Salivary Cortisol and Anxiety in Canadian Dentists over 1 Year of COVID-19.

The dental profession has endured unprecedented disruption amid COVID-19. Novel stressors have included a high risk of occupational exposure to COVID-19, financial losses, and stricter infection prevention and control requirements. The present study investigated the longitudinal impact of COVID-19 on the stress and anxiety levels of a cohort of Canadian dentists (N = 222) between September 2020 and October 2021. Salivary cortisol was selected as a biomarker of mental stress, and 10 sets of monthly saliva samples (2,131 in total) were self-collected, sent to our laboratory in prepaid courier envelopes, and analyzed by enzyme-linked immunosorbent assay. To assess COVID-19 anxiety, 9 monthly online questionnaires were administered, comprising a general COVID-19 anxiety instrument and 3 items regarding the impact of dentistry-related factors. Bayesian log-normal mixed effect models were fitted to estimate the longitudinal trajectory of salivary cortisol levels and their association with the disease burden of COVID-19 in Canada. After accounting for age, sex, vaccination status, and the diurnal rhythm of cortisol secretion, a modest positive association was found between dentists' salivary cortisol levels and the count of COVID-19 cases in Canada (96% posterior probability). Similarly, the self-reported impact of dentistry-related factors, such as fear of getting COVID-19 from a patient or coworker, was greatest during peaks of COVID-19 waves in Canada; however, general COVID-19 anxiety decreased consistently throughout the study period. Interestingly, at all collection points, the majority of participants were not concerned about personal protective equipment. Overall, participants reported relatively low rates of psychological distress symptoms in relation to COVID-19, a result that should be reassuring for the dental community. Our findings strongly suggest a link between self-reported and biochemical measurements of stress and anxiety in Canadian dentists during the COVID-19 pandemic.

Mechanistic Insights into Bioengineered Antibiofilm Enamel Pellicles.

Dental caries remains the most widespread chronic disease worldwide. Basically, caries originates within biofilms accumulated on dental enamel. Despite the nonrenewable nature of the enamel tissue, targeted preventive strategies are still very limited. We previously introduced customized multifunctional proteinaceous pellicles (coatings) for controlling bacterial attachment and subsequent biofilm succession. Stemmed from our whole proteome/peptidome analysis of the in vivo acquired enamel pellicle, we designed these pellicles using hybrid mixtures of the most abundant and complementary-acting antimicrobial and antifouling proteins/peptides for synergetic suppression of early biofilms. In conjugating these domains synthetically, their bioinhibitory efficacy was remarkably boosted. Herein, we sought to explore the key structure-function relationship of these potent de novo hybridized conjugates in comparison with their individual domains, solely or in physical mixtures. Specifically, we interrelated the following facets: physicochemical and 3-dimensional folding characteristics via molecular dynamics simulations, adopted secondary structure by circular dichroism, immobilization capacity on enamel through high-spatial resolution multiphoton microscopy, and biofilm suppression potency. Our data showed consistent associations among the increased preference for protein folding structures, α-helix content, and enamel-immobilization capacity; all were inversely correlated with the attached bioburden. The expressed phenotypes could be explained by the adopted strongly amphipathic helical conformation upon conjugation, mediated by the highly anionic and acidic N-terminal pentapeptide shared region/motif for enhanced immobilization on enamel. In conclusion, conjugating bioactive proteins/peptides is a novel translational approach to engineer robust antibiofilm pellicles for caries prevention. The adopted α-helical conformation is key to enhance the antibiofilm efficacy and immobilization capacity on enamel that are promoted by certain physicochemical properties of the constituent domains. These data are valuable for bioengineering versatile therapeutics to prevent/arrest dental caries, a condition that otherwise requires invasive treatments with substantial health care expenditures.

Salivary Biomarkers in Denture Stomatitis: A Systematic Review.

