Development of a qPCR direct detection method for Listeria monocytogenes in milk

There is a growing movement among consumers in the US and Europe towards minimally processed foods, including raw milk and dairy products. This trend significantly increases exposure to dairy-borne pathogens and indicates a need for rapid, sensitive screening tests for raw dairy products to reduce risk. We recently developed a centrifugation-concentration-qPCR method that can be used to detect and quantify Escherichia coli O157 in raw milk at ~10 cfu/mL. The work presented here describes development of a similar methodology for the detection of Listeria monocytogenes in milk. To circumvent some challenges associated with raw milk, initial efforts utilized pasteurized commercial whole milk. Recovery of L. monocytogenes at each sample clean-up step was studied to optimize overall recovery. Multiple PCR targets and polymerase mixes were tested to optimize sensitivity and resistance to PCR inhibitors. Initial results indicate that L. monocytogenes can be detected in pasteurized milk samples at a level of 10 cfu/mL.