Characterization of the Porphyromonas gingivalis conjugative transposon CTnPg1: determination of the integration site and the genes essential for conjugal transfer

Abstract

In our previous study, extensive genomic rearrangements were found in two strains of the Gram-negative anaerobic bacterium Porphyromonas (Por.) gingivalis, and most of these rearrangements were associated with mobile genetic elements such as insertion sequences and conjugative transposons (CTns). CTnPg1, identified in Por. gingivalis strain ATCC 33277, was the first complete CTn reported for the genus Porphyromonas. In the present study, we found that CTnPg1 can be transferred from strain ATCC 33277 to another Por. gingivalis strain, W83, at a frequency of 10−7 to 10−6. The excision of CTnPg1 from the chromosome in a donor cell depends on an integrase (Int; PGN_0094) encoded in CTnPg1, whereas CTnPg1 excision is independent of PGN_0084 (a DNA topoisomerase I homologue; Exc) encoded within CTnPg1 and recA (PGN_1057) on the donor chromosome. Intriguingly, however, the transfer of CTnPg1 between Por. gingivalis strains requires RecA function in the recipient. Sequencing analysis of CTnPg1-integrated sites on the chromosomes of transconjugants revealed that the consensus attachment (att) sequence is a 13 bp sequence, TTTTCNNNNAAAA. We further report that CTnPg1 is able to transfer to two other bacterial species, Bacteroides thetaiotaomicron and Prevotella oralis. In addition, CTnPg1-like CTns are located in the genomes of other oral anaerobic bacteria, Porphyromonas endodontalis, Prevotella buccae and Prevotella intermedia, with the same consensus att sequence. These results suggest that CTns in the CTnPg1 family are widely distributed among oral anaerobic Gram-negative bacteria found in humans and play important roles in horizontal gene transfer among these bacteria.

Localization of P42 and F 1 -ATPase α-Subunit Homolog of the Gliding Machinery in Mycoplasma mobile Revealed by Newly Developed Gene Manipulation and Fluorescent Protein Tagging

“…The DNA sequences flanking transposon insertion sites were determined using arbitrary PCR (43,44). The genomic DNA of the transformant was isolated by the agarose block method, as previously described (45).…”

Natural Competence Is a Major Mechanism for Horizontal DNA Transfer in the Oral Pathogen Porphyromonas gingivalis

“…Our results here indicate that the traA–Q locus in strain W83 is involved in low-frequency conjugal transfer of chromosomal DNA, and our previous work found that plasmids were not a substrate for the W83 transfer system (18). In contrast, the traA–Q locus in P. gingivalis ATCC 33277 can mobilize plasmids by a DNase I-insensitive mechanism, and a recent study revealed the traA–Q -containing element cTnPg1 to be capable of excision and conjugation (45); both behaviors are similar to those of conjugative Bacteroides transposons. These cTn-like elements may have been shared between the oral and intestinal Bacteroidetes populations at some time in evolutionary history, and similar loci are found in the genome sequences of other members of phylum Bacteroidetes , indicating that DNA conjugation by traA–Q is widespread in this taxonomic group.…”

Genetic exchange and reassignment in Porphyromonas gingivalis

“…They can excise themselves from the genome where they are integrated, transfer themselves to a recipient by conjugation and integrate into the recipient genome [39]. Thus CtnPg1, which was the first complete Ctn found in P. gingivalis , was able to excise itself from chromosomal DNA using an integrase (Int; PGN-0094) encoded in CtnPg1 and produce a circular intermediate [10].…”