Frequency of putative periodontal pathogens among type 1 diabetes mellitus: a case-control study.

OBJECTIVE: The aim of the present study is to compare and assess the risk of periodontitis due to the presence of four putative periodontopathic bacteria viz., Eikenella corrodens, Campylobacter rectus, Prevotella intermedia and Prevotella nigrescens. To fulfil the above objective, polymerase Chain reaction using the primers targeting 16S rRNA gene of the bacterial species was performed with the subgingival plaque collected from the permanent first molars of type 1 diabetic children and age matched healthy children. RESULTS: The prevalence of periodontal pathogens in diabetic and healthy children was 6% and 16% for E. corrodens, 18% and 36% for C. rectus, 2% and 2% for P. intermedia, 4% and 0%, for P. nigrescens respectively. Statistically, significant difference was not observed for the prevalence of all the four periodontal pathogens between type 1 diabetic and healthy children (P = 1.00). The results of the present study thus reveal a negative correlation of type I diabetes to periodontitis in association to Eikenella corrodens, Campylobacter rectus, Prevotella intermedia and Prevotella nigrescens.

Purpura fulminans with Lemierre's syndrome caused by Gemella bergeri and Eikenella corrodens: a case report.

BACKGROUND: Gemella bergeri is one of the nine species of the genus Gemella and is relatively difficult to identify. We herein describe the first case of septic shock due to a Gemella bergeri coinfection with Eikenella corrodens. CASE PRESENTATION: A 44-year-old Asian man with a medical history of IgG4-related ophthalmic disease who was prescribed corticosteroids (prednisolone) presented to our hospital with dyspnea. On arrival, he was in shock, and a purpuric eruption was noted on both legs. Contrast enhanced computed tomography showed fluid retention at the right maxillary sinus, left lung ground glass opacity, and bilateral lung irregular opacities without cavitation. Owing to suspected septic shock, fluid resuscitation and a high dose of vasopressors were started. In addition, meropenem, clindamycin, and vancomycin were administered. Repeat computed tomography confirmed left internal jugular and vertebral vein thrombosis. Following this, the patient was diagnosed with Lemierre's syndrome. Furthermore, he went into shock again on day 6 of hospitalization. Additional soft tissue infections were suspected; therefore, bilateral below the knee amputations were performed for source control. Cultures of the exudates from skin lesions and histopathological samples did not identify any pathogens, and histopathological findings showed arterial thrombosis; therefore it was concluded that the second time shock was associated with purpura fulminans. Following this, his general status improved. He was transferred to another hospital for rehabilitation. The blood culture isolates were identified as Gemella bergeri and Eikenella corrodens. Gemella bergeri was identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry and confirmed by 16S rRNA gene sequencing later. The primary focus of the infection was thought to be in the right maxillary sinus, because the resolution of the fluid retention was confirmed by repeat computed tomography. CONCLUSIONS: Gemella bergeri can be the causative pathogen of septic shock. If this pathogen cannot be identified manually or through commercial phenotypic methods, 16S rRNA gene sequencing should be considered.

Purification and characterization of hemolysin from periodontopathogenic bacterium Eikenella corrodens strain 1073.

Eikenella corrodens 1073 was found to show hemolytic activity when grown on sheep blood agar. A high and dose-dependent hemolytic activity was detected in the cell envelope fraction, which was further purified by ion-exchange and gel-filtration chromatography. Consequently, a 65-kDa protein with hemolytic activity was obtained, suggesting that this protein might be a hemolysin. Its N-terminal amino acid sequence was nearly identical to that of X-prolyl aminopeptidase from E. corrodens ATCC 23834. To confirm that X-prolyl aminopeptidase functions as a hemolytic factor, we expressed the hlyA gene, encoding X-prolyl aminopeptidase, in Escherichia coli. After induction with isopropyl ß-D-1-thiogalactopyranoside, a protein of about 65 kDa was purified on a Ni column, and its hemolytic activity was confirmed. Meanwhile, a strain with a disrupted hlyA gene, which was constructed by homologous recombination, did not show any hemolytic activity. These results suggested that X-prolyl aminopeptidase might function as a hemolysin in E. corrodens.

LuxS affects biofilm maturation and detachment of the periodontopathogenic bacterium Eikenella corrodens.

