When colonies of encapsulated isolates of Streptococcus pneumoniae are viewed with oblique, transmitted light on a transparent surface, they are heterogeneous in appearance because of variation in opacity. There is spontaneous phase variation among at least three discernible phenotypes at frequencies from 10-3 to 10-6. The ability to detect differences in opacity varies according to serotype, but variation is independent of capsule expression. Electron microscopy shows no difference in chain length but suggests that autolysis occurs earlier in the growth of the transparent variant. There was no identifiable difference in membrane protein profiles of opaque and transparent variants of the same strain. In an infant rat model of nasopharyngeal carriage, there was no significant colonization by opaque variants. Efficient and stable colonization by the transparent variants was observed, suggesting a selective advantage for this phenotype in the nasopharynx. In contrast, there was no difference in the incidence of bacteremia or in the 50% lethal dose among the variants following their intraperitoneal inoculation. These results suggest that phase variation which is marked by differences in colonial morphology may provide insight into the interaction of the pneumococcus with its host.

Aims: To validate perfused, inline, filter-based fermentation systems (multiple Sorbarod devices, MSD) for their ability to maintain stable oral bacterial communities. MSD enable replicate (n ¼ 5) microcosm biofilms (BF) to be established and sampled, together with their perfusates (PA, cells in eluted medium). Methods and Results: Fresh saliva from human volunteers was used to inoculate MSD, incubated in an anaerobic cabinet and perfused with artificial saliva at 7 ml h )1 . BF within Sorbarod filters and cells eluted in the PA were analysed at 24-h intervals by differential bacteriological culture and checkerboard DNA-DNA hybridization (CKB, 40 oral species). Dynamic stability was apparent after 2-3 days within both BF and PA as evidenced by culture, CKB data and pH measurements. BF harboured large numbers of anaerobic species and facultative anaerobes [ca 10-11 log 10 colony-forming units (CFU)/filter] comprising considerable numbers of streptococci and Gramnegative species. PA contained ca 9-10 log 10 CFU ml )1 suggesting an apparent mean growth rate of 0AE1 h )1 for the BF, as a whole corresponding to a mean generation time of 10 h. CKB analysis revealed considerable bacterial diversity within the respective MSD. Inter-individual variations in the relative species abundance of inocula was broadly reproduced in the MSD (BF and PA), although considerable variation was apparent between triplicate models established using saliva from one saliva donor or from three individual donors. The dominance of Gramnegative species, indicated by culture was supported by CKB analysis (major species, Prevotella melaninogenica and Fusobacterium nucleatum). Conclusions: Data obtained from the various analytical approaches showed a high degree of congruence. The MSD enables the maintenance of complex, stable salivary microcosms and represents a simple, reproducible tool for modelling individual oral bacterial ecosystems. Significance and Impact of the Study: This study demonstrates the utility of the MSD for studying the microecology of the oral cavity.

Actinobacillus actinomycetemcomitans, an oral bacterial species associated with periodontal disease, was found to invade human cell lines. Invasion was demonstrated by recovery of viable organisms from gentamicin-treated KB cell monolayers and by light and electron microscopy. Internalization occurred through a cytochalasin D-sensitive process. Invasion efficiencies of some A. actinomycetemcomitans strains were comparable to those of invasive members of the family Enterobacteriaceae. Differences in invasiveness were correlated with bacterial colonial morphology. Smooth variants invaded more proficiently than rough variants. A. actinomycetemcomitans can undergo a smooth-to-rough colonial morphology shift which results in the loss of invasiveness. Coordinated regulation of genes involved in the rough-to-smooth phenotypic transitions may play a role in the episodic nature of periodontal disease.

