Molecular diagnostics for dairy-borne pathogens.

Abstract

Advances in diagnostic assays based on nucleic acids will revolutionize the ability of the industry to maintain the safety of dairy foods. Two complementary assay formats are explored, one of which permits the rapid detection of bacterial pathogens and the other the identification of reservoirs of these pathogens. The first format is an assay based on the polymerase chain reaction that employs homogeneous detection (TaqMan polymerase chain reaction detection; Perkin Elmer, Applied Biosystems Division, Foster City, CA) of the target sequence. This assay has been applied to the detection of Listeria monocytogenes. A primary problem with current assays that are based on polymerase chain reaction is the complexity of sample handling and the quantification of the initial target number. This fluorogenic assay takes advantage of the endogenous 5',3'-endonuclease activity in Taq DNA polymerase. Approximately 100 samples can be analyzed in 2 to 3 h with a sensitivity of < 50 cells and a dynamic range of > 1000-fold. The TaqMan polymerase chain reaction detection assay is a robust format that is readily applicable to a wide array of other pathogens found in foods and in the environment. The second format is an instrument for automated ribosomal RNA analysis (Riboprinter; DuPont, Wilmington, DE) that can be used to locate the reservoirs harboring the bacterial pathogen. Use of this typing method it has been shown that, although a number of different ribotypes can be isolated from a single environmental sample, only a selected number of these strains apparently have the ability to cause disease. The future of food microbiology lies in the development and integration of molecular methods that can be automated into a testing regimen that extends from the farm to finished products.