Detection of viable and dead Listeria monocytogenes on gouda-like cheeses by real-time PCR.
OBJECTIVE
Surface contamination by Listeria monocytogenes of gouda-like cheeses during processing represents a potential public health problem. The aim of this work was to develop novel real-time PCR diagnostics to detect the presence of viable, dead or viable but not culturable (VBNC) cells on gouda-like cheeses.


RESULTS
We used ethidium monoazide bromide (EMA)-PCR for direct quantification of viable and dead cells, while semiquantitative detection of culturable cells below the PCR detection limit (c. 100 CFU g(-1)) was obtained by combining growth and real-time PCR. We were able to quantify the fraction of >0.5% viable cells in a background of dead cells by EMA-PCR, given that the viable cell concentration was above the PCR detection limit. The combined growth and real-time PCR complemented the EMA-PCR, and enabled semiquantitative detection of low levels of culturable cells (10 and 100 CFU g(-1)).


CONCLUSIONS
The significance of this work is that we have developed a novel concept for detection of viable and potentially infectious L. monocytogenes.
Rudi
K
K
MATFORSK, Norwegian Food Research Institute, Osloveien, As, Norway. knut.rudi@matforsk.no
Naterstad
K
K
Drømtorp
S M
SM
Holo
H
H
D001386:Azides / Q000378:metabolism; D001431:Bacteriological Techniques; D002611:Cheese / Q000382:microbiology*; D005516:Food Microbiology; D008089:Listeria monocytogenes / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013194:Staining and Labeling
D001386:Azides; C014535:8-azidoethidium