OBJECTIVE: Denture stomatitis (DS) is an oral biofilm-associated inflammation of the denture-bearing mucosa. The objective of this review was to identify and evaluate the quality of evidence on the association between the levels of salivary biomarkers and DS among adults with and without palatal DS. MATERIALS AND METHODS: Following the PRISMA guidelines, Medline, PubMed, EMBASE, and the Cochrane Central Register for Controlled Trials were searched for eligible studies from the beginning of the archives until December 2018. Experimental and observational studies with adult participants were included that had a control group or subgroup analysis and provided data on salivary biomarkers and DS. Articles in languages other than English or French were excluded. The level of evidence and grades of recommendation were established with the 2011 scale of the Oxford Centre for Evidence-Based Medicine. Additionally, the assessment of methodological quality was conducted with the STROBE statement (Strengthening the Reporting of Observational Studies in Epidemiology) and graded according to the Olmos scale. RESULTS: From 1,008 citations, 9 studies were included in the systematic review (8 observational, 1 clinical trial). Seven studies suggested a statistically significant difference in the levels of salivary cytokines (IL-6, CCL3, TGF-ß, CXCL8, GM-CSF, and TNF-α) between participants with DS and controls (P < 0.05). In contrast, 2 studies concluded that the difference in the levels of several salivary cytokines (IL2, IL12, IFN-g, IL-4, IL-8, IL-10, IL-17, TNF-α, and ICAM-1) between the groups was not statistically significant. The level of evidence for the majority of studies was 3, while the grade of recommendation for all the studies was B, interpreted as "favorable." In terms of methodological quality, most studies met 50% to 80% of STROBE criteria and were graded B. CONCLUSION: Palatal inflammation in DS is significantly associated with the levels of salivary cytokines. KNOWLEDGE TRANSFER STATEMENT: The results of this study identified altered levels of specific salivary biomarkers associated with denture stomatitis, which may aid in the early diagnosis and treatment of this disease.

Proteomic profile of root canal contents in teeth with post-treatment endodontic disease.

AIM: To characterize the proteome of 20 root canals in teeth with post-treatment endodontic disease using mass spectrometry and to correlate the identified proteins with clinical features. METHODOLOGY: Twenty patients with radiographic evidence of apical periodontitis and need for root canal re-treatment were selected. Samples from the root canal contents were collected and processed using two-dimensional capillary nano-flow liquid chromatography and electrospray ionization tandem mass spectrometry. The acquired spectra were separately searched against specific protein database. The results obtained were analysed using descriptive statistics. Additionally, Pearson's chi-square test or one-sided Fisher's exact test, as appropriate, was chosen to examine the null hypothesis that there is no relationship between each clinical feature and the presence of specific microbial or human proteins. Significance levels were set at 5% (P < 0.05). RESULTS: A total of 1153 human and 720 microbial UniProt accession numbers corresponding to proteins were recovered. The greater prevalence of proteins was related to biological functions, such as cellular and metabolic processes. A considerable number of microbial proteins with clinical relevance functions, such as pathogenesis/virulence, proteolysis, cell adhesion and drug resistance, were detected. Common endodontic pathogens related to post-treatment endodontic disease such as Enterococcus spp., Propionibacterium spp. and Streptococcus spp. were associated with 23, 40 and 94 distinct proteins, respectively. As for human proteins, many factors related to the immune system process were detected. No significant correlations were found between microbial and human proteins and the clinical features investigated (P > 0.05). CONCLUSIONS: A considerable number of microbial and human proteins were identified using proteomic analyses, being mainly related to processes indicating cell viability. No significant correlation was found between proteins and clinical features. These findings suggest a network of important microbial pathogenic functions that may be responsible for the host immune system response.

Postnatal Identification of Zika Virus Peptides from Saliva.

