Salmonella Enteritidis strains of egg- and non-egg-related origin were characterized molecularly to find markers correlated with the egg-contaminating property of Salmonella Enteritidis. Isolates were examined by random amplified polymorphic DNA (RAPD), plasmid profiling and phage typing. Furthermore, the presence of 30 virulence genes was tested by PCR. In genetic fingerprinting and gene content, only small differences between the strains were found and no correlation was observed with the origin (egg-related versus non-egg-related). A major RADP group was present in both egg- and non-egg-related strains, but other smaller RAPD groups were present as well in both categories of strains. Phage types PT4 and PT21 were predominant. Differential mRNA expression levels of fimA and agfA under conditions of growth simulating the conditions during egg formation were determined by real-time RT-PCR. Although differences in fimA and agfA expression levels were observed between the strains, these could not be correlated with the origin of the strains (egg-related versus non-egg-related). The highest expression levels of agfA and fimA were only found in two non-egg-related strains, which seemed to be correlated with the presence of a 93 kb plasmid instead of the 60 kb virulence plasmid. Our results seem to indicate only a limited role for at least type I fimbriae (encoded by fim operon) in egg contamination by Salmonella Enteritidis.