Previously, we reported that biofilm formation of Eikenella corrodens is regulated by autoinducer-2 (AI-2), based on observations that biofilm-forming efficiency of ΔluxS mutant was greater than that of the wild type (Azakami et al., J. Biosci. Bioeng., 102, 110-117, 2006). To determine whether the AI-2 molecule affects biofilm formation directly, we added purified AI-2 to luxS mutant and wild-type E. corrodens and compared biofilm formations by using a static assay. Results indicated that biofilm formation in E. corrodens was enhanced by the addition of AI-2. We also compared the biofilms formed by flow cell system for the luxS mutant and the wild type by using scanning electron microscopy and confocal laser scanning microscopy. The number of viable bacteria in the luxS mutant biofilm was dramatically reduced and more sparsely distributed than that of the wild type, which suggested that AI-2 might enhance the mature biofilm. Conversely, further analysis by modified confocal reflection microscopy indicated that the wild-type biofilm was matured earlier than that of the luxS mutant, and became thinner and more sparsely distributed with time. These data suggest that LuxS may facilitate the maturation and detachment of biofilm in E. corrodens.

Genomic recombination through plasmid-encoded recombinase enhances hemolytic activity and adherence to epithelial cells in the periodontopathogenic bacterium Eikenella corrodens.

The periodontopathogenic bacterium Eikenella corrodens has an N-acetyl-D-galactosamine (GalNAc)-specific lectin, that contributes significantly to the pathogenicity of the bacterium. Recently, we reported that plasmid-mediated genomic recombination enhances the activity of this lectin. In this study, we investigated the effects of genomic recombination on certain virulence factors. Introduction of the recombinase gene resulted in hemolysis and significantly increased bacterial adhesion to epithelial cells. It was suggested that the enhanced adhesion was attributable to increased lectin activity due to genomic recombination, because it was inhibited by the addition of GalNAc. In contrast, invasion of the epithelial cells was remarkably reduced by genomic recombination. Although we assumed that this decrease in invasion resulted from a loss of type-IV pili, the phase variant did not show any decrease in invasion activity. This suggests that type-IV pili do not contribute to the invasive ability of E. corrodens. Our results suggest that genomic recombination enhances the pathogenicity of E. corrodens.

A pediatric patient with acute suppurative thyroiditis caused by Eikenella corrodens.

A previously healthy 6-year-old boy had continuous fever for 6 days before admission to our hospital. His general condition was good except for pyrexia. The left lobe of the thyroid gland was swollen, red, hot, and tender, and neck movement was limited. The provisional diagnosis was upper respiratory tract infection. We demonstrated the existence of an orifice of the left piriform sinus by esophagography in this case, and made a diagnosis of acute suppurative thyroiditis caused by a piriform sinus fistula in the hypopharynx. The causative organisms of acute suppurative thyroiditis include Peptostreptococcus, Staphylococcus haemolyticus, and α-streptococcus, but the organisms responsible are commonly undetectable in clinical cases. We detected Eikenella corrodens in the present patient. Although Eikenella species occasionally causes acute suppurative thyroiditis in adults, it is rare for this to happen in pediatric patients. Antibiotics were administered for 7 days. We also performed aspiration of the abscess on the 8th day of the illness. The abscess was reduced in size and tenderness was relieved after aspiration. In conclusion, if a pediatric patient has swelling, heat, tenderness, and redness of the anterior neck, we should keep in mind acute suppurative thyroiditis and the possibility of a fistula. If there is an abscess, we should immediately perform aspiration, culture, and isolation, and choose the appropriate antibiotics for the causative bacteria.

Quantification of five putative periodontal pathogens in female patients with and without chronic periodontitis by real-time polymerase chain reaction.

Chronic periodontitis is a highly prevalent endogenous polymicrobial disease. To better understand the etiology of the disease a quantitative approach is mandatory and real-time PCR is the molecular technique currently preferred to achieve this purpose. Taking into account that such a kind of study is still scarce, we aimed to evaluate the association between periodontal microbiota and chronic periodontitis. A total of 60 low-income age-matched female adults, 30 with chronic periodontitis and 30 without periodontal disease, were enrolled. DNA obtained from subgingival specimens was used for quantification of Aggregatibacter actinomycetemcomitans, Eikenella corrodens, Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia by real-time PCR. A. actinomycetemcomitans, E. corrodens, and F. nucleatum were detected in all subjects, P. gingivalis was observed in 70.0% and 46.6% and P. intermedia in 90.0% and 80.0% of chronic periodontitis patients and periodontally healthy subjects, respectively. P. gingivalis mean count was significantly higher in patients with chronic periodontitis than in periodontally healthy individuals. Accurate detection and quantification of five putative periodontal pathogens was feasible using a simple and fast real-time PCR protocol. Although P. gingivalis and P. intermedia have been found more commonly in chronic periodontitis patients, no statistical difference was observed between periodontally diseased and healthy groups. Quantitative data indicated association between P. gingivalis and chronic periodontitis. However, because of its uneven distribution, it should not be solely taken as a marker of periodontal status.