We explored the potential to diagnose Zika virus (ZIKV) infection by analyzing peptides in saliva during a convalescent phase of infection, long after resolution of acute disease. A 25-y-old woman clinically diagnosed with Zika fever in the first trimester was enrolled with her dizygotic twins for a 3-mo postnatal sample of saliva (9-mo after maternal infection). The female baby (A) had microcephaly while the male baby (B) was born healthy. Peptidomic analysis was completed by mass spectrometry (MS/MS), and ZIKV peptides were identified using the National Institutes of Health Zika Virus Resource database, then aligned and mapped to the ZIKV polyprotein to determine proteome coverage and phylogenetic studies. A total of 423 (mother), 607 (baby A), and 183 (baby B) unique ZIKV peptides were identified in saliva by MS/MS, providing a coverage of 67%, 84%, and 45%, respectively, of the entire ZIKV polyprotein (>3,400 amino acids). All peptides were aligned to other flaviviruses that are circulating in Brazil (dengue and yellow fever) to discard false-positive matches. Nine peptides identified were highly conserved to dengue virus. Alignment of a contiguous peptide sequence for mother/babies with the 74 ZIKV sequences suggested that the virus may have entered the oral cavity through the salivary glands, leading to an infection that persists into the postnatal period (vertical transmission). Furthermore, we identified 9 sequence variations that were unique to the baby with microcephaly (not found in the mother or the twin). This sequence information could provide a template for future neuropathogenic studies. A much larger sample size is required to determine whether sequence variation in the envelope protein significantly associates with microcephaly. Finally, from a public health perspective, it will be important to determine whether viral replication is still taking place after birth and whether the virus can be transmitted through salivary contact.

Duplicated or Hybridized Peptide Functional Domains Promote Oral Homeostasis.

Proteins that have existed for millions of years frequently contain repeats of functional domains within their primary structure, thereby improving their functional capacity. In the evolutionary young statherin protein contained within the in vivo-acquired enamel pellicle (AEP), we identified a single functional domain (DR9) located within the protein N-terminal portion that exhibits a higher affinity for hydroxyapatite and more efficient protection against enamel demineralization compared to other native statherin peptides. Thus, we tested the hypothesis that multiplication of functional domains of naturally occurring pellicle peptides amplifies protection against enamel demineralization. In addition, a specific amino acid sequence from histatin 3 (RR-14) was introduced to the hybrid peptides for further testing. Enamel specimens were sectioned to 150-µm thickness and randomly grouped as follows: DR9, DR9-DR9, DR9-RR14, statherin, histatin 1, or distilled water (control). After submersion for 2 h at 37°C, the specimens were placed in 2 mL demineralization solution for 12 d at 37°C. Upon sample removal, the remaining solution was subjected to colorimetric assays to determine the amount of calcium and phosphate released from each specimen. DR9-DR9 amplified protection against enamel demineralization when compared to single DR9 or statherin. Notably, the hybrid peptide DR9-RR14 demonstrated relatively strong protection when the antimicrobial property of these peptides was tested against Candida albicans and Streptococcus mutans. DR9-RR14 was able to maintain 50% of the antifungal activity compared with RR14 for C. albicans and similar values of S. mutans killing activity. This study has pioneered the functional exploration of the natural peptide constituents of the AEP and their evolution-inspired engineered peptides. The knowledge obtained here may provide a basis for the development of stable (proteinase-resistant) synthetic peptides for therapeutic use against dental caries, dental erosion, and/or oral candidiasis.

A Systematic Review Examining the Oral Health Status of Persons with Dementia.

While the oral health of persons with dementia has been shown to be poor, no systematic reviews have been published that examined the topic in depth, including participants with dementia representing the full spectrum of disease severity, and evaluating a broad scope of oral health assessments. The aim of this study was to conduct a current literature review to fill this gap in knowledge. A systematic search of 5 databases (CINAHL, PubMed, EMBASE, Scopus, and ISI Web of Science) was conducted to identify all relevant studies published up to May 2016. There were no exclusions related to study type, severity of dementia, dentate status, or living arrangements. Results were reported descriptively and summarized. Meta-analyses were performed where possible and reported as mean difference (MD) or standardized mean difference (SMD), with a 95% confidence interval (CI). Twenty-eight studies were identified. Assessments were conducted of tooth status, active dental caries, hygiene (plaque/calculus) of natural and artificial teeth, periodontal diseases, denture status (retention, stability, denture-related mucosal lesions), and oral health-related quality of life. Across all evaluations, persons with dementia generally had scores/results suggestive of poor oral health. In meta-analyses, compared with persons without dementia, those with dementia had a significantly fewer number of teeth (MD, -1.52; 95% CI, -0.2.52 to -0.52; P = 0.003; n = 13 studies), more carious teeth (SMD, 0.29; 95% CI, 0.03 to 0.48; P = 0.028; n = 9), significantly worse oral hygiene evaluated using a broad range of assessment tools (SMD, 0.88; 95% CI, 0.57 to 1.19, P < 0.0001; n = 7), and significantly poorer periodontal health (SMD, 0.38; 95% CI, 0.06 to 0.70; P = 0.02; n = 6 studies). The oral health status of persons with mild to severe forms of dementia, who were living in both the community and residential care facilities, was found to be poor across a broad range of dental assessments. Knowledge Transfer Statement: The results of this study define the scope of oral issues and quantify the degree of impairment in individuals with dementia, evaluated using a variety of oral health measures. The results revealed that poor oral health is associated with dementia.

