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Hereditary gelsolin amyloidosis (AGel amyloidosis) is a rare, dominantly inherited systemic disease with worldwide distribution, caused by c.654G > A or c.654G > T gelsolin gene mutation. The disease mainly manifests with late-onset dystrophy of the cornea, laxity of the skin and dysfunction of the cranial nerves whereas the oral manifestations have remained less-studied. To examine if AGel amyloidosis also affects salivary gland function, we studied 27 patients. In a questionnaire, 89% of them reported oral dryness, and 74% oral and ocular dryness. Unstimulated (UWS) and stimulated whole salivary flow (SWS) rates were measured, and salivary proteins were analyzed in the patients and controls. Hyposalivation according to UWS was detected in 67% of the patients, while decreased SWS occurred in 63% of the patients and 19% of the controls (p = 0.001). The secretion rates of salivary total protein and IgA were significantly lower in patients than controls. Histopathological analyses of labial salivary gland biopsies showed deposition of gelsolin amyloid, atrophy and inflammation. This study showed that AGel amyloidosis belongs to the differential diagnostic choices to be kept in mind in the patients presenting with xerostomia, low secretion rates of salivary total protein and IgA and/or deposition of amyloid in the minor salivary glands. AGel amyloidosis patients should be advised for efficient dental care.

The study investigated bacteria sampled from the mandibular third molar pericoronitis with the hypothesis that the same species also can be detected in several sites of the oral cavity. Subgingival plaque samples were taken from 41 patients (mean age 23.2 ± 4.2 years), from the third molar pericoronal pocket and from 3 adjacent gingival pockets, from the tongue and saliva. Aggregatibacter actinomycetemcomitans (A.a.), Porphyromonas gingivalis (P.g.), Tan- ne-rella forsythia (T.f.), Prevotella intermedia (P.i.), Pre-votella nigrescens (P.n.), and Treponema denticola (T.d.) were analyzed by qualitative PCR method. All the species analyzed were found in the pericoronitis samples in the following frequency: T.f. 73%, T.d. 68%, P.i. + P.n. 35%, A.a. 5%, and P.g. 5%. Saliva and tongue samples were also highly positive with T.f. (both 76%) and T.d. (saliva 57%, tongue 76%) and better sources for A.a. (saliva 8%, tongue 14%), and P.g. (saliva 14%, tongue 8%) than the pericoronitis pocket. The gingival pocket samples were less frequently positive the further the site in question was from the pericoronitis site. Contrary to earlier observations T.f. was frequently defected in pericoronitis sites. The results emphasize the importance of pericoronitis as a focus of infection.

The aim of our study was to investigate how teenage smoking affects the prevalence of periodontal bacteria and periodontal health with the hypothesis that smoking increases the prevalence of the bacteria. Oral health of 264 adolescents (15- to 16-year-olds) was clinically examined, and their smoking history was recorded. The participants also filled in a structured questionnaire recording their general health and health habits. Pooled subgingival plaque samples were taken for polymerase chain reaction analysis of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Prevotella intermedia, Prevotella nigrescens, and Treponema denticola. The prevalence of P. intermedia (21% vs. 4%, p = 0.01) and T. forsythia and T. denticola (23% vs. 8%, p

The effects of smoking on periodontal biomarkers in adolescents are unknown. This study investigates matrix metalloproteinase (MMP)-8 and polymorphonuclear leukocyte elastase levels in saliva together with periodontal health indices accounting for body mass index and smoking in a birth cohort from Finland.The oral health of boys (n = 258) and girls (n = 243) aged 15 to 16 years was examined clinically. Health habits were assessed by questionnaire. Saliva samples were collected and analyzed by immunofluorometric and peptide assays for MMP-8 levels and polymorphonuclear leukocyte elastase activities, and investigated statistically with the background factors.Median MMP-8 values of male smokers were 112.03 microg/l compared to 176.89 microg/l of non-smokers (P = 0.05). For female smokers corresponding values were 170.88 microg/l versus 177.92 microg/l in non-smokers (not statistically significant). Elastase values in male smokers were 5.88 x 10(-3) Delta OD(405)/h versus 11.0 x 10(-3) Delta OD(405)/h in non-smokers (P = 0.02), and in female smokers 9.16 x 10(-3) Delta OD(405)/h versus 10.88 x 10(-3) Delta OD(405)/h in non-smokers (P = 0.72). The effect was strengthened by high pack-years of smoking (MMP-8, P = 0.04; elastase, P = 0.01). Both biomarkers increased with gingival bleeding. However, statistically significant associations were observed with bleeding on probing and MMP-8 (P = 0.04); MMP-8 was suggestively associated with probing depth (P = 0.09) in non-smoking boys. In smokers with calculus, MMP-8 increased after adjusting with body mass index (P = 0.03). No corresponding differences were seen in girls.Smoking significantly decreased both biomarkers studied. Compared to girls, boys seem to have enhanced susceptibility for periodontitis as reflected in salivary MMP-8 values.

To assess the prevalence and antifungal drug sensitivity of non-albicans Candida (NAC) species in elderly outpatients. Materials and methods: We investigated oral rinse samples of 194 self-caring elderly population (mean age 83 years) with emphasis on background factors for harbouring NAC. Susceptibility of Candida species to antifungal drugs was determined using standard methodology. Multiple logistic regression analysis was performed taking positive NAC count as the dependent variable and a number of known Candida risk factors as independent variables. Results: Prevalence of candidal carriage of the population was 78.4%, of which 0.5% of the subjects were NAC positive. Candida dubliniensis was the most prevalent NAC species, followed by Candida glabrata and Candida parapsilosis. The NAC positive elderly were more often edentulous with dental prostheses or had fewer teeth than Candida albicans-positive or yeast-negative subjects. Dental caries slightly increased the risk for having NAC strains (odds ratio 1.08), whilst greater age appeared to lower the risk (odds ratio 0.77). Candida species were susceptible to the commonly used antifungal agents in general, but with considerable variation among species. Occasionally, some NAC exhibited lower antifungal susceptibility. Conclusion: The possibility of oral reservoirs of NAC strains which are resistant to common antifungals should be noted in elderly outpatients.