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Significance and Impact of the Study: These in vitro and ex-vivo studies provide a biological rationale for previous clinical studies demonstrating the efficacy of CPC mouthrinses in reducing supragingival plaque and plaque-associated gingivitis.Keywords antimicrobial susceptibility, cetylpyridinium chloride, chlorhexidine, dental plaque, mouthrinse, oral microbiology. AbstractThis study evaluated the antimicrobial activity of two commercially available 0Á05% cetylpyridinium chloride (CPC) mouthrinses with or without alcohol and examined its antimicrobial activity on oral bacterial species including fresh clinical isolates compared to a chlorhexidine mouthrinse and a control fluoride mouthrinse without CPC. Two different approaches were used to evaluate antimicrobial activity. First, the minimum inhibitory concentration (MIC) was determined for each mouthrinse against a panel of 25 micro-organisms including species associated with dental caries, gingivitis and periodontitis. Second, supragingival dental plaque obtained from 15 adults was incubated with the four mouthrinses to evaluate antimicrobial activity on micro-organisms in oral biofilms. Both CPC mouthrinses exhibited lower MIC's, that is, greater antimicrobial activity, against oral Gram-negative bacteria especially periodontal pathogens and species implicated in halitosis such as Aggregatibacter actinomycemcomitans, Campylobacter rectus, Eikenella corrodens, Porphyromonas gingivalis, Prevotella intermedia and Solobacterium moorei than the control mouthrinse. Ex-vivo tests on supragingival plaque micro-organisms demonstrated significantly greater antimicrobial activity by the CPC mouthrinses (>90% killing, P < 0Á001) and the chlorhexidine rinse (>98% killing, P < 0Á05) compared to the control fluoride mouthrinse. Whilst the chlorhexidine mouthrinse was most effective, mouthrinses containing 0Á05% CPC formulated with or without alcohol demonstrated broadspectrum antimicrobial activity against both laboratory strains and supragingival plaque bacteria compared to a control mouthrinse without CPC.

Modern dentistry emphasizes the importance of dental plaque control to improve oral health. The use of oral care formulations with antiplaque biocides plays a crucial role in patient-directed approaches for plaque control. The antiplaque efficacies of these formulations have been extensively studied in many long-term clinical studies designed in accordance with well-accepted guidelines. The results from these studies conclusively demonstrate that long-term use of oral care formulations with well-known antiplaque biocides such as chlorhexidine and triclosan reduce supragingival plaque and gingivitis. This review summarizes microbiological results from clinical studies conducted with oral care formulations containing antiplaque biocides. Results from a number of long-term clinical studies conducted under real-life use conditions indicate no adverse alterations in the bacteria found in dental plaque or emergent microbial resistance. Additionally, microbial sampling of dental plaque subsequent to extended use of antiplaque biocides reveals no increase in resistant microflora. Large numbers of common oral bacteria isolated from patients using chlorhexidine indicate no increase in microbial resistance to chlorhexidine or to commonly used antibiotics. The effects of antiplaque biocides containing oral care formulations on dental plaque that exists naturally as a biofilm are examined. These formulations contain biocide, surfactants, polymers and other components that are effective against the biofilm. In summary, the results of studies on the real-life use of oral care formulations with antiplaque biocides show no emergence of resistant microflora or alterations of the oral microbiota, while such formulations have been found to provide the benefits of reducing plaque and gingivitis.

Actinobacilus actinomycetemcomitans, an oral bacterium implicated in human periodontal disease, was recently demonstrated to invade cultured epithelial cells (D. H. Meyer, P. K. Sreenivasan, and P. M. Fives-Taylor, Infect. Immun. 59:2719-2726. This report characterizes the requirements for invasion of KB cells by A. actinomycetemcomitans. The roles of bacterial and host factors were investigated by using selective agents that influence specific bacterial or host cell functions. Inhibition of bacterial protein synthesis decreased invasion, suggesting the absence of a preformed pool of proteins involved in A. actinomycetemcomitans invasion. Inhibition of bacterial and eukaryotic energy synthesis also decreased invasion, confirming that A. actinomycetemcomitans invasion is an active process. Bacterial adherence to KB cells was indicated by scanning electron microscopy of infected KB cells. Further, the addition ofA. actinomycetemcomitans-specific serum to the bacterial inoculum reduced invasion substantially, suggesting a role for bacterial attachment in invasion. Many of the adherent bacteria invaded the epithelial cells under optimal conditions. Inhibitors of receptor-mediated endocytosis inhibited invasion byA. actinomycetemcomitans. Like that of many facultatively intracellular bacteria, A. actinomycetemcomitans invasion was not affected by eukaryotic endosomal acidification. These are the first published observations describing the requirements for epithelial cell invasion by a periodontopathogen. They demonstrate that A. actinomycetemcomitans utilizes a mechanism similar to those used by many but not all invasive bacteria to gain entry into eukaryotic cells. 1239 Vol. 61, No. 4 on July 16, 2020 by guest http://iai.asm.org/ Downloaded from human periodontal disease.

Twice daily use of a triclosan/copolymer dentifrice may enhance dental implant maintenance by reducing dental plaque and GI.