Microbial Diversity in the Early In Vivo-Formed Dental Biofilm.

Although the mature dental biofilm composition is well studied, there is very little information on the earliest phase of in vivo tooth colonization. Progress in dental biofilm collection methodologies and techniques of large-scale microbial identification have made new studies in this field of oral biology feasible. The aim of this study was to characterize the temporal changes and diversity of the cultivable and noncultivable microbes in the early dental biofilm. Samples of early dental biofilm were collected from 11 healthy subjects at 0, 2, 4, and 6 h after removal of plaque and pellicle from tooth surfaces. With the semiquantitative Human Oral Microbiome Identification Microarray (HOMIM) technique, which is based on 16S rRNA sequence hybridizations, plaque samples were analyzed with the currently available 407 HOMIM microbial probes. This led to the identification of at least 92 species, with streptococci being the most abundant bacteria across all time points in all subjects. High-frequency detection was also made with Haemophilus parainfluenzae, Gemella haemolysans, Slackia exigua, and Rothia species. Abundance changes over time were noted for Streptococcus anginosus and Streptococcus intermedius (P = 0.02), Streptococcus mitis bv. 2 (P = 0.0002), Streptococcus oralis (P = 0.0002), Streptococcus cluster I (P = 0.003), G. haemolysans (P = 0.0005), and Stenotrophomonas maltophilia (P = 0.02). Among the currently uncultivable microbiota, eight phylotypes were detected in the early stages of biofilm formation, one belonging to the candidate bacterial division TM7, which has attracted attention due to its potential association with periodontal disease.

Hydroxyapatite Growth Inhibition Effect of Pellicle Statherin Peptides.

In our recent studies, we have shown that in vivo-acquired enamel pellicle is a sophisticated biological structure containing a significant portion of naturally occurring salivary peptides. From a functional aspect, the identification of peptides in the acquired enamel pellicle is of interest because many salivary proteins exhibit functional domains that maintain the activities of the native protein. Among the in vivo-acquired enamel pellicle peptides that have been newly identified, 5 peptides are derived from statherin. Here, we assessed the ability of these statherin pellicle peptides to inhibit hydroxyapatite crystal growth. In addition, atomistic molecular dynamics (MD) simulations were performed to better understand the underlying physical mechanisms of hydroxyapatite growth inhibition. A microplate colorimetric assay was used to quantify hydroxyapatite growth. Statherin protein, 5 statherin-derived peptides, and a peptide lacking phosphate at residues 2 and 3 were analyzed. Statherin peptide phosphorylated on residues 2 and 3 indicated a significant inhibitory effect when compared with the 5 other peptides (P < 0.05). MD simulations showed a strong affinity and fast adsorption to hydroxyapatite for phosphopeptides, whereas unphosphorylated peptides interacted weakly with the hydroxyapatite. Our data suggest that the presence of a covalently linked phosphate group (at residues 2 and 3) in statherin peptides modulates the effect of hydroxyapatite growth inhibition. This study provides a mechanism to account for the composition and function of acquired enamel pellicle statherin peptides that will contribute as a base for the development of biologically stable and functional synthetic peptides for therapeutic use against dental caries and/or periodontal disease.

The impact of stannous, fluoride ions and its combination on enamel pellicle proteome and dental erosion prevention.

