitative Real-Time PCR for the Detection of Salmonella typhimurium Directly Grown on Tomato Surfaces

A phage-based magnetoelastic (ME) biosensor method was compared with a TaqMan-based quantitative real- time PCR (Q-PCR) method for the detection of Salmonella typhimurium on tomato surfaces. This ME biosensor method utilizes magnetoelastic resonators coated with E2 filamentous phage to bind with and measure the concentration of S. typhimurium . In this study, standard curves, correlations, and limits of detection (LOD) for the ME biosensor and Q-PCR methods were determined by inoculating tomato surfaces with S. typhimurium suspensions in concentrations ranging from 1 to 8 log CFU/tomato. The LOD for the ME biosensor method and Q-PCR were 3 and 2 log CFU/tomato, respectively. In a direct comparison of the detection methods, S. typhimurium suspensions (3 log CFU/tomato) were inoculated on 65 tomato surfaces, then incubated at 37°C and 100% RH for 24 h. After 24 h, S. typhimurium was positively detected by both methods and the quantified concentrations were nearly the same, (6.35 ± 2.03) and (6.34 ± 0.17) log CFU/tomato respectively for the ME biosensor method and the Q-PCR method, which were significantly greater than the concentration determined by the BGS-plate count method (5.33 ± 0.21). Scanning electron microscopy (SEM) was used to confirm the growth of S. typhimurium on the tomato surfaces and the binding of S. typhimurium on the measurement sensors. This study demonstrated that the ME biosensor method was robust and competitive with Q-PCR for S. typhimurium detection on fresh produce.

[1]  S. Garrett,et al.  Detection of meat species using TaqMan real-time PCR assays. , 2004, Meat science.

[2]  R S Lakshmanan,et al.  Rapid and sensitive magnetoelastic biosensors for the detection of Salmonella typhimurium in a mixed microbial population. , 2007, Journal of microbiological methods.

[3]  R. Brackett,et al.  Survival of Salmonellae on and in Tomato Plants from the Time of Inoculation at Flowering and Early Stages of Fruit Development through Fruit Ripening , 2001, Applied and Environmental Microbiology.

[4]  B. Bisha,et al.  Simple Adhesive-Tape-Based Sampling of Tomato Surfaces Combined with Rapid Fluorescence In Situ Hybridization for Salmonella Detection , 2009, Applied and Environmental Microbiology.

[5]  Shin Horikawa,et al.  Direct detection of Salmonella typhimurium on fresh produce using phage-based magnetoelastic biosensors. , 2010, Biosensors & bioelectronics.

[6]  Knut Rudi,et al.  Use of Ethidium Monoazide and PCR in Combination for Quantification of Viable and Dead Cells in Complex Samples , 2005, Applied and Environmental Microbiology.

[7]  V A Petrenko,et al.  Sequential detection of Salmonella typhimurium and Bacillus anthracis spores using magnetoelastic biosensors. , 2009, Biosensors & bioelectronics.

[8]  P. Elizaquível,et al.  A multiplex RTi-PCR reaction for simultaneous detection of Escherichia coli O157:H7, Salmonella spp. and Staphylococcus aureus on fresh, minimally processed vegetables. , 2008, Food microbiology.

[9]  Rhétorique Lettres,et al.  Food and Bioprocess Technology , 2011 .

[10]  R. Brackett,et al.  PCR Detection of Salmonella entericaSerotype Montevideo in and on Raw Tomatoes Using Primers Derived from hilA , 2000, Applied and Environmental Microbiology.

[11]  J. Je,et al.  Quantitative Detection of Salmonella typhimurium Contamination in Milk, Using Real-Time PCR , 2005 .

[12]  M. Uyttendaele,et al.  Evaluation of real‐time PCR vs automated ELISA and a conventional culture method using a semi‐solid medium for detection of Salmonella , 2003, Letters in applied microbiology.

[13]  V. Petrenko,et al.  Thermostability of landscape phage probes , 2005, Analytical and bioanalytical chemistry.

[14]  A. Bhagwat,et al.  Application of a molecular beacon-real-time PCR technology to detect Salmonella species contaminating fruits and vegetables. , 2004, International journal of food microbiology.

[15]  Bryan A. Chin,et al.  Phage coated magnetoelastic micro-biosensors for real-time detection of Bacillus anthracis spores , 2009 .

[16]  A. Gulluce,et al.  Identification of meat species by TaqMan-based real-time PCR assay. , 2009, Meat science.

[17]  Submitted November,et al.  Division of Dockets Management (Docket No. FDA-2009-D-0348) Comments on "Draft Guidance for Industry: Guide to Minimize Microbial Food Safety Hazards of Leafy Greens" , 2009 .

[18]  T. Michailides,et al.  Approaches for eliminating PCR inhibitors and designing PCR primers for the detection of phytopathogenic fungi , 2007 .

[19]  Burkhard Malorny,et al.  Comparison of PCR-ELISA and LightCycler real-time PCR assays for detecting Salmonella spp. in milk and meat samples. , 2004, Molecular and cellular probes.

[20]  R. Heckert,et al.  The effect of pre-enrichment protocol on the sensitivity and specificity of PCR for detection of naturally contaminated Salmonella in raw poultry compared to conventional culture. , 2006, Food microbiology.

[21]  I-Hsuan Chen,et al.  Landscape phage probes for Salmonella typhimurium. , 2005, Journal of microbiological methods.

[22]  L. Heller,et al.  Comparison of Methods for DNA Isolation from Food Samples for Detection of Shiga Toxin-Producing Escherichia coli by Real-Time PCR , 2003, Applied and Environmental Microbiology.

[23]  H. J. Fels-Klerx,et al.  Suitability of Rapid Detection Methods for Salmonella in Poultry Slaughterhouses , 2009 .

[24]  C. Wei,et al.  Growth and Survival of Salmonella montevideo on Tomatoes and Disinfection with Chlorinated Water. , 1995, Journal of food protection.

[25]  Eun-Gyeong Shin,et al.  Direct and quantitative analysis of Salmonella enterica serovar Typhimurium using real-time PCR from artificially contaminated chicken meat. , 2008, Journal of microbiology and biotechnology.

[26]  Bryan A. Chin,et al.  Detection of Salmonella typhimurium in fat free milk using a phage immobilized magnetoelastic sensor , 2007 .