Characterization of Porphyromonas gingivalis insertion sequence-like element ISPg5

Porphyromonas gingivalis, a black-pigmented, gram-negative anaerobe, is fou nd in periodontitis lesions, and its presence in subgingival plaque signifi cantly increases the risk for periodontitis. In contrast to many bacterial pathogens, P. gingivalis strains display considerable variability, which is likely due to genetic exchange and intragenomic changes. To explore the la tter possibility, we have studied the occurrence of insertion sequence (IS) -like elements in P. gingivalis W83 by utilizing a convenient and rapid met hod of capturing IS-like sequences and through analysis of the genome seque nce of P. gingivalis strain W83. We adapted the method of Matsutani et al. (S. Matsutani, H. Ohtsubo, Y. Maeda, and E. Ohtsubo, J. Mol. Biol. 196:445- 455, 1987) to isolate and clone rapidly annealing DNA sequences characteris tic of repetitive regions, within a genome. We show that in P. gingivalis s train W83, such sequences include (i) nucleotide sequence with homology to tRNA genes, (ii) a previously described IS element, and (iii) a novel IS-li ke element. Analysis of the P. gingivalis genome sequence for the distribut ion of the least used tetranucleotide, CTAG, identified regions in many of the initial 218 contigs which contained CTAG clusters. Examination of these CTAG clusters led to the discovery of 11 copies of the same novel IS-like element identified by the repeated sequence capture method of Matsutani et al. This new 1,512-bp IS-like element, designated ISPg5, has features of th e IS3 family of IS elements. When a recombinant plasmid containing much of ISPg5 was used in Southern analysis of several P. gingivalis strains, inclu ding clinical isolates, diversity among strains was apparent. This suggests that ISPg5 and other IS elements may contribute to strain diversity and ca n be used for strain fingerprinting.