Culture-independent detection of Eikenella corrodens and Veillonella parvula in primary endodontic infections.

Eikenella corrodens and Veillonella parvula are normal cultivable inhabitants of the oral cavity but their presence in endodontic infections has not been as common as it could be anticipated. This might have been because of shortcomings of culture techniques when it comes to bacterial isolation or identification. The present study intended to survey samples from primary endodontic infections for the presence of E. corrodens and V. parvula using a culture-independent 16S rRNA gene-based nested PCR protocol. Genomic DNA was isolated directly from samples taken from different forms of periradicular lesions, and the presence of E. corrodens and V. parvula was determined by nested PCR. Specificity for each primer pair was confirmed by sequence analysis of PCR products from positive clinical samples. V. parvula and E. corrodens were, respectively, detected in 33% and 14% of the root canals associated with chronic apical periodontitis. Both V. parvula and E. corrodens were found in 10% of the cases diagnosed as acute apical periodontitis. V. parvula and E. corrodens were detected in 21% and 26% of the samples from acute apical abscesses, respectively. In general, species-specific nPCR allowed the detection of V. parvula in 24% and E. corrodens in 18% of the samples taken from primary endodontic infections. Findings confirmed that V. parvula and E. corrodens can take part in the microbiota of primary endodontic infections, but in prevalence values somewhat higher when compared to most of the previous culture studies that had reported recovery of these species.

Plasmid-mediated genomic recombination at the pilin gene locus enhances the N-acetyl-D-galactosamine-specific haemagglutination activity and the growth rate of Eikenella corrodens.

Eikenella corrodens belongs to a group of periodontopathogenic bacteria and forms unique corroding colonies on solid medium due to twitching motility. It is believed that an N-acetyl-D-galactosamine (GalNAc)-specific lectin on the cell surface contributes significantly to its pathogenicity and can be estimated by its haemagglutination (HA) activity. Recently, a plasmid, pMU1, from strain 1073 has been found; this plasmid affects pilus formation and colony morphology. To identify the gene involved in these phenomena, ORF 4 and ORFs 5-6 on pMU1 were separately subcloned into a shuttle vector, and the resultant plasmids were introduced into E. corrodens 23834. Transformants with the ORF 4 gene, which is identified to be a homologous gene of the type IV pilin gene-specific recombinase, lost their pilus structure and formed non-corroding colonies on a solid medium, whereas transformants with ORFs 5-6 exhibited the same phenotype as the host strain 23834. Southern analysis showed that the introduction of the ORF 4 gene into strain 23834 resulted in genomic recombination at the type IV pilin gene locus. The hybridization pattern of these transformants was similar to that of strain 1073. These results suggest that ORF 4 on pMU1 encodes a site-specific recombinase and causes genomic recombination of the type IV pilin gene locus. Furthermore, the introduction of ORF 4 into strain 23834 increased GalNAc-specific HA activity to a level equivalent to that of strain 1073. Although the morphological colony changes and loss of pilus structure are also observed in phase variation, genomic recombination of the type IV pilin gene locus did not occur in these variants. Moreover, an increase was not observed in the GalNAc-specific HA activity of these variants. These results suggested that the loss of pilus structure, the morphological change in colonies and the increase in HA activity due to plasmid pMU1 might be caused by a mechanism that differs from phase variation, such as a genomic recombination of the type IV pilin gene locus.

A new checkerboard panel for testing bacterial markers in periodontal disease.

