Characterisation and in vitro and in vivo evaluation of supercritical-CO2-foamed ß-TCP/PLCL composites for bone applications.

Most synthetic bone grafts are either hard and brittle ceramics or paste-like materials that differ in applicability from the gold standard autologous bone graft, which restricts their widespread use. Therefore, the aim of the study was to develop an elastic, highly porous and biodegradable ß-tricalciumphosphate/poly(L-lactide-co-ε-caprolactone) (ß-TCP/PLCL) composite for bone applications using supercritical CO2 foaming. Ability to support osteogenic differentiation was tested in human adipose stem cell (hASC) culture for 21 d. Biocompatibility was evaluated for 24 weeks in a rabbit femur-defect model. Foamed composites had a high ceramic content (50 wt%) and porosity (65-67 %). After 50 % compression, in an aqueous environment at 37 °C, tested samples returned to 95 % of their original height. Hydrolytic degradation of ß-TCP/PLCL composite, during the 24-week follow-up, was very similar to that of porous PLCL scaffold both in vitro and in vivo. Osteogenic differentiation of hASCs was demonstrated by alkaline phosphatase activity analysis, alizarin red staining, soluble collagen analysis, immunocytochemical staining and qRT-PCR. In vitro, hASCs formed a pronounced mineralised collagen matrix. A rabbit femur defect model confirmed biocompatibility of the composite. According to histological Masson-Goldner's trichrome staining and micro-computed tomography, ß-TCP/PLCL composite did not elicit infection, formation of fibrous capsule or cysts. Finally, native bone tissue at 4 weeks was already able to grow on and in the ß-TCP/PLCL composite. The elastic and highly porous ß-TCP/PLCL composite is a promising bone substitute because it is osteoconductive and easy-to-use and mould intraoperatively.

Lack of serotype antigen in A. actinomycetemcomitans.

Aggregatibacter actinomycetemcomitans is divided into 6 serotypes. Occurrence of non-serotypeable strains is known, but background reasons are unclear. We hypothesized that non-serotypeable strains represent new serotypes or have altered expression of serotype-specific polysaccharide antigen (S-PA). We first characterized 311 strains from 189 individuals using both immunoassay- and PCR-based serotyping. Next, using natural human infection and rabbit immunization approaches, we clarified whether the phenotypically non-serotypeable strains expressed S-PA. Immunoassay identified serotypes a-f among 216 strains from 159 individuals. The remaining 95 strains from 30 individuals were phenotypically non-serotypeable. Yet, all these strains were identified by PCR-typing as serotype a-, b-, c-, or f. Non-serotypeability was confirmed by Western immunoblot with respective rabbit antisera. Patient sera remained non-reactive with autologous non-serotypeable strains at the serotype-specific region. Rabbit immunization with a phenotypically non-serotypeable strain induced no antibody production against S-PA. Thus, phenotypically non-serotypeable strains did not include novel serotypes, but lacked S-PA expression.

Lymphotoxin alpha LTA+496C allele is a risk factor for periodontitis in patients with coronary artery disease.

