Sulfate-reducing bacteria in relation with other potential periodontal pathogens.

BACKGROUND, AIMS: Oral sulfate-reducing bacteria are involved in several clinical categories of periodontitis. The aim of this cross-sectional study was to compare the presence of sulfate-reducing bacteria (SRB) with other putative pathogens including spirochetes, Actinobacillus actinomycetemcomitans, Bacteroides forsythus, Porphyromonas gingivalis, and Treponema denticola in periodontal lesions. METHOD: Periodontal SRB were detected by enrichment culture and compared with a microscopic spirochete count (n=168). Species-specific oligonucleotide probes directed against the 16S rRNA were employed to determine the presence of A. actinomycetemcomitans, P. gingivalis, B. forsythus, and T. denticola (n=55). RESULTS: A significant positive correlation was observed between the presence of SRB and the proportions of spirochetes in subgingival plaque, although the 2 bacterial groups also occurred separately. SRB tended to be negatively correlated with the presence of A. actinomycetemcomitans. In contrast, all pockets with SRB harbored either T. denticola, or both T. denticola and B. forsythus (12/14) before therapy. Interestingly, the combination of SRB with P. gingivalis occurred in 32% of the periodontal pockets before treatment. After initial periodontal therapy, the prevalence of this combination was reduced to 2% of the sites, and to 25% of the sites in recall patients. CONCLUSION: The presence of SRB was positively correlated with T. denticola, B. forsythus, and P. gingivalis in periodontal lesions. These suspected pathogens form a complex strongly associated with destructive periodontitis.

Decrease of sulfate-reducing bacteria after initial periodontal treatment.

Sulfate-reducing bacteria (SRB) are associated with periodontitis, but it is unknown if elimination of these potential pathogens accompanies clinical improvement. This longitudinal study examined the occurrence of SRB and clinical effects following scaling and root planing. In this study, the presence of periodontal SRB was determined in 38 selected patients before and six months after mechanical therapy. SRB were detected by the enrichment culture technique. Mechanical periodontal treatment resulted in elimination of SRB in 89% of the patients, and 95% of the sites (n = 76). SRB were significantly reduced in patients with progressive, adult, and refractory periodontitis. The elimination of SRB was accompanied by clinical improvement. The mean gain of attachment of these pockets was 3 mm (p < 0.001). The reductions in pocket depth (p < 0.001) and bleeding were significant (p < 0.001). Persistence of SRB correlated with the initial pocket depth (p < 0.02) and attachment level (p < 0.02), and with bleeding of the site after treatment (p < 0.05). In conclusion, mechanical debridement is generally effective for the elimination of SRB.

The djlA gene acts synergistically with dnaJ in promoting Escherichia coli growth.

The DnaK chaperone of Escherichia coli is known to interact with the J domains of DnaJ, CbpA, and DjlA. By constructing multiple mutants, we found that the djlA gene was essential for bacterial growth above 37 degrees C in the absence of dnaJ. This essentiality depended upon the J domain of DjlA but not upon the normal membrane location of DjlA.

DjlA is a third DnaK co-chaperone of Escherichia coli, and DjlA-mediated induction of colanic acid capsule requires DjlA-DnaK interaction.

DjlA is a 30-kDa type III membrane protein of Escherichia coli with the majority, including an extreme C-terminal putative J-domain, oriented toward the cytoplasm. No other regions of sequence similarity aside from the J-domain exist between DjlA and the known DnaK (Hsp70) co-chaperones DnaJ (Hsp40) and CbpA. In this study, we explored whether and to what extent DjlA possesses DnaK co-chaperone activity and under what conditions a DjlA-DnaK interaction could be important to the cell. We found that the DjlA J-domain can substitute fully for the J-domain of DnaJ using various in vivo functional complementation assays. In addition, the purified cytoplasmic fragment of DjlA was shown to be capable of stimulating DnaK ATPase in a manner indistinguishable from DnaJ, and, furthermore, DjlA could act as a DnaK co-chaperone in the reactivation of chemically denatured luciferase in vitro. DjlA expression in the cell is tightly controlled, and even its mild overexpression leads to induction of mucoid capsule. Previous analysis showed that DjlA-mediated induction of the wca capsule operon required the RcsC/RcsB two-component signaling system and that wca induction by DjlA was lost when cells contained mutations in either the dnaK or grpE gene. We now show using allele-specific genetic suppression analysis that DjlA must interact with DnaK for DjlA-mediated stimulation of capsule synthesis. Collectively, these results demonstrate that DjlA is a co-chaperone for DnaK and that this chaperone-co-chaperone pair is implicated directly, or indirectly, in the regulation of colanic acid capsule.

Identification of Tn10 insertions in the rfaG, rfaP, and galU genes involved in lipopolysaccharide core biosynthesis that affect Escherichia coli adhesion.

Escherichia coli was used as a model to study initial adhesion and early biofilm development to abiotic surface. Tn10 insertion mutants of Escherichia coli K-12 W3110 were selected for altered abilities to adhere to a polystyrene surface. Seven insertion mutants that showed a decrease in adhesion harbored insertions in genes involved in lipopolysaccharide (LPS) core biosynthesis. Two insertions were located in the rfaG gene, two in the rfaP gene, and three in the galU gene. These adhesion mutants were found to exhibit a deep-rough phenotype and to be reduced, at different levels, in type 1 fimbriae production and motility. The loss of adhesion exhibited by these mutants was associated with either the affected type 1 fimbriae production and/or the dysfunctional motility. Apart from the pleiotropic effect of the mutations affecting LPS on type 1 fimbriae and flagella biosynthesis, no evidence for an involvement of the LPS itself in adhesion to polystyrene surface could be observed.

Identification of Tn10 insertions in the dsbA gene affecting Escherichia coli biofilm formation.

Escherichia coli was used as model to study initial adhesion and early biofilm development to an abiotic surface. Tn10 insertion mutants with reduced attachment to a polystyrene surface were isolated. Three adhesion mutants harbored the transposon in the dsbA gene, whose product, DsbA, catalyses folding of numerous extracytoplasmic disulfide bond-containing proteins. All three mutants were weakly adherent and grew poorly. Cell surface structure analysis showed that motility. type 1 fimbriation and lipopolysaccharide structure were affected in these mutants. The pleiotropic effect of the dsbA mutations on biofilm formation is discussed.

A rapid screening procedure to identify mini-Tn10 insertion mutants of Escherichia coli K-12 with altered adhesion properties.

A rapid screening procedure was developed for detection of Escherichia coli mutants with altered adhesion abilities using polystyrene 96-well microtiter plates as attachment surfaces. During this assay, bacterial strains grew and adhered simultaneously, and attached cells were measured after crystal violet staining. Starting with a total of 7000 W3110::Tn10 insertion mutants of E. coli K-12 W3110, 50 adhesion-deficient mutants were isolated which showed less than 40% attachment, and 22 mutants were found with an attachment of 40-75%. Motility assays were performed on these 72 mutants, and 34 displayed altered motility.