OBJECTIVES: To compare the effects of stannous (Sn) and fluoride (F) ions and their combination on acquired enamel pellicle (AEP) protein composition (proteome experiment), and protection against dental erosion (functional experiment). METHODS: In the proteome experiment, bovine enamel specimens were incubated in whole saliva supernatant for 24h for AEP formation. They were randomly assigned to 4 groups (n=10), according to the rinse treatment: Sn (800ppm/6.7mM, SnCl2), F (225ppm/13mM, NaF), Sn and F combination (Sn+F) and deionized water (DIW, negative control). The specimens were immersed 3× in the test rinses for 2min, 2h apart. Pellicles were collected, digested, and analyzed for protein content using liquid chromatography electrospray ionization tandem mass spectrometry. In the functional experiment, bovine enamel specimens (n=10) were similarly treated for pellicle formation. Then, they were subjected to a five-day erosion cycling model, consisting of 5min erosive challenges (15.6 mM citric acid, pH 2.6, 6×/d) and 2min treatment with the rinses containing Sn, F or Sn+F (3×/d). Between the treatments, all specimens were incubated in whole saliva supernatant. Surface loss was determined by profilometry. RESULTS: Our proteome approach on bovine enamel identified 72 proteins that were common to all groups. AEP of enamel treated with Sn+F demonstrated higher abundance for most of the identified proteins than the other groups. The functional experiment showed reduction of enamel surface loss for Sn+F (89%), Sn (67%) and F (42%) compared to DIW (all significantly different, p<0.05). CONCLUSION: This study highlighted that anti-erosion rinses (e.g. Sn+F) can modify quantitatively and qualitatively the AEP formed on bovine enamel. Moreover, our study demonstrated a combinatory effect that amplified the anti-erosive protection on tooth surface.

Nanoscale adhesion forces between enamel pellicle proteins and hydroxyapatite.

The acquired enamel pellicle (AEP) is important for minimizing the abrasion caused by parafunctional conditions as they occur, for instance, during bruxism. It is a remarkable feature of the AEP that a protein/peptide film can provide enough protection in normofunction to prevent teeth from abrasion and wear. Despite its obvious critical role in the protection of tooth surfaces, the essential adhesion features of AEP proteins on the enamel surface are poorly characterized. The objective of this study was to measure the adhesion force between histatin 5, a primary AEP component, and hydroxyapatite (HA) surfaces. Both biotinylated histatin 5 and biotinylated human serum albumin were allowed to adsorb to streptavidin-coated silica microspheres attached to atomic force microscope (AFM) cantilevers. A multimode AFM with a Nanoscope IIIa controller was used to measure the adhesion force between protein-functionalized silica microspheres attached to cantilever tips and the HA surface. The imaging was performed in tapping mode with a Si3N4 AFM cantilever, while the adhesion forces were measured in AFM contact mode. A collection of force-distance curves (~3,000/replicate) was obtained to generate histograms from which the adhesion forces between histatin 5 or albumin and the HA surface were measured. We found that histatin 5 exhibited stronger adhesion forces (90% >1.830 nN) to the HA surface than did albumin (90% > 0.282 nN). This study presents an objective approach to adhesion force measurements between histatin 5 and HA, and provides the experimental basis for measuring the same parameters for other AEP constituents. Such knowledge will help in the design of synthetic proteins and peptides with preventive and therapeutic benefits for tooth enamel.

Effect of dialyzed saliva on human enamel demineralization.

BACKGROUND: Saliva is supersaturated with respect to calcium and phosphate ions. Salivary ions may well play a role in the subsequent adsorption of proteins and consequently in the formation of the acquired enamel pellicle. Among several biological functions, the enamel pellicle forms a selectively permeable barrier that regulates demineralization processes. AIM: The aim of this study was to evaluate the importance of salivary proteins when adsorbed on enamel surface and the resultant protective effect against demineralization without the presence of salivary ions. METHODS: Enamel surfaces were coated with whole saliva, parotid saliva, dialyzed whole saliva or dialyzed parotid saliva (molecular weight cutoff 1 kDa). Adsorption was allowed to proceed for a period of 2 h. Enamel specimens were then washed with deionized water and immersed into a demineralization solution of pH 4.5 for 12 days. This solution was used to measure the amount of calcium and phosphate released from enamel specimens after the demineralization period. RESULTS: All coated specimen groups showed a significantly higher protection than those not coated with any type of saliva. In addition, undialyzed saliva (whole saliva and parotid saliva) was more effective in protecting the enamel against demineralization than dialyzed saliva. CONCLUSION: The present investigation indicates that the ionic composition of saliva can amplify the demineralization protection effect by reducing acid-induced enamel demineralization. Moreover, a protective effect of salivary proteins without presence of ions was demonstrated in this study.