Periodontitis and coronary artery disease (CAD) are inflammatory diseases and associated with each other. The major histocompatibility complex (MHC) region carries genes involved in immune response and inflammation. We investigated whether the MHC genes correlate with the presence of periodontitis or with the occurrence of periodontal pathogens in patients with CAD. Blood and saliva samples from CAD patients (n = 106) were collected at the time of hospitalization. Nine MHC genetic markers [human leukocyte antigen (HLA)-A, HLA-B, HLA-DRB1, lymphotoxin alpha (LTA) +253(a/g), +496(C/T), +633(c/g), +724(C/A), C4A and C4B)] were typed. Based on panoramic tomography, patients were categorized into nonperiodontitis and periodontitis groups. Two major periodontal pathogens, Aggregatibacter (Actinobacillus) actinomycetemcomitans and Porphyromonas gingivalis, were cultivated and polymerase chain reaction-amplified from salivary samples. Serum immunoglobulin (Ig)A and IgG antibody levels to these pathogens were measured. In the univariate analysis, LTA+496C allele (OR = 5.29; 95% CI = 2.07-13.51, P = 0.00027), and the occurrence of P. gingivalis in saliva (OR = 4.74; 95% CI = 1.64-13.70; P = 0.002) were more frequent in periodontitis when compared with nonperiodontitis. Similarly, serum IgA antibody level against the pathogen was increased in periodontitis (P = 0.048). In the multiple logistic regression analysis, when a wide range of covariates was included, the LTA+496C allele (OR = 10.87; 95% CI = 3.23-36.60; P = 0.00012) and the elevated serum IgA antibody level against P. gingivalis (OR = 1.56; 95% CI = 1.05-2.30; P = 0.026) remained as significant risk factors for periodontitis. In conclusion, the major finding of this study is that the LTA+496C allele is associated with periodontitis in patients with CAD.

Complement and c4 null alleles in severe chronic adult periodontitis.

Severe forms of chronic periodontitis affect up to 10% of adults. Tumour necrosis factor and lymphotoxin-alpha genes in the major histocompatibility complex are associated with severe periodontitis. Complement factor C4 is a nearby, polymorphic, functionally relevant gene region. Although associated with chronic mucosal infections, C4 deficiencies have not been assessed in adult periodontitis patients. We tested whether complement levels are systemically altered and C4 deficiencies associated with severe chronic periodontitis. In a case-control study, we analysed levels of plasma C3, and C4, serum classical pathway haemolytic activity, C4 allotypes and C4 gene numbers in 37 patients with severe chronic periodontitis and in 150 voluntary controls. Plasma levels of C3 were higher, and classical pathway haemolytic activity was lower in patients than in controls. Partial C4 gene deficiencies were more frequent in patients than in controls (odds ratio 2.4, 95% confidence interval 1.1-5.5, P = 0.032). Changes in complement levels may reflect chronic, recurring inflammation. C4 gene deficiencies are associated with predisposition to chronic periodontitis.

Clonal distribution of natural competence in Actinobacillus actinomycetemcomitans.

The competence for natural transformation was investigated in 67 Actinobacillus actinomycetemcomitans strains. The transformation assays were performed with both cloned DNA fragments and chromosomal markers of A. actinomycetemcomitans. Competence was found in 12 of 18 serotype a strains, 0 of 21 serotype b strains, 0 of 14 serotype c strains, 3 of 6 serotype d strains, 3 of 4 serotype e strains, 0 of 3 serotype f strains, and 0 of 1 nonserotypeable strain. The transformation frequencies varied from 5 x 10(-3) to 4 x 10(-6) (median 1.5 x 10(-4)). The distribution pattern of natural competence is concordant with the major clonal lineages of A. actinomycetemcomitans. Serotype a strains are predominantly competent for transformation, while serotypes b and c strains are apparently non-competent.

Relationship of Actinobacillus actinomycetemcomitans serotype b to aggressive periodontitis: frequency in pure cultured isolates.

BACKGROUND: To our knowledge, the association of the five serotypes of Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans) to the new diagnostic classification scheme defined by the American Academy of Periodontology in 1999 has not yet been described. The goal of this study was to characterize the frequencies of the five serotypes of A. actinomycetemcomitans in A. actinomycetemcomitans isolates from various forms of periodontitis using both old and new diagnostic classifications and to determine the relationships between serotype and age and clinical diagnosis. METHODS: A total of 345 A. actinomycetemcomitans isolates from 115 A. actinomycetemcomitans culture-positive subjects (mean age 38.0 +/- 18.3 years, 59% female) were collected. Based on the new classifications, 33 subjects had aggressive periodontitis and 82 chronic periodontitis. According to old classifications, there were six prepubertal periodontitis (PPP), 12 localized juvenile periodontitis (LJP), 15 post-localized juvenile periodontitis (PLJP), 28 refractory periodontitis (Ref-P), and 54 adult periodontitis (AP) cases. Serotypes of A. actinomycetemcomitans were determined by an indirect immunofluorescence assay using serotype-specific polyclonal antisera to A. actinomycetemcomitans strains ATCC 29523, ATCC 43728, ATCC 33384, IDH 781 and IDH 1705 (serotype a, b, c, d, and e, respectively). Proportions of serotype b were examined between different diagnostic and age groups with a Z-test for proportions. RESULTS: Most subjects (n = 100, 86.96%) were infected with a single serotype (22 serotype a, 44 serotype b, 30 serotype c, 1 serotype d, and 3 serotype e). There were 11 subjects (9.57%) with two serotypes and two subjects (1.74%) with 3 serotypes. Two individuals had isolates lacking any detectable serotype antigen. Serotype b was the predominant serotype in children under 18 years of age and young adults between 19 to 35 years, although serotype b status was not significantly associated with age. Serotypes d and e were not found in patients under 35 years old. In 62 adult patients, one subject had serotype d and three had serotype e. Serotype b was the most common serotype in aggressive periodontitis (60.61%). The proportion of cases with serotype b was significantly higher in aggressive periodontitis compared to chronic periodontitis (P = 0.031). Other serotypes were not significantly associated with new diagnostic categories. Serotypes d and e were not detected in aggressive periodontitis. CONCLUSION: The results of this study show that proportions of serotype b of A. actinomycetemcomitans are significantly greater in culture-positive patients with aggressive periodontitis than those with chronic periodontitis.

Multiserotype enzyme-linked immunosorbent assay as a diagnostic aid for periodontitis in large-scale studies.

Periodontitis is a common chronic oral infection caused by gram-negative bacteria, including Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis. Periodontitis evokes inflammatory host response locally in the periodontium but also systemically. The systemic humoral antibody response against oral pathogens can conveniently be measured by an immunoassay. The aim of the study was to measure serum immunoglobulin G class antibodies against A. actinomycetemcomitans and P. gingivalis by an enzyme-linked immunosorbent assay (ELISA) in which mixtures of several serotypes of the pathogens were used as antigens to avoid biasing of the results in favor of a particular strain. For A. actinomycetemcomitans the antigen consisted of six strains representing serotypes a, b, c, d, and e and one nonserotypeable strain. In the P. gingivalis ELISA, antigens representing serotypes a, b, and c were used. Serum samples from 90 subjects, including 35 samples from patients with diagnosed periodontitis, 10 samples from periodontally healthy controls, and 45 samples from randomly selected apparently healthy volunteers (referred to as "healthy subjects"), were tested. For both pathogens the antibody levels (means +/- standard deviations) of the patients--xpressed as area under the dilution curve--were significantly higher than those for healthy controls or healthy subjects, with values for A. actinomycetemcomitans and P. gingivalis, respectively, as follows: patients, 22.60 +/- 9.94 mm(2) and 26.72 +/- 11.13 mm(2); healthy controls, 9.99 +/- 3.92 mm(2) and 6.90 +/- 3.38 mm(2); and healthy subjects, 16.85 +/- 6.67 mm(2) and 8.51 +/- 4.23 mm(2). The serotype mixture ELISA is suitable for measuring antibodies against periodontal pathogens in large epidemiological studies in order to evaluate the role of periodontitis as a risk factor for other diseases.

Subgingival strains of Candida albicans in relation to geographical origin and occurrence of periodontal pathogenic bacteria.