Objectives: Oral yeast infections are an emerging problem among medically compromised and frail elderly. Antifungal drug resistance is also increasing because of an increase in non-albicans Candida strains in these populations. We therefore set out to study, in the randomised-controlled trial setting if the use of a topical amine fluoride-stannous luoride combination (AmF-SnF 2) could control oral Candida growth in the elderly. The hypothesis was based on earlier findings showing that in vitro this combination had antifungal efficacy. Methods: A total of 194 nursing home residents were randomised to receive either the test mouth rinse and toothpaste or a placebo twice daily for 8 months. Of these, 136 completed the trial. Saliva samples were taken using the oral rinse method, cultivated and the strain level identified with routine microbial methods. Compliance and use of preparations was assessed by a nurse. Results: Significantly at the end of the trial, less mucosal lesions were observed in the test group in comparison to controls. Total bacterial count decreased in both the groups during the trial. Candida albicans was the most prevalent strain detected both at baseline and 8 months later. Only a few subjects carried non-albicans strains. The AmF-SnF 2 did not significantly affect mean oral Candida counts, but median Candida counts were reduced in the AmF-SnF 2 group while an increase was seen in the placebo group. However, the differences observed were not statistically significant. Compliance among the regular elderly users slightly increased during the trial for both the groups. Conclusion: The number of subjects with high Candida counts decreased in the AmF-SnF 2 group. Hence, the fluoride combination might be useful as a support therapy for oral candidiasis. Prevalence of non-albicans Candida strains was low in this population.

Aim: The aim of the study was to evaluate the adhesion of Lactobacillus delbrueckii subsp. bulgaricus and Lactobacillus rhamnosus strain GG to saliva-coated surfaces in vitro. Methods: Fifteen radiolabeled dairy L. delbrueckii subsp. bulgaricus strains and L. rhamnosus GG were tested for their ability to adhere to saliva-coated hydroxyapatite beads and polystyrene microtiter plates and the radioactivity was measured by liquid scintillation counter. The effects of lysozyme on the adhesion of lactobacilli and of pretreatment with lactobacilli on the adhesion of Streptococcus sanguinis were also assessed. Results: All strains tested adhered to saliva-coated surfaces but with significantly different binding frequencies. The adhesion of the L. delbrueckii subsp. bulgaricus strains remained lower in comparison to L. rhamnosus strain GG. One L. delbrueckii subsp. bulgaricus strain showed binding frequency comparable to S. sanguinis. Lysozyme pretreatment of the samples significantly increased lactobacillus adhesion to saliva-coated surfaces. Conclusion: The present results showed significant variations in the adhesion capacity of the Lactobacillus strains studied. Adhesion to oral surfaces is of primary importance for bacterial colonization in the mouth. Only one of the L. delbrueckii subsp. bulgaricus dairy starter culture strains investigated had a high adhesion percentage. This strain might then be considered for further investigations in the oral environment.

Tissue reactions to bacteria lead to proinflammatory reactions involving matrix metalloproteinases (MMPs). Synthetic protease inhibitors may offer new possibilities to regulate bacterial proteases. We investigated proteolytic activities of certain periodontal bacteria, their effects on the latent proMMP-9, and the effects of synthetic MMP inhibitors and a serine protease inhibitor Pefabloc. The strains studied werePorphyromonas gingivalis, Prevotella intermedia, Peptostreptoccus micros, Prevotella nigrescens, Fusobacterium nucleatum,and5 Aggregatibacter actinomycetemcomitansserotypes. Their gelatinolytic activities and the effects of certain synthetic MMP inhibitors and Pefabloc were analyzed by zymography. Bacterial effects on proMMP-9 conversion were investigated by Western immunoblot. All investigated periodontal bacteria produced gelatinolytic cell-bound and extracellular proteinases which could fragment latent proMMP-9, suggesting co-operative processing cascades in oral tissue remodeling. A. actinomycetemcomitans produced the weakest gelatinolytic activity. Synthetic proteinase inhibitors exhibited slight but clear reductive effects on the bacterial proteolytic activities. We conclude that targeted anti-proteolytic treatment modalities against bacterial-host proteolytic cascades can be developed.

Background: Human laminin-332 (Lm-332) degradation by 12 Candida strains and effects of synthetic proteinase inhibitors [Ilomastat (ILM), EDTA, chemically modified tetracycline-3(CMT-3), CMT-308, synthetic peptide CTT-2, and Pefabloc] were studied. Materials and methods: Laminin-332 was incubated with sonicated cell fractions and 10 times concentrated cell-free fractions of reference and clinical strains of C. albicans, C. dubliniensis, C. guilliermondii, C. glabrata, C. krusei, and C. tropicalis. Proteolysis, pH effects, and inhibitors were analyzed by fluorography and zymography. Results: Cell fractions of all species except C. guilliermondii and cell-free fractions of C. albicans, and C. dubliniensis showed 20–70 kDa gelatinases at pH 5.0 and 6.0. At pH 7.6, C. glabrata, C. krusei, and C. tropicalis cell fractions and C. tropicalis cell-free fractions showed 55–70 kDa gelatinases. CMT-3, CMT-308, and CTT-2 inhibited Candida gelatinases slightly better than Pefabloc, ILM, and EDTA. No Candida fractions degraded Lm-332 at pH 7.6, but at pH 5.0, 100 kDa bands were generated by cell fractions of C. dubliniensis and C. tropicalis; C. albicans and C. glabrata clinical strains; and C. guilliermondii reference strain. C. krusei reference strain yielded three 100–130 kDa bands. C. albicans, C. dubliniensis, and C. tropicalis reference and clinical strain’s cell-free fractions generated 100 kDa band. Conclusions: Laminin-332 degradation is pH-dependent and differences exist between studied Candida strains. Lm-332 degradation can exert functional disturbances on basement membrane integrity, possibly aiding Candida cell invasion into tissues. Certain synthetic matrix metalloproteinase inhibitors (CMTs, CTT) can inhibit Candida proteinases and may be therapeutically useful in future.

