Pulsed-field gel electrophoresis (PFGE) was used to compare the limited number of large restriction fragments generated by digesting DNA of Listeria monocytogenes strains with restriction enzymes characterized by rare recognition sequences. Sixteen macro-restriction patterns were observed with ApaI and SmaI, and 7 with NotI, among 42 strains of serovar 4b, the most important serovar in human listeriosis epidemiology. Analysis of these restriction fragment length polymorphisms enabled a rapid differentiation of genetically closely related strains, revealing differences between strains which initially appeared similar by other typings. The results of this study suggested that the PFGE protocol could be a useful addition to methods currently available for epidemiological analysis of human listeriosis.