Clonal diversity of subgingival yeast strains was determined in relation to geographical location and coexistence of selected periodontal pathogenic bacteria. A total of 60 dental patients from Finland, the United States and Turkey each contributed five Candida albicans isolates. C. albicans isolates were serotyped using slide agglutination and genotyped using polymerase chain reaction (PCR) amplification and a random sequence primer. In general, each study subject yielded C. albicans isolates belonging to the same serotype and genotype. C. albicans serotype A occurred more frequently in subjects from Finland and Turkey than in subjects from the United States. A total of 27 PCR-based C. albicans genotypes were identified. One C. albicans genotype occurred with particularly high frequency in subjects from Turkey and another genotype in subjects from the United States. Relationships were identified between C. albicans serotypes and genotypes. Further studies are needed to determine environmental factors of importance for subgingival colonization and persistence of C. albicans.

Neutrophil free oxygen radical production and blood total antioxidant capacity in patients with coronary heart disease using various medications.

Despite convincing results of studies in vitro, less is known about the effects of antioxidants on in vivo redox balance in humans. We developed a novel parameter of in vivo redox balance, and studied it and its relation to dental infections in 51 patients on medication for coronary heart disease (CHD) and 39 random controls matched for age group, sex, social class and locality. In vivo redox balance was the ratio of plasma antioxidant capacity, as measured with radical-trapping assay, to neutrophil respiratory burst capacity, as measured with whole blood chemiluminescence assay. Dental infections were quantitated with four rating scales. CHD patients had higher values than controls. Patients on acetosalicylic acid (ASA), diuretics or beta blockers, but not the ones on calcium channel blocker, had significantly higher redox balance than non-users. Combination of calcium channel blockers and ASA was associated with redox balance similar to taking beta blockers or diuretics. Diuretics and ASA were independent determinants of redox balance in multivariate analyses. Redox balance did not correlate with severity of dental infections (Spearman's r 0.06 to 0.11). The results contrast experimental data indicating that calcium channel blockers are as antioxidants superior to other cardiovascular drugs. Total antioxidant capacity in parallel with oxygen species production capacity should be considered in attempts to solve the antioxidant paradox.

Age, dental infections, and coronary heart disease.

Epidemiological and intervention studies have suggested that infections are risk factors for coronary heart disease (CHD). Dental infections have appeared as cardiovascular risk factors in cross-sectional and in follow-up studies, and the association has been independent of the "classic" coronary risk factors. This case-control study aimed at detailed assessment of the dental pathology found in various CHD categories (including elderly patients). Altogether, 85 patients with proven coronary heart disease and 53 random controls, matched for sex, age, geographic area, and socio-economic status, were compared with regard to dental status, assessed blindly with four separate scores, and to the "classic" coronary risk factors (seven of the controls had CHD, and they were not included in the analyses). The dental indices were higher among CHD patients than in the controls, but, contrary to previous studies, the differences were not significant (between the CHD patients and their matched controls or among the different CHD categories). This result could not be explained by potential confounding factors. The participants in the present study were older and had more often undergone recent dental treatment in comparison with subjects in our earlier studies. Age correlated with the severity of dental infections only in the random controls but not in the coronary patients who, although young, already had high dental scores. We believe that the higher age of the participants in the present study is the most likely reason for the results. Other possible explanations include an age-related selection bias among older CHD patients, and the fact that those participating in studies like this may have better general health and thus also less severe dental infections. Thus, the role of dental infections as a coronary risk factor varies according to the characteristics of the population studied.

Localization of heat shock proteins in clinical Actinobacillus actinomycetemcomitans strains and their effects on epithelial cell proliferation.

Actinobacillus actinomycetemcomitans is an important pathogen in periodontitis. In the present study we localized the GroEL- and DnaK-like heat shock proteins (Hsp) in subcellular fractions of 12 A. actinomycetemcomitans strains of various clinical origin and compared their effects on periodontal epithelial cell proliferation and viability. In all strains, GroEL-like protein was found in the membrane, cytoplasm, and periplasm, whereas DnaK-like protein was present in the cytoplasm and periplasm. No correlation was observed between the Hsp expression and the serotype or origin of A. actinomycetemcomitans strains. The bacterial membrane fractions that expressed the GroEL-like protein moderately or strongly induced epithelial cell proliferation more strongly than strains that expressed the protein weakly. The results suggest that GroEL-like Hsp may play a role in the virulence of A. actinomycetemcomitans by increasing epithelial proliferation.

