PCR detection of Listeria monocytogenes: a study of multiple factors affecting sensitivity.
OBJECTIVE
To test, under comparable conditions, several parameters affecting sensitivity of PCR detection in order to establish a PCR procedure suitable for the routine detection of Listeria monocytogenes in food.


RESULTS
Beef samples artificially inoculated were used to determine sensitivity of PCR detection under different parameters. As few as 1 CFU g(-1) were detected by DNA extraction using a DNeasy Tissue Kit (Qiagen GmhH, Hilden, Germany) of 1 ml aliquot and PCR amplification with primers directed to the hlyA gene. This PCR protocol was applied in 60 naturally contaminated foods, comparing two enrichment procedures with the traditional culture method. The highest number of positives was recorded by PCR following a 24-h pre-enrichment step at 30 degrees C and a 24-h enrichment step at 37 degrees C. Afterwards, it was applied in 217 naturally contaminated foods and 56 of them tested positive for L. monocytogenes in which only 17 tested positive using the culture method.


CONCLUSIONS
The PCR procedure described has proved to be a rapid and sensitive method suitable for the routine analysis of different types of food.


CONCLUSIONS
The method proposed for the detection of L. monocytogenes, has been validated in naturally contaminated food and is suitable to implement in the food industry.
Aznar
R
R
Departamento de Microbiología y Ecología, Universitat de València, Valencia, Spain. rosa.aznar@uv.es
Alarcón
B
B
D000818:Animals; D001431:Bacteriological Techniques / Q000379:methods; D002417:Cattle; D015169:Colony Count, Microbial; D004269:DNA, Bacterial / Q000032:analysis; D005516:Food Microbiology; D008089:Listeria monocytogenes / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D008460:Meat / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial