POLYMERASE CHAIN-REACTION ASSAY COUPLED WITH FLUORESCENCE DETECTION ON MICROWELL PLATES FOR LISTERIA-MONOCYTOGENES IN FOODS

This study evaluates a polymerase chain-reaction assay coupled with fl uorescence detection (PCR-FD) in microwell plates for Listeria monocyt ogenes in dairy and food samples. Guanidinium thiocyanate/silica-extra cted cultures and milk and dairy-food samples were used as templates f or a PCR assay in microwell plates, with a primer pair that amplifies a 221 bp segment of the internal transcribed spacer (ITS), in the pres ence of 0.5 mu g/ml of ethidium bromide. Fluorescence of the PCR produ cts was measured with a CytoFluor 2300 fluorescence measurement system (Millipore Corporation, Bedford, MA). The lowest level of detection o f the assay was 10 to 100 CFU. A total of 326 food samples were tested , both by culture and by PCR-FD, The overall sensitivity of the PCR-FD assay was 95.2% and the specificity was 89.9%. Positive and negative predictive values were 74.8% and 99.4%, respectively. Based on the res ults obtained in this study it appears that the PCR-FD assay described here can be useful for screening a large number of milk and dairy-foo d samples for contamination by Listeria monocytogenes.