BACKGROUND/AIMS: Various microbiological methods have been used for testing bacterial markers for periodontitis and periodontal disease progression. Most studies have used only a limited number of well recognized bacterial species. The purpose of the present study was to evaluate the association of 13 more recently identified bacterial species in a new panel in comparison with 12 previously more recognized periodontotopathogens ('old panel') using the 'checkerboard' DNA-DNA hybridization method. METHODS: Fifty individuals were chosen who showed at least one site with a probing pocket depth of 6 mm or more (disease) and bleeding on probing and at least one site with a probing pocket depth of 3 mm and without bleeding on probing (health). One diseased and one healthy site on each individual were sampled with the paperpoint technique and the samples were processed in the checkerboard technique against deoxigenin-labeled whole genomic probes to 25 subgingival species representing 12 well recognized and 13 newly identified periodontitis associated species. RESULTS: Twenty-four (out of 25) species were detected more frequently in the subgingival plaque of diseased than healthy sites both at score 1 (> 10(4)) and score 3 (> 10(5)). A significant difference at the higher score (score 3) was noticed for all species of the old panel except for three (Streptococcus intermedius, Selenomonas noxia, and Eikenella corrodens). Of the species in the new panel only Prevotella tannerae, Filifactor alocis, and Porphyromonas endodontalis showed a statistical significant difference between diseased and healthy sites. CONCLUSION: It was concluded that P. tannerae, F. alocis, and P. endodontalis should be added to the 12 species used for routine diagnostics of periodontitis-associated bacterial flora.

Isolation and characterization of a plasmid DNA from periodontopathogenic bacterium, Eikenella corrodens 1073, which affects pilus formation and colony morphology.

Eikenella corrodens (Ec) is one of a group of periodontopathogenic bacteria. A plasmid DNA (8.7 kb) isolated from Ec 1073 was designated pMU1. Agarose gel electrophoresis and Southern analysis suggested that pMU1-like plasmids were carried in 2 Ec strains, including 1073, with higher hemagglutination (HA) activity than other strains. We determined the nucleotide sequence of this plasmid and identified 7 ORFs. A homology search revealed that 4 ORFs of pMU1 were homologous to ORFs in pJTPS1, found in a spontaneous avirulent mutant of the phytopathogenic bacterium, Ralstonia solanacearum. pJTPS1 is a putative hypovirulent plasmid, which is thought to control the virulence of R. solanacearum. We also found the ORF to be homologous to the recombinase specific to the type IV pilin gene. We introduced a part of pMU1 into the Ec 23834 strain, which has a pilus structure on its cell surface and forms corroding colonies on solid medium. No pilus structure was observed on the surface of transformants, most of which formed non-corroding colonies. When such transformants (or Ec 1073) were cured of pMU1 with acridine orange, they remained non-foliated and non-corroding. The results suggest that pMU1 might irreversibly affect pilus formation and colony morphology, and might be involved in the pathogenicity and virulence of Ec.

Clonal diversity and stability of subgingival Eikenella corrodens.

Eikenella corrodens is a commensal subgingival bacterium commonly found in both periodontally nondiseased and diseased subjects. The present study examined the clonal diversity and stability of subgingival E. corrodens over time. Ninety-five subjects were enrolled at the baseline examination, including 44 periodontally nondiseased subjects and 51 subjects with aggressive periodontitis. Twenty-two nondiseased subjects and 11 subjects with aggressive periodontitis were subsequently reexamined after an average interval of 14 months. Two subgingival plaque samples were obtained from each subject to determine the total cultivable bacteria. In addition, multiple E. corrodens isolates from each sample were recovered for clonal analysis by arbitrarily primed PCR. The mean numbers of distinct E. corrodens clones harbored by nondiseased subjects and subjects with aggressive periodontitis were 1.3 and 3.0, respectively. Thirty-nine percent of the nondiseased subjects and 63% of the subjects with aggressive periodontitis harbored multiple clones of E. corrodens. The numbers of distinct E. corrodens clones increased significantly (Mann-Whitney ranking test, P < 0.05) in sites from patients with aggressive periodontitis, in sites with pocket depths of 4 mm or greater, in sites with a clinical attachment loss of 2 mm or greater, and in sites coinfected with Porphyromonas gingivalis. Comparison of E. corrodens clones recovered at the baseline and those recovered at the follow-up examination showed that E. corrodens colonization was not stable. Thirty-eight of the 66 follow-up samples (58%) showed a complete change (including de novo colonization of the sites or complete elimination of the organism from the sites) of the subgingival E. corrodens clonal types between the baseline and the follow-up examinations. Our results suggest a complexity of subgingival microbiota not seen previously.

Characterization and expression of adjacent proline iminopeptidase and aspartase genes from Eikenella corrodens.

