The foodborne pathogen Listeria monocytogenes has the ability to develop biofilm in the food-processing environment, which becomes a major concern for food safety. PrfA, a key transcriptional activator that regulates most of the known listerial virulence gene expression, has been shown to promote L. monocytogenes biofilm formation. In this study, the whole-genome microarray was used to identify differentially expressed genes associated with the putative interaction between biofilm formation and PrfA in L. monocytogenes. Comparative transcriptome analyses indicated that over 21.9 % of the L. monocytogenes EGDe genes (627 out of 2,857 predicted) were altered in their expression of biofilm compared to the planktonic phase. These genes were classified into different functional categories which cover most of the biochemical functions encountered in bacterial cells, indicating that L. monocytogenes biofilm formation is probably controlled by a complex regulation network involved in variable genes required for the different biological pathways. Further comparison of gene expression profiles of biofilms between L. monocytogenes EGDe and its PrfA deletion mutant revealed 185 genes associated with PrfA and biofilm formation. Except for 10 genes, transcription levels of 175 genes were completely opposite between Delta prfA and wild-type during the biofilm formation, i.e., up-regulated genes in Delta prfA were down-regulated in the wild-type strain, and vice versa, indicating that loss of PrfA dramatically altered gene expression patterns in L. monocytogenes biofilm and resulted in reduced ability of the biofilm formation.