Altered antigenicity is seen in the lipopolysaccharide profile of non-serotypeable Actinobacillus actinomycetemcomitans strains.

Non-serotypeable Actinobacillus actinomycetemcomitans strains may be derived from the serotypeable ones. In the present study, we compared the outer membrane proteins (OMPs) and lipopolysaccharides (LPSs) of serotypeable and non-serotypeable A. actinomycetemcomitans strains (n=24) of the same genotype in the same subject (n=6) to find out if alterations on the cell-surface contribute to the non-serotypeability. Serotypeable and non-serotypeable A. actinomycetemcomitans strains showed great similarity in the OMP patterns both within and between subjects. Using serotype-specific antisera, clear immunoblotting LPS profiles in the O-antigenic region were seen in serotype b and c strains but not in non-serotypeable strains from the same subjects. The results suggest that changes in LPS lead to the altered antigenicity of non-serotypeable A. actinomycetemcomitans strains.

Heterogeneity of Actinobacillus actinomycetemcomitans strains in various human infections and relationships between serotype, genotype, and antimicrobial susceptibility.

Actinobacillus actinomycetemcomitans, an oral pathogen, only occasionally causes nonoral infections. In this study 52 A. actinomycetemcomitans strains from 51 subjects with nonoral infections were serotyped and genotyped by arbitrarily primed PCR (AP-PCR) to determine whether a certain clone(s) is specifically associated with nonoral infections or particular in vitro antimicrobial susceptibility patterns. The promoter structure of leukotoxin genes was additionally investigated to find the deletion characteristic of highly leukotoxic A. actinomycetemcomitans strains. The nonoral A. actinomycetemcomitans strains included all five known serotypes and nonserotypeable strains, the most common serotypes being b (40%) and c (31%). AP-PCR distinguished 10 different genotypes. A. actinomycetemcomitans serotype b strains were more frequently found in blood samples of patients with bacteremia or endocarditis than in patients with focal infections. One AP-PCR genotype was significantly more frequently found among strains originating in focal infections than in blood samples. Resistance to benzylpenicillin was significantly more frequent among A. actinomycetemcomitans serotype b strains than among strains of other serotypes. No differences in the leukotoxin gene promoter region or benzylpenicillin resistance between nonoral and oral A. actinomycetemcomitans strains were observed. Nonoral A. actinomycetemcomitans strains showed great similarity to the oral strains, confirming that the oral cavity is the likely source of nonoral A. actinomycetemcomitans infections. The predominance of serotype b strains in endocarditis and bacteremia supports the hypothesis of a relationship between certain A. actinomycetemcomitans clones and some nonoral infections. The mechanisms behind the exceptionally high rate of occurrence of benzylpenicillin resistance among A. actinomycetemcomitans serotype b strains are to be elucidated in further studies.

Comparison of virulence factors of oral Candida dubliniensis and Candida albicans isolates in healthy people and patients with chronic candidosis.

We determined differences in the expression of certain virulence factors between oral Candida dubliniensis and Candida albicans species. In addition, clonal differences were sought among C. albicans isolates recovered from patients with and without compromised immune system. The material comprised 93 clinical yeast isolates originated in 40 subjects (1-5 isolates per subject). All 26 C. dubliniensis isolates and 46 C. albicans isolates originated from healthy routine dental clinic patients. Additionally, 21 C. albicans isolates were collected from patients with autoimmune polyendocrinopathy-candidosis-ectodermal dystrophy (APECED), who have chronic candidosis as one manifestation of their immunocompromising disease. Polymerase chain reaction amplification using the random sequence primer OPE-03 enabled grouping of the C. dubliniensis isolates in 2 genotypes (I and II) and C. albicans isolates in 15 genotypes (I-XV). No significant difference was found in the distribution of genotypes between the patients with APECED and the healthy subjects. C. dubliniensis isolates exhibited high-frequency phenotypic switching significantly more frequently than did C. albicans isolates, and vice versa regarding phospholipase and proteinase production. Proteinase production was significantly more frequent among C. albicans genotype V than genotype IX isolates. No significant difference was found in expression of virulence factors of C. albicans isolates between the patients with APECED and the healthy subjects.

