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PCR detection of Listeria monocytogenes : a study of multiple factors affecting sensitivity
Author(s) -
Aznar R.,
Alarcón B.
Publication year - 2003
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1046/j.1365-2672.2003.02066.x
Subject(s) - listeria monocytogenes , polymerase chain reaction , biology , dna extraction , contamination , false positive paradox , food microbiology , food science , real time polymerase chain reaction , microbiology and biotechnology , chromatography , bacteria , gene , chemistry , mathematics , genetics , ecology , statistics
Aims: 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. Methods and 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°C and a 24‐h enrichment step at 37°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. Significance and Impact of the Study: 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.