Aims: Listeria monocytogenes Delta gadD1 and Delta lisK mutants display enhanced and reduced sensitivity, respectively, to the food preservative nisin in laboratory media. However, the behaviour of these strains in a nisin-containing food has not been assessed. Here we use cottage cheese as a model food to address this issue. Materials and Results: Antibiotic-resistant forms of the wild-type and mutant strains were employed to investigate the behaviour of multiple strains in a single food sample, thereby eliminating the problem of intersample variation. Using this approach, it was established that percentage survival of the Delta lisK mutant was greater than the parent strain in the absence of nisin and that this relative difference became even more dramatic in cottage cheese supplemented with nisin. The numbers of the Delta gadD1 mutant decreased more rapidly than the parent in cottage cheese without nisin, but surprisingly this trend was reversed in nisin-supplemented cheese. Upon the addition of 10 mmol l-1 monosodium glutamate, a substrate for the glutamate decarboxylase (GAD) system, the wild-type LO28 strain regained its relative advantage over Delta gadD1. Conclusions: Care needs to be taken when predicting the behaviour of mutants of L. monocytogenes with altered resistance to nisin in food as experiments in laboratory media are not always a good indicator of how the strains will behave in such food environments. Significance and impact of the Study: This study further emphasizes the importance of utilizing food matrices to confirm observations made using laboratory media.