Complete Genome Sequence of the Bacterium Porphyromonas gingivalis TDC60, Which Causes Periodontal Disease

Abstract

Porphyromonas gingivalis is a black-pigmented asaccharolytic anaerobe and a major causative agent of periodontitis. Here, we report the complete genome sequence of P. gingivalis strain TDC60, which was recently isolated from a severe periodontal lesion in a Japanese patient.

GENOME ANNOUNCEMENT

Porphyromonas gingivalis is a Gram-negative asaccharolytic bacterium which causes destructive periodontal diseases that result in a weakening of the tooth-supporting tissue and loss of teeth (7). Complete genomic sequences of two P. gingivalis laboratory strains, W83 and ATCC 33277, have been available for several years (11, 12). However, there is a need to analyze recently isolated strains without extensive subculture, because reports indicate a loss of virulence by a number of pathogenic bacteria after several passages in laboratory culture (1). P. gingivalis TDC60 was isolated from a severe periodontal lesion at Tokyo Dental College in Japan. Strain TDC60 exhibited higher pathogenicity in causing abscesses in mice than strains W83 and ATCC 33277 and other strains tested in the college. Thus, analyzing the genomic information of TDC60 is expected to provide new insights into the mechanisms of the onset and progression of periodontitis.

The complete genome sequence of TDC60 was determined by a combination of pyrosequencing (22,277,778-bp sequences; 9-fold coverage) and the Sanger method (16,934,400-bp sequences; 7-fold coverage). The pyrosequencing reads and the Sanger reads were assembled using the Newbler and Phred/Phrap/Consed, respectively. Gaps between adjacent contigs were closed by sequencing PCR amplicons from genomic DNA. Protein-coding sequences (CDSs) over length of 90 bp were predicted using a combination of MetaGeneAnnotator (13), GLIMMER (4), and the IMCGE software (In Silico Biology Co., Ltd., Japan). Functional annotation of CDSs was based on the results of the BLASTP searches against the NCBI nonredundant protein database. Insertion sequences (ISs), miniature inverted-repeat transposable elements (MITEs), conjugative transposons (CTns), and clustered regularly interspaced short palindromic repeats (CRISPRs) were identified using ISfinder (15), a combination of MUST (2) and MITE-hunter (8), a combination of Mauve (3) and GenomeMatcher (14), and CRISPRFinder (6), respectively. Nontranslated genes were predicted using the tRNAscan-SE (10), RNAmmer (9), and Rfam (5).

The genome of P. gingivalis TDC60 contained a single circular chromosome (2,339,898 bp; 48.34% GC content). The chromosome contained 2,220 CDSs, four rRNA operons, 53 tRNA sequences, and nine noncoding RNAs.

All-to-all BLASTP analysis of W83 and ATCC 33277 protein sequences showed that TDC60 possessed 382 strain-specific CDSs, and approximately two-thirds were annotated as hypothetical proteins. Of the CDSs encoding hypothetical proteins, 87 CDSs were on mobile elements, while some showed high homology to proteins encoded by genes of the Cytophaga-Flavobacteria-Bacteroides (CFB) group of bacteria. Strain TDC60 may have acquired these CDSs via horizontal gene transfer from multiple species of periodontal bacteria, and this might account for the unique features of this strain. Dot plot analysis comparison of TDC60 with W83 and ATCC 33277 genome sequences indicated a high degree of genome rearrangement among the three strains. It is speculated that mobile elements dispersed on the genome might have contributed to rearrangement of the genomic structure and led to the diversification of this species. Thus, the genomic information of P. gingivalis TDC60 could inform a comprehensive overview of the ecophysiology of this important oral pathogen and help develop new treatment methods for preventing periodontitis.

Nucleotide sequence accession number.

The completed genome sequence of P. gingivalis TDC60 was deposited in the DDBJ/EMBL/GenBank databases under accession no. AP012203.