The Prevotella intermedia group organisms in young children and their mothers as related to maternal periodontal status.

Currently, the Prevotella intermedia group includes three biochemically and phylogenetically related species: Prevotella intermedia, Prevotella nigrescens, and the newly described Prevotella pallens. The two first-named species are mentioned with varying emphasis in connection with periodontal diseases, while such a connection of P. pallens is not known. Mothers serve as a plausible source of bacteria to their children, and conceivably, a mother with periodontitis as a recurrent reservoir of periodontally infecting organisms. In the present study, 23 mothers and their young children were examined for the presence of the P. intermedia group organisms in relation to maternal periodontal status (I: periodontal health, II: initial periodontitis, and III: advanced periodontitis). Species differentiation was based on established biochemical methods, electrophoretic mobility patterns, SDS-PAGE, and DNA hybridization. P. intermedia was not recovered from children but nearly exclusively from mothers in group III, thus confirming its association with periodontitis. P. nigrescens and P. pallens were frequently found in mothers and children. To determine bacterial transmission between a mother and her child, 72 isolates from 13 mother-child pairs were analyzed by arbitrarily primed PCR (AP-PCR). Similar AP-PCR types of P. nigrescens and/or P. pallens were recovered from 3/4 pairs in group I, 2/5 pairs in group II, and none in group III. Our results indicate that different species within the P. intermedia group have a different colonization pattern in childhood and that the periodontal status reflects qualitatively their presence in maternal saliva. Intra-familial transmission of P. nigrescens and P. pallens can occur in early childhood, however similar AP-PCR types were most obvious within periodontally healthy mother-child pairs.

Bacterial adhesion of Actinobacillus actinomycetemcomitans serotypes to titanium implants: SEM evaluation. A preliminary report.

BACKGROUND: In this study, the adherence ability of Actinobacillus actinomycetemcomitans serotypes to titanium implant surfaces was evaluated to demonstrate if any selective adherence occurs according to the serotypes of the microorganism. METHODS: The study material included 3 reference strains of A. actinomycetemcomitans serotypes a, b, and c (ATCC 29523, ATCC 43718, ATCC 33384) and 2 clinical isolates of A. actinomycetemcomitans serotypes d and e (IDH 781, IDH 1705), together with commercially available titanium blade implants. For each strain, bacterial suspensions with identical concentrations (5 x 10(7) cells/ml) were prepared and 0.5 ml of each was added on to the implant surfaces, which had been precoated with glycine-bovine serum albumin (BSA). After incubation at 37 degrees C for 60 minutes in 5% CO2 in air, the implants with attached bacteria were prepared for scanning electron microscopic (SEM) observations. Bacterial adhesion was quantified on the textured body surfaces of the implants, and results were statistically analyzed with analysis of variance followed by Duncan's test. The surface ultrastructure of the bacterial cells was also evaluated descriptively. RESULTS: The tested strains adhered to implant surfaces in different quantities. Serotype a (ATCC 29523) showed the highest adherence affinity (statistically significant, P <0.01). When compared with each other, serotypes b, c, and d (ATCC 43718, ATCC 33384, and IDH 781) attached equally well, whereas serotype e (IDH 1705) had a statistically significant low adherence capability. CONCLUSIONS: It is suggested that in vitro A. actinomycetemcomitans adhesion to implant surfaces is strain dependent.

Oral ecology and person-to-person transmission of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis.

