Interspecies interactions within oral microbial communities.

While reductionism has greatly advanced microbiology in the past 400 years, assembly of smaller pieces just could not explain the whole! Modern microbiologists are learning "system thinking" and "holism." Such an approach is changing our understanding of microbial physiology and our ability to diagnose/treat microbial infections. This review uses oral microbial communities as a focal point to describe this new trend. With the common name "dental plaque," oral microbial communities are some of the most complex microbial floras in the human body, consisting of more than 700 different bacterial species. For a very long time, oral microbiologists endeavored to use reductionism to identify the key genes or key pathogens responsible for oral microbial pathogenesis. The limitations of reductionism forced scientists to begin adopting new strategies using emerging concepts such as interspecies interaction, microbial community, biofilms, polymicrobial disease, etc. These new research directions indicate that the whole is much more than the simple sum of its parts, since the interactions between different parts resulted in many new physiological functions which cannot be observed with individual components. This review describes some of these interesting interspecies-interaction scenarios.

Systematic approach to optimizing specifically targeted antimicrobial peptides against Streptococcus mutans.

Previously we reported a novel strategy of "targeted killing" through the design of narrow-spectrum molecules known as specifically targeted antimicrobial peptides (STAMPs) (R. Eckert et al., Antimicrob. Agents Chemother. 50:3651-3657, 2006; R. Eckert et al., Antimicrob. Agents Chemother. 50:1480-1488, 2006). Construction of these molecules requires the identification and the subsequent utilization of two conjoined yet functionally independent peptide components: the targeting and killing regions. In this study, we sought to design and synthesize a large number of STAMPs targeting Streptococcus mutans, the primary etiologic agent of human dental caries, in order to identify candidate peptides with increased killing speed and selectivity compared with their unmodified precursor antimicrobial peptides (AMPs). We hypothesized that a combinatorial approach, utilizing a set number of AMP, targeting, and linker regions, would be an effective method for the identification of STAMPs with the desired level of activity. STAMPs composed of the Sm6 S. mutans binding peptide and the PL-135 AMP displayed selectivity at MICs after incubation for 18 to 24 h. A STAMP where PL-135 was replaced by the B-33 killing domain exhibited both selectivity and rapid killing within 1 min of exposure and displayed activity against multispecies biofilms grown in the presence of saliva. These results suggest that potent and selective STAMP molecules can be designed and improved via a tunable "building-block" approach.

Dentistry and dental education in the context of the evolving health care system.

This article is intended to stimulate dialogue within the intertwined dental practice and dental education communities about our evolving health care system and dentistry's role within this system as it reconfigures in response to a complex interplay of influences. The changing dental disease burden in the United States is analyzed with consideration of how evolution in disease prevalence influences societal need for dental services and the resulting potential impact on the types of services provided and the education of future dental practitioners. The article concludes with discussion of a potential future scenario for practice and education in which one or both of the two health abnormalities (dental caries and periodontal diseases) most closely associated with dentistry as an area of medical specialization go away as a consequence of transformational technologies.

Dental insurance: will it help or hinder adoption of caries management practices?

Whether public or private dental insurance will provide benefits for caries management practices is a business decision. The foundation for this decision is multifactorial and continually changing as the values of the purchasers and health care consumers evolve. Understanding the dynamics involved in allocating finite health care resources will help those who advocate for caries management inform decision makers about the potential benefits of these strategies.

In situ and non-invasive detection of specific bacterial species in oral biofilms using fluorescently labeled monoclonal antibodies.

