Rapid detection of enterotoxigenic clostridium perfringens by real-time fluorescence resonance energy transfer PCR.
Clostridium perfringens is one of the etiologic agents of gas gangrene that can occur when a wound is contaminated with soil. Type A C. perfringens can cause foodborne and nonfoodborne gastrointestinal illnesses due to an enterotoxin (CPE) produced by some strains during sporulation. We developed a quantitative real-time PCR assay based on fluorescence resonance energy transfer hybridization chemistry that targets the C. perfringens-specific phospholipase C (plc) gene and the enterotoxigenic gene (cpe) with the LightCycler and the Ruggedized Advanced Pathogen Identification Device (R.A.P.I.D.). The assay can detect as few as 20 copies of target sequences per PCR. The total assay time, from extraction to PCR analysis, is 90 min. This assay is rapid, sensitive, and specific and will allow direct detection of C. perfringens in water, food, and stool samples. It should prove helpful in investigating foodborne illnesses due to C. perfringens and can be used as a tool to ensure the safety of food and water supplies.
dela Cruz
Wilfred P
WP
Molecular Biology Research Element, Clinical Investigation Facility, David Grant USAF Medical Center, Travis Air Force Base, California 94535, USA. wilfred.delacruz@travis.af.mil
Gozum
Mary M A
MM
Lineberry
Sarah F
SF
Stassen
Sarah D
SD
Daughtry
Marianne
M
Stassen
Nicholas A
NA
Jones
Morris S
MS
Johnson
Oswald L
OL
D001483:Base Sequence; D003016:Clostridium perfringens / Q000145:classification / Q000302:isolation & purification*; D004269:DNA, Bacterial / Q000032:analysis*; D004768:Enterotoxins / Q000032:analysis / Q000235:genetics; D005453:Fluorescence; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013171:Spores, Bacterial; D013997:Time Factors; D014871:Water Microbiology
D004269:DNA, Bacterial; D004768:Enterotoxins; C035598:enterotoxin, Clostridium