The ecological characteristics of the oral cavity are dissimilar for A. actinomycetemcomitans and for P. gingivalis, as judged by differences in their colonization preferences and patterns, associations with periodontal disease parameters, relationships with the subgingival microbiota and the type of periodontitis and their clonal persistence in the oral cavity. These features also suggest that as a periodontal pathogen, A. actinomycetemcomitans is different from P. gingivalis. Probably in most infected individuals, low levels of A. actinomycetemcomitans can persist for years in equilibrium with the host and the resident oral microbiota. However, it is well established that A. actinomycetemcomitans can cause disease in some individuals or in some circumstances when the regulatory mechanisms are unable to maintain homeostasis in the ecosystem. Elevated A. actinomycetemcomitans proportions of the biota can be regarded as a sign of ecological imbalance, leading to increased risk of periodontal destruction. There is also evidence showing elevated pathogenic potential of certain A. actinomycetemcomitans clones. Although A. actinomycetemcomitans seems to be relatively rarely transmitted between cohabiting adults, transmission can occur to periodontally healthy children of A. actinomycetemcomitans-positive parents. Parents and children may share factors that promote successful oral colonization of A. actinomycetemcomitans, or the window of opportunity is in childhood. Therefore, to prevent parent-child transmission of A. actinomycetemcomitans, bacterium-positive parents of young children are optimal targets for enhanced information and treatment. In selected populations, screening for specific clones of A. actinomycetemcomitans has been employed in prevention of peridontitis. Future research aiming at finding the reasons which cause the changes in the oral homeostasis to allow the growth of A. actinomycetemcomitans may give insight into novel prevention strategies for A. actinomycetemcomitans-associated periodontitis. Compared with A. actinomycetemcomitans, P. gingivalis shows a different pattern of coexistence with the host. In periodontal health or in children, P. gingivalis is absent or only rarely detected. When present, P. gingivalis is commonly recovered in high numbers from dentitions exhibiting inflamed periodontitis and poor oral hygiene. Contrary to A. actinomycetemcomitans, the data on the vertical transmission of P. gingivalis are limited. The major infection route of P. gingivalis seems to be between adults, indicating that P. gingivalis commonly colonizes in an established oral microbiota. These characteristics suggest that the degree of tolerance between P. gingivalis and the host is inferior to that between A. actinomycetemcomitans and the host. It appears that the association of P. gingivalis with disease is a rule rather than an accidental incident. On these grounds, it seems that the host-P. gingivalis relationship approaches antibiosis. Since P. gingivalis infection is related to a typical periodontal eco-pathology, the susceptibility to person-to-person transmission of this pathogen may be controlled by periodontal treatment and emphasizing the significance of high standard oral hygiene.

Beta-lactamase production in Prevotella intermedia, Prevotella nigrescens, and Prevotella pallens genotypes and in vitro susceptibilities to selected antimicrobial agents.

The present study investigated the beta-lactamase production of 73 Prevotella intermedia, 84 Prevotella nigrescens, and 14 Prevotella pallens isolates and their in vitro susceptibilities to six antimicrobial agents. The P. intermedia and P. nigrescens isolates were recovered from oral and extraoral samples obtained from subjects in two geographic locations from 1985 to 1995. The clonality of the beta-lactamase-positive and beta-lactamase-negative isolates and the clustering of the genotypes were studied by arbitrarily primed-PCR fingerprinting. beta-Lactamase production was detected in 29% of P. intermedia isolates, 29% of P. nigrescens isolates, and 57% of P. pallens isolates. No difference in the frequencies of beta-lactamase production by P. intermedia and P. nigrescens between isolates from oral and extraoral sites, between isolates obtained at different time periods, or between P. intermedia isolates from different geographic locations was observed. However, the P. nigrescens isolates from the United States were significantly more frequently (P = 0.015) beta-lactamase positive than those from Finland. No association between the genotypes and beta-lactamase production or between the genotypes and the sources of the isolates was found. The penicillin G MICs at which 90% of the isolates were inhibited were 8 microg/ml for P. intermedia, 8 microg/ml for P. nigrescens, and 16 microg/ml for P. pallens. For the beta-lactamase-negative isolates, the corresponding values were 0.031, 0.031, and 0.125 microg/ml, and for the beta-lactamase-positive isolates, the corresponding values were 16, 8, and 32 microg/ml. All isolates were susceptible to amoxicillin-clavulanate, cefoxitin, metronidazole, azithromycin, and trovafloxacin. The MICs of amoxicillin-clavulanate and cefoxitin were relatively higher for the beta-lactamase-positive population than for the beta-lactamase-negative population.

