Real-time FRET PCR assay for Salmonella enterica serotype detection in food.
Salmonella enterica subsp. enterica serotypes are leading etiological agents of food-borne gastroenteritis. Traditional identification is laborious and time intensive. Faster molecular methods may allow early identification in contaminated food products. We developed a real-time, fluorescence resonance energy transfer hybridization probe polymerase chain reaction (PCR) assay for S. enterica serotypes on the basis of the exclusive presence of the apeE gene in Salmonella Typhimurium. Assay sensitivity for 12 S. enterica serotypes was as low as 1.87 x 10(2) genomic equivalents per milliliter. PCR efficiency was 94% and the dynamic range was linear over six orders of magnitude from 10(0) to 10(6) copies. The lower limit of detection for 12 different food matrices was between 1.5 x 10(2) and 1.5 x 10(5) CFU/mL without pre-enrichment. When combined with high-throughput automated DNA extraction, 32 food specimens were processed and assayed in less than 2 hours, allowing rapid, specific, sensitive detection of S. enterica serotypes in food products.
David Grant USAF Medical Center, Clinical Investigation Facility, 101 Bodin Circle, Travis Air Force Base, CA 94535, USA.
Eric V
EV
Olsen
Carl S
CS
Gibbins
J Kevin
JK
Grayson
D000818:Animals; D004269:DNA, Bacterial / Q000032:analysis; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005516:Food Microbiology; D008460:Meat / Q000382:microbiology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012478:Salmonella Food Poisoning / Q000175:diagnosis*; D019779:Salmonella enterica / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D012703:Serotyping / Q000379:methods*; D014675:Vegetables / Q000382:microbiology
D004269:DNA, Bacterial