Two adjacent genes involved in nitrogen metabolism from Eikenella corrodens, with a potential role in pathogenesis, were studied. Proline iminopeptidase (Pip) activity, which may be essential for energy production and protection against host immune mechanisms, is exhibited by E. corrodens. Analysis of Pip-expressing clones revealed an ORF of 939 bases with a predicted amino acid sequence identity of 67% to the Pip of Neisseria gonorrhoea. 200 bp downstream from pip, an ORF of 1395 bases, encoding a protein with 87% identity to a putative aspartase from the Neisseria meningitidis genome sequence, was identified. Enzymatic function was confirmed with a complemented Escherichia coli aspartase deficient mutant. The E. corrodens aspartase was found to be 77% identical to the Haemophilus influenzae aspartase sequence, which was originally identified on the basis of its ability to bind plasminogen. However, the E. corrodens aspartase had no such activity. Southern hybridization indicated both genes to be single copy and conserved within the genomes of a diverse panel of E. corrodens isolates from health and disease.

[Eikenella corrodens-induced spondylitis. Detection with 16s-RNA polymerase chain reaction]. / Eikenella-corrodensinduzierte Spondylitis. Nachweis mittels 16s-RNA-Polymerasekettenreaktion.

Isolation of the relevant organism in patients with spondylitis even after an open biopsy is successful only in 75-90%. The rare case of an Eikenella corrodens-induced spondylitis is presented, which could only be identified using 16S ribosomal DNA polymerase chain reaction following unsuccessful microbiological cultivation. Eikenella corrodens is a facultative anaerobic gram-negative organism, which is mostly found in the oropharynx of healthy patients.

Role of the Eikenella corrodens pilA locus in pilus function and phase variation.

The human pathogen Eikenella corrodens expresses type IV pili and exhibits a phase variation involving the irreversible transition from piliated to nonpiliated variants. On solid medium, piliated variants form small (S-phase), corroding colonies whereas nonpiliated variants form large (L-phase), noncorroding colonies. We are studying pilus structure and function in the clinical isolate E. corrodens VA1. Earlier work defined the pilA locus which includes pilA1, pilA2, pilB, and hagA. Both pilA1 and pilA2 predict a type IV pilin, whereas pilB predicts a putative pilus assembly protein. The role of hagA has not been clearly established. That work also confirmed that pilA1 encodes the major pilus protein in this strain and showed that the phase variation involves a posttranslational event in pilus formation. In this study, the function of the individual genes comprising the pilA locus was examined using a recently developed protocol for targeted interposon mutagenesis of S-phase variant VA1-S1. Different pilA mutants were compared to S-phase and L-phase variants for several distinct aspects of phase variation and type IV pilus biosynthesis and function. S-phase cells were characterized by surface pili, competence for natural transformation, and twitching motility, whereas L-phase cells lacked these features. Inactivation of pilA1 yielded a mutant that was phenotypically indistinguishable from L-phase variants, showing that native biosynthesis of the type IV pilus in strain VA1 is dependent on expression of pilA1 and proper export and assembly of PilA1. Inactivation of pilA2 yielded a mutant that was phenotypically indistinguishable from S-phase variants, indicating that pilA2 is not essential for biosynthesis of functionally normal pili. A mutant inactivated for pilB was deficient for twitching motility, suggesting a role for PilB in this pilus-related phenomenon. Inactivation of hagA, which may encode a tellurite resistance protein, had no effect on pilus structure or function.

Eikenella corrodens phase variation involves a posttranslational event in pilus formation.

The human pathogen Eikenella corrodens synthesizes type IV pili and exhibits a phase variation involving the irreversible transition from piliated to nonpiliated variants. On solid medium, piliated variants form small (S-phase), corroding colonies whereas nonpiliated variants form large (L-phase), noncorroding colonies. We are studying the molecular basis of this phase variation in the clinical isolate E. corrodens VA1. A genomic fragment encoding the major type IV pilin was cloned from the S-phase variant of strain VA1. Sequence analysis of the fragment revealed four tandemly arranged potential open reading frames (ORFs), designated pilA1, pilA2, pilB, and hagA. Both pilA1 and pilA2 predict a type IV pilin. The protein predicted by pilB shares sequence identity with the Dichelobacter nodosus FimB fimbrial assembly protein. The protein predicted by hagA resembles a hemagglutinin. The region containing these four ORFs was designated the pilA locus. DNA hybridization and sequence analysis showed that the pilA locus of an L-phase variant of strain VA1 was identical to that of the S-phase variant. An abundant pilA1 transcript initiating upstream of pilA1 and terminating at a predicted hairpin structure between pilA1 and pilA2 was detected by several assays, as was a less abundant read-through transcript encompassing pilA1, pilA2, and pilB. Transcription from the pilA locus was nearly indistinguishable between S- and L-phase variants. Electron microscopy and immunochemical analysis showed that S-phase variants synthesize, export, and assemble pilin into pili. In contrast, L-phase variants synthesize pilin but do not export and assemble it into pili. These data suggest that a posttranslational event, possibly involving an alteration in pilin export and assembly, is responsible for phase variation in E. corrodens.

