The use of immuno-magnetic separation (IMS) as a tool in a sample preparation method for direct detection of L. monocytogenes in cheese

Abstract

A sample preparation procedure was developed for direct detection of L. monocytogenes in cheese. The sample preparation protocol consisted of a 10-fold dilution and homogenization, a centrifugation step to precipitate large food particles, passage of the supernatant over a sieve and through a separatory funnel to further eliminate food particles and fat, a centrifugation step to recover the bacterial pellet and finally enzymatic digestion of the suspension to degrade the remaining small food particles. Recovery of L. monocytogenes was confirmed by plating on Oxford medium and confirmation of suspected colonies. This protocol enabled direct detection (without prior enrichment) of low numbers of L. monocytogenes (0.5–1.5 cfu/g cheese) from different types of cheese. The performance of Dynabeads ® Anti-Listeria (Dynal, Oslo, Norway) for selective recovery of L. monocytogenes and their applicability in the above mentioned procedure for direct detection of low numbers of L. monocytogenes from cheese was evaluated. IMS could not separate and recover L. monocytogenes from the food particles in the concentrated suspension. The use of IMS after a 24 h enrichment procedure (as recommended by the manufacturer) allowed for the detection of low numbers of L. monocytogenes (<10 cfu/g). However, experiments in broth cultures showed that although the detection limit of IMS with Dynabeads ® Anti-Listeria was 40–100 cfu/ml, the ratio of L. monocytogenes to non- Listeria flora was not increased. Thus, selective enrichment or concentration of L. monocytogenes was not obtained.