Rapid detection of Listeria monocytogenes in food using culture enrichment combined with real-time PCR.
A rapid method for the detection of Listeria monocytogenes in foods combining culture enrichment and real-time PCR was compared to the ISO 11290-1 standard method. The culture enrichment component of the rapid method is based on the ISO standard and includes 24h incubation in half-Fraser broth, 4h incubation in Fraser broth followed by DNA extraction and real-time PCR detection of the ssrA gene of L. monocytogenes. An internal amplification control, which is co-amplified with the same primers as the L. monocytogenes DNA, was also included in the assay. The method has a limit of detection of 1-5CFU/25g food sample and can be performed in 2 working days compared to up to 7days for the ISO standard. A variety of food samples from retail outlets and food processing plants (n=175) and controls (n=31) were tested using rapid and conventional methods. The rapid method was 99.44% specific, 96.15% sensitive and 99.03% accurate when compared to the standard method. This method has the potential to be used as an alternative to the standard method for food quality assurance providing rapid detection of L. monocytogenes in food.
O'Grady
Justin
J
Molecular Diagnostics Laboratory, Department of Microbiology, National University of Ireland Galway, University Road, Galway, Ireland. justin.ogrady@nuigalway.ie
Ruttledge
Margaret
M
Sedano-Balbás
Sara
S
Smith
Terry J
TJ
Barry
Thomas
T
Maher
Majella
M
D015169:Colony Count, Microbial / Q000379:methods* / Q000592:standards; D003470:Culture Media / Q000737:chemistry; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D005504:Food Analysis / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology; D005784:Gene Amplification; D008089:Listeria monocytogenes / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D012329:RNA, Bacterial / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013997:Time Factors
D003470:Culture Media; D004269:DNA, Bacterial; D012329:RNA, Bacterial; C096222:tmRNA