Subgingival microbial profile of Papillon-Lefèvre patients assessed by DNA-probes.

The prevalence of 18 selected bacterial species was assessed by means of "checkerboard" DNA-DNA hybridisation in a group of 12 Saudi-Arabian adolescents with Papillon-Lefèvre syndrome. A total of 36 tooth sites were investigated. The patients exhibited severe periodontal disease with deep pockets. All 12 patients harboured the putative bacterial pathogens P. intermedia, F. nucleatum, P. micros and S. intermedius while T. denticola, B. forsythus, P. nigrescens, E. corrodens, S. noxia and C. rectus were recovered from 11 patients. P. gingivalis was recovered from 9 patients and 18 sites while corresponding figures for A. actinomycetemcomitans were 8 and 19, respectively. A number of the investigated species (B. forsythus, T. denticola, P. intermedia, C rectus) reached high levels (> or =10(6) cells) in more than 1/2 of the patients. On the other hand, bacteria such as A. actinomycetemcomitans and P. gingivalis were infrequently encountered at high levels in these subgingival samples. In conclusion, the analysis failed to demonstrate a PLS-specific profile of the subgingival infection, since the bacterial composition of the sampled sites closely resembled that characterising deep pockets in adult periodontitis patients.

Variable serum immunoglobulin G immune response to genetically distinct Eikenella corrodens strains coexisting in the human oral cavity.

This study examined the variable serum immunoglobulin G (IgG) levels to genetically distinct autologous Eikenella corrodens strains by enzyme-linked immunosorbent assay (ELISA). Twenty subjects, including 10 adult periodontitis patients, 5 juvenile periodontitis patients and 5 periodontally healthy subjects were examined. Each subject was colonized by 2-8 genetically distinct E. corrodens strains. The serum IgG levels to autologous E. corrodens within individuals were significantly different in 7 adult periodontitis patients, 4 juvenile periodontitis patients and a periodontally healthy subject. Poor correlation was found in diseased subjects between serum IgG levels to autologous strains and to reference strains ATCC 23834 or FDC 373. Four adult periodontitis patients and two juvenile periodontitis patients exhibited significant serum IgG levels to autologous E. corrodens strains (two standard deviations above the mean for periodontally healthy subjects); two of these six diseased subjects exhibited low serum IgG levels to reference strains and would have been classified as low immune responders if only reference strains had been used in ELISA. This study showed the importance of using autologous E. corrodens strains in the assessment of serum IgG immune responses to this organism.

"Checkerboard" versus culture: a comparison between two methods for identification of subgingival microbiota.

The present study compared the "checkerboard" DNA-DNA hybridization methodology with culture techniques for the analysis of the composition of the subgingival microbiota. 70 subjects, presenting with a variety of periodontal conditions, contributed with a total of 283 subgingival plaque samples analyzed with respect to the following species: Porphyromonas gingivalis, Prevotella intermedia/Prevotella nigrescens, Fusobacterium nucleatum, Campylobacter rectus, Eikenella corrodens, Bacteroides forsythus, Actinobacillus actinomycetemcomitans, Streptococcus sanguis and Streptococcus mutans. Species identification and quantification was performed by (i) the checkerboard method, using whole genomic, digoxigenin labeled DNA probes; and (ii) culture, including non-selective and selective media in combination with routine biochemical testing using commercial test panels. We found that the checkerboard technology resulted in higher prevalence figures for half of the species tested when compared to culture data. If the latter were used as the reference, checkerboard detection sensitivities ranged from 0.17 to 0.86, specificities from 0.17 to 1.0, and diagnostic accuracies from 0.51 to 0.81, depending on bacterial species. The use of the checkerboard data as the reference resulted in detection sensitivities for the culture procedures between 0.24 and 1.0 and specificities between 0.21 and 0.87. The checkerboard methodology resulted in statistically significant higher bacterial counts for the majority of the species. It was further observed that, for most species, the higher the total number colony-forming units in the sample, the higher the discrepancy between the results obtained by the two techniques.