Enumeration of Viable Listeria Species and Listeria monocytogenes in Foods

Abstract

A direct plating procedure was developed for the enumeration of Listeria spp. and Listeria monocytogenes in foods. Both naturally contaminated foods and foods spiked with L. innocua, L. seeligeri, and L monocytogenes were studied. The enhanced hemolysis agar (EHA) developed by Cox and modified in our study resulted in two types of agar, referred to as listeria enumeration agar (LEA) no. 1 and 2, used for products of lighter and heavier background microbial populations, respectively. On LEA plates, total Listeria spp. counts were determined by fluorescence caused by the breakdown of 4-methylumbelliferyl-β-d-glucoside contained in EHA. L. monocytogenes counts were determined by picking a representative number of hemolytic colonies and stabbing them into a xylose agar plate to distinguish L. monocytogenes from L seeligeri. Contamination levels of >200 Listeria cells per g of food can be accurately quantified by this procedure with >80% recovery. Counts of <200 Listeria cells per g of food were considered estimates. When the level of contamination was < 100 Listeria cells per g of food, the recovery was <58%. Occasionally, with low-level inocula, Listeria was not detected. Nevertheless, when the procedure was combined with incubation of the enrichment mixture (used for the 1:10 direct plating dilution) and subsequent streaking, Listeria contamination could still be detected and the level, therefore, was determined to be between 1 and 150/g.