Statement of the Problem: Oligodontia (severe partial anodontia) is a developmental dental anomaly. It is either an isolated trait or part of a syndrome. Oligodontia is characterized by the congenital absence of more than six permanent teeth except the third molars. Treatment often calls for facilitated surgical techniques, but less severe cases can be treated conventionally in a normally equipped dental office. Case Presentation: A case of oligodontia treated with prosthetic rehabilitation is presented in this article to remind clinicians of the anomaly. A medical history was taken and clinical and radiographic examinations were made. It is emphasized that conventional prosthetic treatment can lead to a satisfactory result. The patient was treated with fully extending denture prostheses. An overdenture was designed in the maxillary arch, while a removable partial denture was made in the mandibular arch. Results: The patient’s speech and masticatory function improved greatly. He was also pleased with better facial esthetics. Observed temporomandibular joint dysfunction also ameliorated after the treatment. Conclusion: Dental clinicians should keep in mind that there are good possibilities with conventional prosthodontic techniques to help patients with dental anomalies. Treatment not only improves speech and masticatory function but also has psychological implications that may greatly help in regaining self-confidence. CLINICAL SIGNIFICANCE Patients suffering from oligodontia may have severe psychological, esthetic, and functional problems. Thus, early diagnosis and treatment of these patients are necessary.

Alcohol is a well documented risk factor for upper digestive tract cancers. It has been shown that acetaldehyde, the first metabolite of ethanol is carcinogenic. The role of microbes in the production of acetaldehyde to the oral cavity has previously been described in several studies. In the present study, the aim was to investigate the capability of viridans group streptococci of normal oral flora to produce acetaldehyde in vitro during ethanol incubation. Furthermore, the aim was to measure the alcohol dehydrogenase (ADH) activity of the bacteria. Eight clinical strains and eight American Type Culture Collection (ATCC) strains of viridans group streptococci were selected for the study. Bacterial suspensions were incubated in two different ethanol concentrations, 11 mM and 1100 mM and the acetaldehyde was measured by gas chromatography. ADH-activity was measured by using a sensitive spectroscopy. The results show significant differences between the bacterial strains regarding acetaldehyde production capability and the detected ADH-activity. In particular, clinical strain of Streptococcus salivarius, both clinical and culture collection strains of Streptococcus intermedius and culture collection strain of Streptococcus mitis produced high amounts of acetaldehyde in 11 mM and 1100 mM ethanol incubation. All these four bacterial strains also showed significant ADH-enzyme activity. Twelve other strains were found to be low acetaldehyde producers. Consequently, our study shows that viridans group streptococci may play a role in metabolizing ethanol to carcinogenic acetaldehyde in the mouth. The observation supports the concept of a novel mechanism in the pathogenesis of oral cancer.

Introduction: Most probiotic products are consumed orally and hence it is feasible that the bacteria in these products may also attach to oral surfaces; however, the effects of these bacteria on the oral ecosystem are mostly unknown. Our aim was to evaluate the oral colonization potential of different probiotic, dairy, and fecal Lactobacillus and Bifidobacterium strains in vitro. Methods: The binding of 17 Lactobacillus and seven Bifidobacterium strains to hydroxyapatite and microtitre wells coated with human saliva was tested. Binding of selected strains to human buccal epithelial cells and co-adherence with Fusobacterium nucleatum were also investigated. In addition, the survival in sterilized human whole saliva was examined. Results: There was a large variation in binding to saliva-coated surfaces and buccal epithelial cells but all strains survived in saliva. The binding pattern of the probiotics did not differ from the binding of the fecal strains. F. nucleatum altered the binding of both the low-binding bifidobacteria and the high-binding lactobacilli. Conclusion: The differences in binding in vitro may indicate that there are also differences in the persistence of the different probiotic strains in the oral cavity in vivo.

Background and Objectives: To investigate the levels of matrix metalloproteinase (MMP) -8 and -9 with the simultaneous presence of periodontal pathogens in gingival crevicular fluid (GCF) as well as MMP-9 and cholesterol in blood. Although bacterial pathogens are required to initiate the periodontal disease process, in some individuals the reaction to bacteria may lead to an excessive host response, resulting in a general inflammatory response. Methods: MMP-9 and lipids were analyzed from the blood samples of 33 subjects with a 16-year history and oral health records of periodontal disease as well as from 31 periodontally healthy controls. Information was obtained on education, body mass index, and family history of atherosclerosis. GCF was taken to determine MMP-8 and MMP-9 levels, and bacterial samples were simultaneously collected for polymerase chain reaction assessment of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Prevotella nigrescens, Tannerella forsythia, and Treponema denticola. Analysis of variance, chi-squared test, and multiple logistic regression analysis were used to analyze the results. Results: Demographic data showed significant differences between patients and controls in smoking (P

The aim of this study was to investigate the simultaneous presence of periodontal microbiota on inflammatory markers in gingival crevicular fluid from individuals with periodontal diseases.A total of 82 individuals with periodontal disease (mean age: 54.3 +/- 3.0 years) and 31 periodontally healthy individuals (mean age: 53.2 +/- 3.0 years) were randomly chosen and underwent clinical oral examinations in 2003 with the determination of the dental plaque index (PI), gingival index (GI), and periodontal probing depth (PD). The simultaneous presence of polymerase chain reaction (PCR)-assessed periodontal bacteria, levels of prostaglandin E(2) (PGE(2)), granulocyte elastase, interleukin 1-beta (IL-1beta), and total protein concentration were determined from the pockets. Marginal bone height percent was measured on x-rays. Analysis of variance and chi(2) tests were used to analyze the results.In sites with Tannerella forsythensis, levels of PGE(2) (pg/site), granulocyte elastase (monoclonal antibodies (mAbs)/site), and total protein (mg/ml) were significantly higher than in sites without T. forsythensis (P0.05, P0.01, and P0.05, respectively). Those with periodontal disease with simultaneous presence of T. forsythensis and Porphyromonas gingivalis, or T. forsythensis and Prevotella nigrescens, showed significantly higher PI and GI, deeper PD, more loss of attachment, and more release of PGE(2) and granulocyte elastase than did periodontitis patients without these bacteria.The simultaneous presence of T. forsythensis and P. gingivalis, or T. forsythensis and P. nigrescens, seemed to promote the release of subgingival inflammatory mediators and seemed to be associated with more severe periodontal disease.