Noninvasive in situ detection of suspected cariogenic bacterial species within dental biofilms could facilitate monitoring of the dynamic change of oral microbial flora and assist in the assessment of the treatment efficacy of therapeutic agents. In this study, we explore the possibility to use three well-characterized monoclonal antibodies (MAbs) against Streptococcus mutans, Actinomyces naeslundii, and Lactobacillus casei to identify these three important members of the oral microbial community in the complex environment of oral biofilms. These MAbs, which were conjugated to different fluorescent labels and visualized with confocal laser scanning microscopy (CLSM), proved to be an useful tool to identify the three species of interest (S. mutans, A. naeslundii, and L. casei) under various experimental conditions including in vitro and in vivo derived oral biofilms. Manifold addition of the MAbs on consecutive days did not alter the biofilm structure thus allowing monitoring of the same biofilm over extended time periods. Using this MAb-based method the effect of sucrose challenge on the biofilm composition and the distribution of S. mutans, A. naeslundii, and L. casei were examined. S. mutans was found to be the predominant species under the various biofilm conditions tested. These studies indicate that MAbs based bacterial detection with CLSM is a versatile tool which permits new insights into the ecology of oral biofilm development.

Reimbursement for dental implants: dispelling some popular myths.

BACKGROUND AND OVERVIEW: Dental implants are being placed with increasing regularity. Many dentists do not take advantage of insurance coverages for this treatment because they are under the impression that no aspect of implant therapy is covered. This article discusses a number of insurance benefits that may be available to dental patients but not readily apparent to treating dentists and their staff members. CONCLUSIONS AND PRACTICE IMPLICATIONS: Some dentists and patients may assume incorrectly that a dental insurance plan does not reimburse for any implant therapy when, in fact, there may be some benefit available for at least a portion of the treatment. In addition, some dentists and patients may not consider implant therapy even when it is the preferred treatment option because of the assumed lack of reimbursement. Knowing that some reimbursement is available may make the difference in the patient's accepting the best treatment for his or her condition. Furthermore, since an increasing number of patients now make some contribution toward their dental insurance plan premiums, they deserve to know their options and to receive appropriate benefits.

Future trends in dental benefits.

Dentistry and dental payment systems as we know them today will continue to evolve. Dentistry as practiced today and the prepayment systems of dentistry are substantially different than they were fifty years ago when dental insurance as we know it was first developed. Dentistry has always changed with the development of our science and the expression of dentistry's diseases in the populations we serve. The changes that are likely to occur in the future will be focused on improving health outcomes across risk-analyzed populations with the goals of providing optimal health outcomes at reasonable costs. Dentists will increasingly become engaged in the whole health of their patients. Where sufficient correlations can be leveraged between dentistry and overall health, medical plans will play an increasing role in dentistry's future for two reasons. Given favorably altered therapeutic outcomes for medical systems that preserve scarce resources, it will be an economic imperative to engage the dental system. It will also be the right thing to do from a total health perspective. In the final analysis, this elevates the role of dentistry and empowers the dentist to participate in the total health of their patients.

Patterns of oral care in dental school and general dental practice.

This study compared patterns of oral care provided by predoctoral dental students for patients seeking treatment at the University of Washington (UW) with patterns reported for general dental offices by the Washington Dental Service (WDS). Dental care included about 5 million services provided to 880,317 patients by 2,803 WDS general dentists and about 45,600 dental services provided to 9,488 patients by 155 UW dental students during 1999. There was high fidelity between databases and randomly surveyed patient records for treatment provided in both UW (95 percent) and WDS (97 percent) populations. While patient age patterns were generally similar, UW students completed more procedures for young children and for adults older than seventy-four years but completed fewer procedures for age groups of from thirteen to eighteen and from forty-five to fifty-four than general dental offices. The relative mix of all services completed by UW and WDS providers was similar (ANOVA, P=0.82). Within categories of service, the percentage of total services completed by students compared to those submitted by community dentists to WDS was about the same for examinations, radiographs, fluoride and sealants, amalgams, composites, single crowns, and endodontics. The percentage of total procedures completed showed a greater emphasis by UW students on inlays/onlays, dentures, extractions, and periodontal maintenance, and lesser experience with implants, orthodontics, sedation, and emergency procedures than general dental offices. We conclude that the relative distribution of clinical services provided by UW dental students is comparable to those procedures reported to WDS by dental offices in the adjacent community.

A review of the efficacy of chlorhexidine on dental caries and the caries infection.

