Quantification detection of Listeria monocytogenes and Listeria innocua by real-time PCR: assessment of hly, iap, and lin02483 targets and AmpliFluor technology

We developed and assessed real-time PCR (RTi-PCR) assays for the detection and quantification of the food-borne pathogen Listeria monocytogenes and the closely related nonpathogenic species L. innocua. The target genes were hly and iap for L. monocytogenes and lin02483 for L. innocua. The assays wer...

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Bibliographic Details
Published in:Applied and environmental microbiology 2004, Vol.70 (3), p.1366-1377
Main Authors: Rodriguez-Lazaro, D, Hernandez, M, Scortti, M, Esteve, T, Vazquez-Boland, J.A, Pla, M
Format: Article
Language:English
Subjects:
Ampifluor technology
bacterial proteins
DNA primers
fluorescence
food microbiology
food pathogens
genes
genetic polymorphism
hemolysins
hly gene
iap gene
invasion-associated surface protein p60
lin02483 gene
Listeria innocua
Listeria monocytogenes
listeriolysin O
microbial detection
molecular sequence data
nucleotide sequences
polymerase chain reaction
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Summary:We developed and assessed real-time PCR (RTi-PCR) assays for the detection and quantification of the food-borne pathogen Listeria monocytogenes and the closely related nonpathogenic species L. innocua. The target genes were hly and iap for L. monocytogenes and lin02483 for L. innocua. The assays were 100% specific, as determined with 100 Listeria strains and 45 non-Listeria strains, and highly sensitive, with detection limits of one target molecule in 11 to 56% of the reactions with purified DNA and 3 CFU in 56 to 89% of the reactions with bacterial suspensions. Quantification was possible over a 5-log dynamic range, with a limit of 15 target molecules and R2 values of >0.996. There was an excellent correspondence between the predicted and the actual numbers of CFU in the samples (deviations of
ISSN:0099-2240
1098-5336