Establishment of oral anaerobes during the first year of life.

Anaerobic species constitute a significant part of the bacterial community of the mouth. Although the time and species involved in the primary colonization of infants are of great importance by forming the basis for further colonization, the development of the oral anaerobic microflora with age is still inadequately understood. In the present study, time and succession of colonization of oral anaerobes were longitudinally examined in 44 healthy Caucasian infants at 2, 6, and 12 months of age. Unstimulated saliva samples were quantitatively cultured on non-selective Brucella blood agar and several selective media for the isolation of anaerobic micro-organisms. The most frequent anaerobic finding in two-month-old infants was Veillonella spp. The Prevotella melaninogenica group also represented early colonizing species, and the frequency increased remarkably during the first year of life, whereas the Prevotella intermedia group organisms seemed to be late colonizers. Fusobacterium nucleatum, non-pigmented Prevotella spp., and Porphyromonas catoniae were occasional findings in subjects at 2 months but frequent findings in those at one year of age. F. nucleatum was the most frequent strictly anaerobic species in one-year-old infants; other fusobacteria were also occasionally found. The frequency of facultative/micro-aerophilic corroding rods and Capnocytophaga spp. started to increase toward the end of the first year. Except for the common presence of facultative/micro-aerophilic Actinomyces spp., other anaerobic gram-positive species were only occasionally present in these infants. Once established, early-colonizing species tended to persist in the mouth. Our longitudinal study demonstrated the establishment of several anaerobic species with steadily increasing frequencies during the first year of life.

Age-related acquisition of oral and nasopharyngeal yeast species and stability of colonization in young children.

The occurrence and stability of colonization of oral yeast species and strains was determined from 40 healthy children during a 22-month follow-up at the ages 2, 6, 12, 18 and 24 months. In addition, salivary samples were obtained from the mothers at baseline (2 months) to study the role of the mother as the source of yeasts for the child. Yeasts were recovered at least once from 17/40 (43%) children by the age of 2 years. Of the 40 children, 11 (28%) were yeast-positive at multiple sampling occasions. No significant differences were found in recovery frequency of yeasts at different ages. Candida parapsilosis was isolated in 18/33 (55%) yeast-positive samples, and it predominated (share of positive findings 76%) at ages 12 to 24 months. The same yeast species was rarely detected in successive follow-up samples and thus on species level yeasts were transient colonizers in the developing oral flora of the children. Of the mothers 20/40 (50%) harbored yeasts. Candida albicans was recovered from 19/20 (95%) of the yeast-positive mothers and C. parapsilosis from none. Only 7/20 (35%) of the mothers with a yeast-positive finding had a yeast-positive child. In 5/7 (71%) of these mother-child pairs, both harbored the same yeast species (C. albicans) and in 3/5 (60%) of the pairs the AP-PCR profiles of the yeast isolates were identical suggesting possible transmission. In children, significant relationships (Fisher's exact-test, P < 0.05) were found between recovery of yeasts and use of pacifier at age over 12 months, eruption of first teeth at age over 6 months, mother cooling the child's food by blowing and mother cleaning the child's pacifier in her own mouth. In mothers, a significant relationship existed between recovery of yeasts and use of antibiotics.