Adhesion of Candida species to prosthetic acrylic resins is an essential first step in the pathogenesis of denture stomatitis. Data on the relative adhesion of pathogenic non-albicans Candida species to different denture base materials are sparse. The purpose of the present study was to investigate in vitro adhesion of C. albicans, C. glabrata, C. krusei and C. dubliniensis to four different denture base materials. Specimens of both heat-cured resins (Vertex(TM) Rapid Simplified and ProBase Hot) and cold-cured resins (Paladur A and Paladur B) were prepared using a novel method and the adhesion of four strains each of the foregoing Candida species evaluated microscopically using a soft imaging system. There was a significant difference in yeast adherence between Vertex and the other resins. Only C. glabrata attached to Vertex, while all the remainder of the tested species adhered to all other resins tested except ProBase, which resisted C. krusei adhesion. There was a significant difference in candidal adhesion between cold-cured and heat-cured resins for three Candida species (C. albicans, P = 0.039; C. glabrata, P = 0.002 and C. krusei, P = 0.000). The type of denture base material and whether they are heat-cured or cold-cured play an important role in modifying candidal adhesion.

Objective: The combination of amine fluoride and stannous fluoride (AmF/SnF2) was, by chance, found to be antifungal in a clinical trial. This study investigated its effect on pathogenic Candida species with the hypothesis that the antifungal action on different species is variable. Materials and methods: Growth inhibition effect of Meridol® mouth rinse which contains 250 ppm AmF/SnF2 was evaluated on 43 reference and clinical strains of Candida albicans, C. dubliniensis, C. glabrata, C. guilliermondii, C. krusei, C. parapsilosis, and C. tropicalis. Meridol® base solution without AmF/SnF2 was used as a negative control. Results: Undiluted Meridol® mouth rinse killed most study strains within a few minutes. In ascending order, C. parapsilosis, C. tropicalis, C. albicans, C. glabrata, C. krusei and C. dubliniensis showed higher resistance against AmF/SnF2 than C. guilliermondii. Conclusion: AmF/SnF2 could be used as a potent adjunct to antifungal therapy for oral yeasts. Although different Candida species demonstrated variable sensitivity the most prevalent oral yeast C. albicans appeared sensitive to the AmF/SnF2 combination.

Objectives: The increased plaque formation observed in gingival inflammation is not fully understood. Receptor proteins in the dental pellicle might influence bacterial adhesion and subsequent plaque formation. The purpose of the present study was to examine proteins and microorganisms in dental biofilms, at healthy and inflamed gingival margins. Material and methods: To see whether marginal inflammation affects the composition of the pellicle and the early dental plaque, samples were taken from the gingival and incisal parts of teeth in periodontally healthy subjects, both in gingival health and during experimental gingivitis. Pellicle proteins were analysed using gel-electrophoresis, immunoblotting and image analysis. Bacteria were analysed by culturing and the PCR technique. Results: During gingivitis, the rate of plaque formation increased significantly. The semiquantitative amounts of proteins and the numbers of bacteria varied considerably between individuals and surfaces. The amount of total and individual pellicle proteins and the total numbers of bacteria were, however, increased during gingivitis and the increase in proteins was statistically significant on the incisal parts of tooth surfaces. Compared to a healthy gingiva, reduced numbers for Actinomyces spp. (incisal parts only) and streptococci and increased numbers of periodontopathogens in the 4 h dental biofilms were seen at the inflamed gingiva. Conclusion: Increased gingival crevicular fluid flow during gingivitis affects pellicle formation and increased plasma proteins in the pellicle may modify bacterial attachment and early dental plaque composition.

Treponema denticola is an important periodontal pathogen capable of tissue invasion. Its chymotrypsin-like proteinase (CTLP) can degrade a number of basement membrane components in vitro, thus suggesting a contribution to tissue invasion by the spirochete. The aim of this study was to analyze the localization of CTLP in chronic periodontitis tissues ex vivo. A polyclonal antibody specific to T. denticola cell-bound CTLP was used to detect the spirochetes in the gingival tissues of patients with moderate to severe chronic periodontitis (n=25) by immunohistochemistry and periodic acid-Schiff staining (PAS). The presence of T. denticola in the periodontal tissue samples was analyzed by PCR. Periodontal tissue samples of 12 of the 25 patients were found to be positive for T. denticola by PCR. Moreover, CTLP could be detected in the periodontal tissues of all these patients by immunohistochemistry. In the epithelium, the CTLP was mostly intracellular. Typically, the positive staining could be seen throughout the whole depth of the epithelium. When detected extracellularly, CTLP was localized mainly as granular deposits. The connective tissue stained diffusely positive in four cases. The positive staining co-localized with the PAS stain in nine cases. T. denticola and its CTLP could be detected in diseased human periodontium both intra- and extracellularly. The granular staining pattern was suggestive of the presence of T. denticola bacteria, whereas the more diffused staining pattern was indicative of the recent presence of the bacterium and shedding of the cell-bound proteinase.Keywords: dentilisin; spirochetes; major outer sheath protein(Published: 4 July 2014)Citation: Journal of Oral Microbiology 2014, 6: 24349 - http://dx.doi.org/10.3402/jom.v6.24349

