Direct detection of Listeria monocytogenes using paramagnetic bead DNA extraction and enzymatic DNA amplification.
The potential of the polymerase chain reaction (PCR) as a tool for direct detection of Listeria monocytogenes in food and clinical samples was investigated. A specific oligonucleotide, directed against the listeriolysin 0 gene of L. monocytogenes, was coupled to paramagnetic beads and used to isolate listerial DNA from food homogenates and blood. The isolated DNA was amplified with a seminested PCR procedure, which recognized the hly gene. The detection limit for L. monocytogenes in 50 ml of a buffer solution was between 1 and 10 colony forming units (cfu). In food homogenates consisting of 10 g food and 40 ml 0.85% NaCl solution, artificially spiked with an overnight culture of L. monocytogenes, the detection limit was 1-10 cfu. The method was further evaluated by application to naturally contaminated food samples. In 250 microliters whole human blood artificially spiked with L. monocytogenes 10,000 cfu could be detected.
Laboratory of Food Chemistry, University of Berne, Switzerland.
C
C
Niederhauser
J
J
Lüthy
U
U
Candrian
D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis* / Q000235:genetics*; D005516:Food Microbiology; D005784:Gene Amplification; D006801:Humans; D018189:Immunomagnetic Separation; D008089:Listeria monocytogenes / Q000235:genetics* / Q000302:isolation & purification*; D008722:Methods; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial