The aim of the present study was to apply descriptive, phylogenetic, recombination, and selection analyses on alignments of the Listeria Pathogenicity Island 1 (LIPI-1) of 1/2a and 4b Listeria monocytogenes isolates of different origin in order to gain insights into the evolution of this virulence gene cluster. For that purpose, a total of 19 L. monocytogenes isolates (9 meat isolates, serotype 1/2a; 5 meat isolates, serotype 4b; 5 strawberry isolates, serotype 4b) that have been previously separated at strain level were subjected to sequencing of their LIPI-1. Descriptive analysis revealed extensive nucleotide diversity mostly in the intragenic regions. The actA gene of 1/2a and 4b meat isolates and the hly gene of the 4b strawberry isolates exhibited the higher diversity; limited diversity was observed in prfA and plcA genes of the 4b isolates and mpl gene of the 1/2a isolates. Phylogenetic analysis of the complete island resulted in two major clusters that were consistent with serotype assignment of the isolates. Moreover, effective discrimination between serotypes was obtained by plcA, plcB, mpl, actA and the intergenic regions plcA-prfA and plcA-hly. In all cases but plcB and plcA-prfA 4b isolates were also differentiated according to their source of isolation as well. Selection analysis revealed that the island consisted of randomly evolving DNA with the exception of prfA gene of 1/2a isolates and actA gene of 4b meat isolates for which purifying selection or population expansion was indicated. Finally, no statistically significant evidence for recombination has been observed.