Nonopsonic phagocytosis of Bacillus cereus by human polymorphonuclear leukocytes (PMNs) with particular attention to bacterial surface properties and structure was studied. Two reference strains (ATCC 14579 T and ATCC 4342) and two clinical isolates (OH599 and OH600) from periodontal and endodontic infections were assessed for adherence to matrix proteins, such as type I collagen, fibronectin, laminin, and fibrinogen. One-day-old cultures of strains OH599 and OH600 were readily ingested by PMNs in the absence of opsonins, while cells from 6-day-old cultures were resistant. Both young and old cultures of the reference strains of B. cereus were resistant to PMN ingestion. Preincubation of PMNs with the phagocytosis-resistant strains of B. cereus did not affect the phagocytosis of the sensitive strain. Negatively stained cells of OH599 and OH600 studied by electron microscopy had a crystalline protein layer on the cell surface. In thin-sectioned cells of older cultures (3 to 6 days old), the S-layer was observed to peel off from the cells. No S-layer was detected on the reference strains. Extraction of cells with detergent followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a major 97-kDa protein from the strains OH599 and OH600 but only a weak 97-kDa band from the reference strain ATCC 4342. One-day-old cultures of the clinical strains (hydrophobicity, 5.9 to 6.0%) showed strong binding to type I collagen, laminin, and fibronectin. In contrast, reference strains (hydrophobicity, −1.0 to 4.2%) as well as 6-day-old cultures of clinical strains (hydrophobicity, 19.0 to 53.0%) bound in only low numbers to the proteins. Gold-labelled biotinylated fibronectin was localized on the S-layer on the cell surface as well as on fragments of S-layer peeling off the cells of a 6-day-old culture of B. cereus OH599. Lactose, fibronectin, laminin, and antibodies against the S-protein reduced binding to laminin but not to fibronectin. Heating the cells at 84°C totally abolished binding to both proteins. Benzamidine, a noncompetitive serine protease inhibitor, strongly inhibited binding to fibronectin whereas binding to laminin was increased. Overall, the results indicate that changes in the surface structure, evidently involving the S-layer, during growth of the clinical strains of B. cereus cause a shift from susceptibility to PMN ingestion and strong binding to matrix and basement membrane proteins. Furthermore, it seems that binding to laminin is mediated by the S-protein while binding to fibronectin is dependent on active protease evidently attached to the S-layer.

Background/aims: Lactobacillus rhamnosus GG (LGG) is one of the most widely studied probiotic bacterial strain. The benefits of LGG treatment in gastrointestinal disorders are well documented. The aim of the present study was to investigate whether LGG can be detected in the oral cavity after discontinuation of administration of a product prepared with this bacterium. Material and methods: 56 volunteers consumed Gefilus® juice (Valio Ltd, Helsinki, Finland) containing LGG during a 14-day trial period. Saliva samples were collected and cultured onto MRS agar after a clearance period and then daily after a 2-week intervention period for as long as LGG was found. LGG-like colonies were analyzed in saliva samples, identified by characteristic colony morphology, a lactose fermentation test, and PCR with specific primers. Results: LGG was not able to colonize the oral cavity. It could only be temporarily detected. In one female subject, however, whose medical history revealed use of LGG in childhood, the bacterium was detected in all saliva samples taken up to 5 months. (She was excluded from the intervention trial). Conclusion: Permanent colonization of LGG in the oral cavity is improbable but seems possible in individual cases.

Although S-layers are being increasingly identified on Bacteria and Archaea, it is enigmatic that in most cases S-layer function continues to elude us. In a few instances, S-layers have been shown to be virulence factors on pathogens (e.g. Campylobacter fetus ssp. fetus and Aeromonas salmonicida), protective against Bdellovibrio, a depository for surface-exposed enzymes (e.g. Bacillus stearothermophilus), shape-determining agents (e.g. Thermoproteus tenax) and nucleation factors for fine-grain mineral development (e.g. Synechococcus GL 24). Yet, for the vast majority of S-layered bacteria, the natural function of these crystalline arrays continues to be evasive. The following review up-dates the functional basis of S-layers and describes such diverse topics as the effect of S-layers on the Gram stain, bacteriophage adsorption in lactobacilli, phagocytosis by human polymorphonuclear leukocytes, the adhesion of a high-molecular-mass amylase, outer membrane porosity, and the secretion of extracellular enzymes of Thermoanaerobacterium. In addition, the functional aspect of calcium on the Caulobacter S-layer is explained.

In our previous study, we reported that only 13 of 46 adult patients with advanced periodontitis responded well to initial non-surgical periodontal therapy. In the present follow-up study, the remaining 33 patients were randomly treated further using either modified Widman flap surgery or systemic metronidazole. The patients responding unsatisfactorily to this 2nd treatment phase, received supplementary systemic chemotherapy or surgery, respectively. By using this study design, we determined which baseline clinical variables and/or laboratory findings predicted the treatment outcome in these study patients. Clinical variables included the assessment of bleeding, suppuration, probing pocket depth, furcation lesions, relative attachment level and radiographic infrabony defects. Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis were cultured from subgingival plaque samples. The specific IgG and IgA antibody levels against 5 serotypes of A. actinomycetemcomitans were determined in serum and saliva. Elastase-like, trypsin-like and general protease activities were assessed from saliva. The bivariate statistical analyses showed that the most pronounced difference between the patients responding well to initial non-surgical therapy (group MC, n = 13), to either supplementary surgery or chemotherapy (group FT1, n = 11), or those responding to the complex therapy (group FT2, n = 17), was the prior extent of periodontal destruction expressed as the proportion of > or = 6 mm deep periodontal pockets. When multiple linear regression was used to investigate the influence of clinical and laboratory findings on the variation of treatment response between the 3 groups, the most significant explanatory factor was the simultaneous presence of subgingival A. actinomycetemcomitans and multiple deep periodontal pockets. None of the immunological or biochemical variables used had any further influence in the model. Pretreatment microbiological examination, especially for the detection of A. actinomycetemcomitans, seems to be a valuable laboratory screening method for identifying complex treatment need in adult patients with advanced periodontitis. However, the evaluation of the extent and pattern of periodontal breakdown remains crucial for choosing the treatment strategy including surgery and/or chemotherapy in A. actinomycetemcomitans-infected adult periodontitis patients.

