Based an the findings that plastids and cyanobacteria have similar group I introns inserted into tRNA(UAA)(Leu) genes, these introns have been suggested to be immobile and of ancient origin. In contrast, recent evidence suggests lateral transfer of cyanobacterial group I introns located in tRNA(UAA)(Leu) genes. In light of these new findings, we have readdressed the evolution and lateral transfer of tRNA(UAA)(Leu) group I introns in cyanobacteral radiation, We determined the presence of introns in 38 different strains, representing the major cyanobacterial lineages, and characterized the introns in 22 of the strains, Notably, two of these strains have two tRNA(UAA)(Leu) genes, with each of these genes interrupted by introns, while three of the strains have bath interrupted and uninterrupted genes. Two evolutionary distinct clusters of tRNA genes, with the genes interrupted by introns belonging to two distinct intron clusters, were identified. We also compared 16S rDNA and intron evolution for both closely and distantly related strains. The distribution of the introns in the clustered groups, as defined from 16S rDNA analysis, indicates relatively recent gain and/or loss of the introns in some of these lineages. The comparative analysis also suggests differences in the phylogenetic trees for 16S rDNA and the tRNA(UAA)(Leu) group I introns, Taken together, our results show that the evolution of the intron is considerably more complex than previous studies found to be the case, We discuss, based on our results, evolutionary models involving lateral intron transfer and models involving differential loss of the intron.