Chlorhexidine has been used for the past 35 years in the treatment of the two primary diseases of dentistry with varying degrees of success. The purpose of this paper is to review the literature for both the caries reduction potential and the microbiologic reduction of the pathogens associated with dental caries. The literature remains mixed on the success of chlorhexidine for the reduction of dental caries. Its performance as an antimicrobial against Streptococcus mutans is more consistent and favorable.

Rapid probing of biological surfaces with a sparse-matrix peptide library.

Finding unique peptides to target specific biological surfaces is crucial to basic research and technology development, though methods based on biological arrays or large libraries limit the speed and ease with which these necessary compounds can be found. We reasoned that because biological surfaces, such as cell surfaces, mineralized tissues, and various extracellular matrices have unique molecular compositions, they present unique physicochemical signatures to the surrounding medium which could be probed by peptides with appropriately corresponding physicochemical properties. To test this hypothesis, a naïve pilot library of 36 peptides, varying in their hydrophobicity and charge, was arranged in a two-dimensional matrix and screened against various biological surfaces. While the number of peptides in the matrix library was very small, we obtained "hits" against all biological surfaces probed. Sequence refinement of the "hits" led to peptides with markedly higher specificity and binding activity against screened biological surfaces. Genetic studies revealed that peptide binding to bacteria was mediated, at least in some cases, by specific cell-surface molecules, while examination of human tooth sections showed that this method can be used to derive peptides with highly specific binding to human tissue.

Development and evaluation of a safe and effective sugar-free herbal lollipop that kills cavity-causing bacteria.

Dental caries (tooth decay) is caused by a specific group of cariogenic bacteria, like Streptococcus mutans, which convert dietary sugars into acids that dissolve the mineral in tooth structure. Killing cariogenic bacteria is an effective way to control or prevent tooth decay. In a previous study, we discovered a novel compound (Glycyrrhizol A), from the extraction of licorice roots, with strong antimicrobial activity against cariogenic bacteria. In the current study, we developed a method to produce these specific herbal extracts in large quantities, and then used these extracts to develop a sugar-free lollipop that effectively kills cariogenic bacteria like Streptococcus mutans. Further studies showed that these sugar-free lollipops are safe and their antimicrobial activity is stable. Two pilot human studies indicate that a brief application of these lollipops (twice a day for ten days) led to a marked reduction of cariogenic bacteria in oral cavity among most human subjects tested. This herbal lollipop could be a novel tool to promote oral health through functional foods.

Novel synthetic antimicrobial peptides against Streptococcus mutans.

Streptococcus mutans, a common oral pathogen and the causative agent of dental caries, has persisted and even thrived on the tooth surface despite constant removal and eradication efforts. In this study, we generated a number of synthetic antimicrobial peptides against this bacterium via construction and screening of several structurally diverse peptide libraries where the hydrophobicity and charge within each library was varied incrementally in order to generate a collection of peptides with different biochemical characteristics. From these libraries, we identified multiple peptides with robust killing activity against S. mutans. To further improve their effectiveness, the most bactericidal peptides from each library were synthesized together as one molecule, in various combinations, with and without a flexible peptide linker between each antimicrobial region. Many of these "fusion" peptides had enhanced killing activities in comparison with those of the original nonconjoined molecules. The results presented here illustrate that small libraries of biochemically constrained peptides can be used to generate antimicrobial peptides against S. mutans, several of which may be likely candidates for the development of anticaries agents.

Comparative analysis of a monoclonal antibody-based Streptococcus mutans detection method with selective culture assays using polymerase chain reaction as a gold standard.

The aim of this study was to compare a recently developed monoclonal antibody (MAb)-based salivary Streptococcus mutans detection method with various selective media using polymerase chain reaction (PCR) as the gold standard. Salivary S. mutans cells were enumerated with a MAb-based method, along with three commonly used selective media, mitis-salivarius-bacitracin agar (MSB), trypticase yeast-extract cystine sucrose bacitracin agar (TYCSB), and glucose-sucrose-potassium tellurite-bacitracin (GSTB) agar. Statistical analysis showed no significant correlations between each method. With PCR as the standard, a MAb-based detection method was found to provide the highest sensitivity (91%) and specificity (96%) among these four methods. This study demonstrates that the MAb-based detection method may provide more accurate enumeration of salivary S. mutans than selective media.