Probiotic consumption and repeated probiotic intake have shown promising results as adjunct therapies in prevention and alleviation of some chronic disease conditions in the gastrointestinal tract. Recent evidence suggests that probiotics may also be beneficial in preventing oral diseases. An efficient probiotic candidate in the mouth, however, should not impose any risk to oral tissues, such as acid demineralisation of tooth enamel because of sugar fermentation. The aim of the present in vitro study was to evaluate the utilisation of some sugars and sugar alcohols by yogurt starter Lactobacillus delbrueckii subsp. bulgaricus strains and to assess the influence of these carbohydrate sources on cell surface properties. For comparsion, a commercially available probiotic, Lactobacillus rhamnosus GG, was used. The results showed that lactose, glucose and fructose were readily metabolised by all strains tested. However, strain-specific metabolic patterns were observed when other sugars and sugar alcohols were used as sole carbohydrate source in the growth medium. Surface properties of the bacteria such as hydrophobicity and surface-associated proteins appeared to vary with the carbohydrate content of the growth medium. Based on these results it can be concluded that among the L. delbrueckii subsp. bulgaricus strains probiotic candidate strains are available that warrant further studies due to their inability to ferment sugars with pronounced cariogenic properties.

Objective We investigated in subjects with and without periodontitis, the levels of certain salivary proteins and matrix metalloproteinase-8 (MMP-8) in gingival crevicular fluid (GCF), in relation to the presence of specific periodontal pathogens. Methods Clinical parameters were recorded at baseline, in 1985 and in 2009 from 99 subjects; 56 with and 43 without periodontitis (mean age 59.2 ± SD 2.9). Saliva samples collected in 2009 were analysed for salivary albumin, total protein and immunoglobulins A, G and M. GCF was collected for analysis of MMP-8 levels and for the PCR-analysis of the microorganisms Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Treponema denticola and Tannerella forsythia. Results Periodontitis patients were more often infected by P. gingivalis (p

To assess the prevalence and antifungal drug sensitivity of non-albicans Candida (NAC) species in elderly outpatients.We investigated oral rinse samples of 194 self-caring elderly population (mean age 83 years) with emphasis on background factors for harbouring NAC. Susceptibility of Candida species to antifungal drugs was determined using standard methodology. Multiple logistic regression analysis was performed taking positive NAC count as the dependent variable and a number of known Candida risk factors as independent variables.Prevalence of candidal carriage of the population was 78.4%, of which 0.5% of the subjects were NAC positive. Candida dubliniensis was the most prevalent NAC species, followed by Candida glabrata and Candida parapsilosis. The NAC positive elderly were more often edentulous with dental prostheses or had fewer teeth than Candida albicans-positive or yeast-negative subjects. Dental caries slightly increased the risk for having NAC strains (odds ratio 1.08), whilst greater age appeared to lower the risk (odds ratio 0.77). Candida species were susceptible to the commonly used antifungal agents in general, but with considerable variation among species. Occasionally, some NAC exhibited lower antifungal susceptibility.The possibility of oral reservoirs of NAC strains which are resistant to common antifungals should be noted in elderly outpatients.

Background Despite conflicting results on the use of hormone replacement therapy (HRT) there is no doubt that many women benefit from it. Women using HRT are known to be more health conscious in general with putative positive implications in the mouth. However, we observed recently in our cohort hardly any difference in oral health status between HRT-users and non-users. There are only a few studies about HRT and oral microbiota. We hypothesized that counts of periodontal micro-organisms are lower in health-conscious HRT-users than non-users. Methods Two-year open follow-up study was conducted on originally 200 HRT-users and 200 non-users from age cohorts of 50-58 years. After clinical examination pooled subgingival plaque samples were taken for polymerase chain reaction analyses. The results of finally 135 women meeting the inclusion criteria were analyzed with cross-tabulation and chi-square test. Explanatory factors were studied by step-wise logistic regression analysis. Results In HRT group, the numbers of positive samples for Porphyromonas gingivalis (P. gingivalis, p < 0.07), Prevotella intermedia (P. intermedia, p < 0.05)and Tannerella forsythia (T. forsythia, p < 0.01) decreased in women with ≥ 4-mm-deep pockets. Respectively in HRT users with ≥ 6-mm-deep pockets the numbers of positive samples for P. gingivalis (p < 0.05) and T. forsythia (p < 0.01) were decreased. No corresponding differences were observed in the non-HRT group. In logistic regression, the existence of deep periodontal pockets explained the majority of cases harboring specific micro-organisms in both groups. Conclusion Although use of HRT did not correlate with periodontal health status, HRT led to decreasing numbers of positive samples of the periodontal pathogens P. gingivalis and T. forsythia. Further studies with longer observation time are needed to observe the clinical relevance of the results.

In oral medicine and dentistry probiotics have shown promising results in controlling dental diseases and yeast infections. This study was made to investigate the effect of eight strains of Lactobacillus bulgaricus and their effects on human matrix metalloproteinase-9 (MMP-9). The hypothesis was that these bacteria used in yoghurt production for centuries are not proteolytic and thus can be safely used in the development of probiotic preparations. Bacterial cell fractions and supernatant specimens were prepared and studied with gelatinase zymography and MMP-9 activation was assessed by immunoblotting. The effect of synthetic MMP inhibitors and a serine protease inhibitor (Pefabloc) on bacterial proteinases was studied with zymography. The results showed very low gelatinolytic activity. There was a slight difference between the supernatant and cell fractions so that the supernatant specimens produced weak gelatinolytic bands in zymography while hardly anything was seen in the cell fraction series. The tested synthetic MMP inhibitors and Pefabloc did not affect the proteolytic activity of the lactobacilli strains. The lactobacilli did not seem to induce the conversion of proMMP-9 to its active form. Consequently, our study hypothesis was confirmed and the studied Lactobacillus strains are not likely to degrade host tissue components.

OBJECTIVES: Porphyromonas gingivalis is a significant periodontal pathogen that has been shown in vitro to be able to invade gingival epithelial cells and grow intracellularly. The aim of the present study was to detect P. gingivalis in gingival tissues from chronic periodontitis (CP) patients. MATERIALS AND METHODS: Monoclonal antibodies specific to a cell membrane-bound thiol proteinase of P. gingivalis were used to detect the microbe in gingival tissues of CP patients (n = 13) by immunohistochemistry. The presence of P. gingivalis was also analysed by polymerase chain reaction (PCR). RESULTS: Immunohistochemical analysis of the periodontal tissues revealed positive staining for P. gingivalis thiol proteinase in 11 of the 13 patients. Positive staining was mainly located intracellularly in the perinuclear region of the cytoplasm in the periodontal epithelial cells and it could be detected throughout the whole depth of both pocket and oral epithelium. The sensitivity of immunohistochemistry was found to be comparable with that of PCR. CONCLUSIONS: Our results provide in vivo evidence of the ability of P. gingivalis to enter human gingival epithelial cells. Intracellular localization of P. gingivalis contributes to its evasion of the host immune surveillance and eventually increases its resistance to conventional treatments of periodontal diseases.

Due to old age and poor general health, the institutionalized elderly have often not been used as research subjects for the investigation of oral health protocols. Because the number of elderly is increasing, they are an important target group for the development of oral hygiene aids. This trial was designed to investigate whether the institutionalized elderly could successfully use an amine-fluoride- and stannous-fluoride-containing mouthwash and toothpaste during a 12-month period. A dental hygienist visited the subjects weekly, instructed them, and recorded their self-reported compliance. Plaque and saliva samples were taken at three-month intervals. Some salivary biochemical constituents reflecting mucosal integrity were analyzed. The carrier status of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Bacteroides forsythus, Prevotella intermedia, and Prevotella nigrescens was studied by the polymerase chain-reaction method, and acidogenic bacteria and yeasts by dip-slide methods (Orion Diagnostica, Espoo, Finland). Of those 98 subjects originally included, 44 elderly (30 women and 14 men; mean age, 81.7 +/- 5.9 yrs) completed the trial. Their observations were positive or very positive regarding the use of mouthwash and toothpaste. The number of residents with P. gingivalis decreased statistically significantly throughout the trial, while the carrier status of the other periodontal bacteria and mutans streptococci remained on the same level. The number of subjects with high counts (> or = 10(5) cfu/mL) of yeasts decreased during the study: 26.2% at baseline, 9.1% one year later, respectively. Biochemical analyses of salivary immunoglobulins showed no effect on IgA and IgM levels, but IgG reduced significantly. Albumin and other salivary protein concentrations were found to have a decreasing tendency. Thus, the overall effect of the protocol was beneficial with regard to subjects' oral health as measured in this study. The residents were pleased to participate, but the number of drop-outs was very high, confirming the expected difficulties with the utilization of a group of frail elderly as research subjects.

HIV infection reduces oral defensive mechanisms and may affect mucosal integrity. Differences in salivary protein concentrations and periodontopathogenic bacteria were studied in 56 HIV-infected patients with respect to their disease phase. Thirty-three patients were followed up for 2 years. Fifty-three healthy subjects of corresponding age and sex were studied as controls. At baseline, salivary albumin, total protein, IgA, and IgM levels were significantly higher (P

SUMMARY The surface ultrastructure of three anaerobic Gram-positive cocci frequently encountered in oral infections, Peptostreptococcus micros, P. magnus and P. anaerobius, was studied. The type strains of P. micros (DSM 20468) and P. anaerobius (ATCC 27337), several clinical isolates of both species and the type strain of P. magnus (DSM 20470) were included. Thin-sectioned cells studied by electron microscopy revealed a homogeneous layer outside the peptidoglycan layer in P. anaerobius. In P. micros and P. magnus a more amorphous layer was present. No periodic structures were seen in negatively stained whole cells of these three species. However, in freeze-etched cells of P. anaerobius a crystalline surface protein layer (S-layer) was detected. No periodicity was seen in any of the P. micros strains or the magnus type strain by the methods used, but a periodic pattern was observed in negatively stained specimens of cell wall fragments of sonicated P. anaerobius cells. No capsular material was visible outside the S-layer in P. anaerobius. The cells of the Peptostreptococcus spp. were extracted for 30 min with detergents and urea. One er cent SDS and 6 M urea both extracted a major 78 kDa protein from all strains of P. anaerobius. Extraction of P. micros and P. magnus cells did not reveal any major protein bands comparable to that of P. anaerobius. Surface biotinylation of cells followed by Western blotting and detection by alkaline-phosphatase-conjugated extravidin showed strong staining of the 78 kDa band in P. anaerobius, further indicating that this molecule is located on the surface of the cell and is the S-protein. Another protein, of 127 kDa, was also detected by surface biotinylation in all P. anaerobius strains. No differences were detected in colony morphology or cell surface structures between the four strains. Isoelectric focusing of the proteins of P. anaerobius revealed that the S-proteins of individual strains have different pl values, ranging from 5.3 to 6.9.

The aim of the present study was to discover any possible correlation between specific antibodies against Actinobacillus actinomycetemcomitans (A.a.) in serum and saliva. The test group consisted of 38 patients aged 31-68 yr (mean 49) with advanced periodontitis. Twenty-nine subjects aged 23-67 yr, without periodontal destruction, formed a control group with a reference level of specific salivary antibodies against A.a. A subgingival plaque sample for culturing A.a., a specimen of stimulated whole saliva, and a sample of venous blood were taken from each subject of the test group. Specific IgG and IgA antibodies against A.a. were determined from serum and stimulated whole saliva by means of the ELISA test. Fifteen of the patients (39%) had cultivable A.a. Six of the 15 A.a. culture-positive patients and one of the 29 reference subjects exhibited very high antibody titers against A.a. in saliva. Specific IgG and IgA antibodies in saliva correlated highly significantly with the corresponding antibody values in serum among the patients in the test group. It was concluded that among patients with severe adult periodontitis, the less invasive saliva sample has a diagnostic value equal to that of the serum sample concerning specific antibodies against A.a.

Nitrogenase activity (C2H2 reduction) in root-associated Azospirillum lipoferum, Klebsiella pneumoniae, Enterobacter agglomerans and Pseudomonas sp. isolated from roots of Finnish grasses was assayed in the presence of glyphosate, the phenoxy acid herbicides 2-methyl-4-chlorophenoxy acetic acid (MCPA), 2,4-dichlorophenoxy acetic acid (2,4-D), (±)-2-(2-methyl-4-chlorophenoxy)propionic acid (mecoprop) and (±)-2-(2,4-dichlorophenoxy)propionic acid (dichlorprop), and the commercial products Roundup, Nurmikko-Hedonal, Mepro, and Dipro. In the presence of the phenoxy acid herbicides the nitrogenase activity of K. pneumoniae was significantly inhibited, but that of E. agglomerans was stimulated. With the exception of Mepro and mecoprop no phenoxy acid herbicides inhibited the nitrogenase activity of A. lipoferum and none that of Pseudomonas sp. Nurmikko-Hedonal considerably stimulated the nitrogenase activity of E. agglomerans, and Pseudomanas sp. On the other hand, the nitrogenase activity of both K. pneumoniae and E. agglomerans was considerably repressed by glyphosate and Roundup, which also inhibited the growth of the bacteria. These chemicals had no effect on the growth of A. lipoferum and Pseudomonas sp., but stimulated their nitrogenase activity.