Targeted killing of Streptococcus mutans by a pheromone-guided "smart" antimicrobial peptide.

Within the repertoire of antibiotics available to a prescribing clinician, the majority affect a broad range of microorganisms, including the normal flora. The ecological disruption resulting from antibiotic treatment frequently results in secondary infections or other negative clinical consequences. To address this problem, our laboratory has recently developed a new class of pathogen-selective molecules, called specifically (or selectively) targeted antimicrobial peptides (STAMPs), based on the fusion of a species-specific targeting peptide domain with a wide-spectrum antimicrobial peptide domain. In the current study, we focused on achieving targeted killing of Streptococcus mutans, a cavity-causing bacterium that resides in a multispecies microbial community (dental plaque). In particular, we explored the possibility of utilizing a pheromone produced by S. mutans, namely, the competence stimulating peptide (CSP), as a STAMP targeting domain to mediate S. mutans-specific delivery of an antimicrobial peptide domain. We discovered that STAMPs constructed with peptides derived from CSP were potent against S. mutans grown in liquid or biofilm states but did not affect other oral streptococci tested. Further studies showed that an 8-amino-acid region within the CSP sequence is sufficient for targeted delivery of the antimicrobial peptide domain to S. mutans. The STAMPs presented here are capable of eliminating S. mutans from multispecies biofilms without affecting closely related noncariogenic oral streptococci, indicating the potential of these molecules to be developed into "probiotic" antibiotics which could selectively eliminate pathogens while preserving the protective benefits of a healthy normal flora.

Mutation of luxS affects biofilm formation in Streptococcus mutans.

Quorum sensing is a bacterial mechanism for regulating gene expression in response to changes in population density. Many bacteria are capable of acyl-homoserine lactone-based or peptide-based intraspecies quorum sensing and luxS-dependent interspecies quorum sensing. While there is good evidence about the involvement of intraspecies quorum sensing in bacterial biofilm, little is known about the role of luxS in biofilm formation. In this study, we report for the first time that luxS-dependent quorum sensing is involved in biofilm formation of Streptococcus mutans. S. mutans is a major cariogenic bacterium in the multispecies bacterial biofilm commonly known as dental plaque. An ortholog of luxS for S. mutans was identified using the data available in the S. mutans genome project (http://www.genome.ou.edu/smutans.html). Using an assay developed for the detection of the LuxS-associated quorum sensing signal autoinducer 2 (AI-2), it was demonstrated that this ortholog was able to complement the luxS negative phenotype of Escherichia coli DH5alpha. It was also shown that AI-2 is indeed produced by S. mutans. AI-2 production is maximal during mid- to late-log growth in batch culture. Mutant strains devoid of the luxS gene were constructed and found to be defective in producing the AI-2 signal. There are also marked phenotypic differences between the wild type and the luxS mutants. Microscopic analysis of in vitro-grown biofilm structure revealed that the luxS mutant biofilms adopted a much more granular appearance, rather than the relatively smooth, confluent layer normally seen in the wild type. These results suggest that LuxS-dependent signal may play an important role in biofilm formation of S. mutans.

Analyses of Streptococcus mutans in saliva with species-specific monoclonal antibodies.

Three species-specific monoclonal antibodies (MAbs) against Streptococcus mutans were used to detect and quantify S. mutans levels in saliva. This study shows that MAb-based salivary S. mutans tests exhibit significantly higher specificity and sensitivity than the commonly used selective culture method. Examination of nearly 2,000 human saliva samples shows that S. mutans counts in human saliva vary from less than 10,000 to a high 36 million cells/mL. Over 15% of the saliva samples examined have salivary S. mutans counts over 500,000 cells/mL. When saliva samples were collected at different time points during a day, the number of salivary S. mutans in the same human subject varied, especially before and after sugar uptake. Additionally, data obtained from stimulated versus unstimulated saliva in the same human subjects differed greatly and appear to be completely uncorrelated. This study provides useful information and tools for analyzing the role of S. mutans in human dental caries.