A comprehensive view of the diazotrophic bacterial flora of plants requires that attention be paid to the appropriate carbon and oxygen requirements during isolation of the bacteria. Twenty compounds (monosaccharides, disaccharides, polyols, and organic acids) were therefore examined as carbon and energy sources for nitrogenase activity in semisolid stab cultures at pO 2 values of 0.21, 0.02, and ≤0.002 with 12 strains of diazotrophic root-associated bacteria. With the facultatively anaerobic bacteria of the genera Klebsiella and Enterobacter , the best substrate was sucrose, followed by fructose and mannitol, whereas among the organic acids, only malic and fumaric acids supported any activity. With the obligately aerobic bacteria of the genera Azospirillum and Pseudomonas , disaccharides were not utilized for nitrogen fixation, but several organic acids were accepted in addition to monosaccharides and polyols; malate and glucose were the best substrates. The patterns of the carbon sources utilized for nitrogen fixation were coherent within the species, with the exception of one Klebsiella pneumoniae and one Enterobacter agglomerans strain, both isolated from the same individual grass plant, which were unable to utilize lactose. Anaerobic conditions (pO 2 value of ≤0.002) were required for maximum nitrogenase activity with the facultatively anaerobic bacteria, with the exception of one strain of E. agglomerans , which required atmospheric oxygen (pO 2 value of 0.21). Also, the obligately aerobic diazotrophs required atmospheric oxygen for maximum nitrogenase activity. The maximum specific nitrogenase activities (expressed as micromoles of C 2 H 4 · milligram of bacterial protein −1 · hour −1 ) noted during the exponential growth phase of the bacteria were the following: 2.68 with Azospirillum lipoferum on malate, 2.41 with K. pneumoniae and 1.58 with E. agglomerans on sucrose, and 0.95 with Pseudomonas sp. on malate.

Our previous work showed that strains of dinitrogen fixing enterobacter and Klebsiella were found associated with the roots of uncultivated grasses in Finland more commonly than other species of diazotrophic bacteria. In this paper we compare E. agglomerans strains to K. pneumoniae and K. terrigena strains, and show that the E. agglomerans strains fall into two biogroups. The groups differ not only in the utilization of different carbon sources and other physiological characteristics such as the production of indole, but also in the physiology and genetics of nitrogenase activity. Biotype 1 (isolated from Achillea millefolium, Calamagrostis arundinacea, and Phleum pratense) showed active nitrogenase in atmospheric oxygen, whereas biotype 2 (from Phalaris arundinacea) resembled K. pneumoniae in that it was active at reduced oxygen pressure (pO2

The composition of subgingival flora was correlated with clinical periodontal conditions in 100 teenagers aged 12 to 17 years. The Community Periodontal Index of Treatment Needs (CPITN) was used for the clinical examination. Subgingival bacterial samples were taken from the mesial surface of each first molar, two samples for dark-field microscopy and two samples for Actinobacillus actinomycetemcomitans (A.a.) cultivation. Fifty-nine subjects had at least three healthy sextants. Score 1 was the highest CPITN recording in 61 subjects and Score 2 in 30 subjects. None had scores 3 or 4. In dark-field microscopy, cocci predominated in most samples. Straight rods, fusiforms and motile rods correlated negatively to the number of healthy sextants per subject. Straight rods and fusiforms showed a positive correlation to gingival bleeding tendency at the sampled site. A.a. was isolated in four subjects. Motile microorganisms and A.a. were detected rarely in subjects with good periodontal conditions.

20 patients with juvenile periodontitis (JP) were treated with oral hygiene instruction, scaling and root planing, possibly with flap surgery but with no antibiotics. The patients were monitored after 6 to 12 years. Re-examination revealed that no probing depth of 7 mm or more existed any longer, and that sites with probing depth of 4 to 6 mm had decreased from 237 to 46. The bone loss scores had changed from 18% (range 2.0 to 48.1) to 14% (range 0 to 44.4) and the bleeding on probing scores from 39% (range 0 to 100) to 10% (range 0 to 40). Actinobacillus actinomycetemcomitans (A.a.) had neither been cultivated nor serologically tested at the initial examination. At the re-examination, it was found in 2 patients out of 20 at 5 sites. 7 patients, the 2 with positive cultures included, had elevated titers to A.a. strain Y4 in whole or parotid saliva or both. It is concluded that there is a marked improvement in the periodontal condition of these patients, and that good periodontal health in patients with JP can be reached without antibiotics.

The technique of15N isotope dilution was used to verify that nitrogen was fixed and transferred to the plant byKlebsiella pneumoniae strain Pp in association withPoa pratensis orTriticum aestivum. Surface sterilized, sprouting seeds were inoculated withK. pneumoniae and grown in sand in modified Leonard jars. Potassium nitrate enriched with15N was used to provide N concentrations ranging from 10–40 mg Nl−1 nutrient solution. After 10–18 weeks the shoots and roots were analyzed separately for dry matter, N content, total N, and atom %15N excess. The acetylene reduction technique was used to test for the presence of N2-fixing organisms on the roots. The data from15N isotope dilution demonstrated that up to 33.8% of N in the shoots ofP. pratensis and 15.9% in those ofT. aestivum were derived from associative N2 fixation byK. pneumoniae. In most experiments the dry matter yield, N content, and total N yield of the shoots ofP. pratensis were increased byK. pneumoniae inoculation, whereas inoculation had no significant effect on the dry matter yield, N content or total N of the shoots ofT. aestivum.