The effect of histatin 5, adsorbed on PMMA and hydroxyapatite, on Candida albicans colonization.

The limited number of treatments for oral candidiasis resulted in the emergence of azole-resistant Candida albicans strains, thus enforcing the need for novel antifungal treatments. Although histatin 5 (H5) demonstrates antifungal activity, its inhibitory effect when adhered to hydroxyapatite and Polymetylmethacrylate (PMMA) surfaces, resembling conditions of the in vivo pellicle, remains unexplored. The objective of this in vitro study was to determine whether surface-adhered H5 inhibits the colonization of C. albicans on hydroxyapatite and/or PMMA. The C. albicans assay involved developing a mono-protein pellicle (either H5 or albumin) on hydroxyapatite and PMMA discs, introducing C. albicans and counting the number of adhered cells, throughout time, using scanning electron microscopy. A negative binomial statistical model and the Tukey-Kramer test were used for statistical analysis, with p < 0.01 indicating significance. H5-coated PMMA had significantly reduced number of cells compared to albumin-coated PMMA at 30, 90 and 1440 min (p < 0.0001), with the number of cells decreasing significantly in 90 and 1440 min (p < 0.0001). Similarly, H5-coated hydroxyapatite had significantly fewer cells compared to the albumin-coated surface at 90 and 1440 min (p < 0.0001), with the number of cells decreasing significantly at 30, 90 and 1440 min (p < 0.0001). In conclusion, C. albicans colonization was most inhibited by PMMA and hydroxyapatite-adhered H5 after 1440 min, illustrating the time-dependent effect of H5. In addition, yeast cells colonized albumin-coated PMMA, while dense hyphal networks formed on albumin-coated hydroxyapatite, suggesting that C. albicans morphology is influenced by the surface available for albumin adhesion.

New insights into the composition and functions of the acquired enamel pellicle.

The acquired enamel pellicle (AEP) is a thin acellular film that forms on tooth surfaces upon exposure to the oral environment. It consists predominantly of salivary proteins, but also includes non-salivary-derived proteins, carbohydrates, and lipids. Since it is the interface between teeth and the oral environment, the AEP plays a key role in the maintenance of oral health by regulating processes including lubrication, demineralization, and remineralization and shaping the composition of early microbial flora adhering to tooth surfaces. Knowledge of the 3D structure of the AEP and how that correlates with its protective functions may provide insight into several oral pathological states, including caries, erosion, and periodontal disease. This review intends to update readers about the latest discoveries related to the formation, ultrastructure, composition, and functions of the AEP, as well as the future of pellicle research, with particular emphasis on the emerging role of proteomic and microscopy techniques in oral diagnosis and therapeutics.

Whole-saliva proteolysis and its impact on salivary diagnostics.

There is growing interest in the use of human whole saliva for diagnostics and disease monitoring as an alternative to blood samples. In contrast to blood, whole saliva is a non-sterile body fluid. Proper hand-ling and storage are required to preserve the integrity of potential biomarkers. We investigated salivary autoproteolytic degradation using a variety of approaches. We determined inhibition of protease activities by monitoring the endogenous proteome. In addition, the stability of highly protease-susceptible proteins-histatin 5, statherin, and PRP1-was assessed. Experimental variables included (a) protease inhibitors, (b) salivary pH, (c) incubation temperatures, and (d) sample heating. A cocktail containing AEBSF, aprotinin, pancreatic trypsin inhibitor, leupeptin, antipain, and EDTA could not prevent histatin 5, statherin, or PRP1 degradation in whole saliva. Among the other treatments evaluated, short-term storage of freshly collected samples on ice was effective without interfering with the chemistry of the proteome. In conclusion, whole saliva contains a unique mixture of enzymes as evidenced from their resilience to protease inhibition. Analytical evidence on protein stability is needed to ensure the validity of salivary biomarker study outcomes. Analysis of the data presented will provide help and guidance for the use of saliva samples for diagnostic purposes.

Clinical performance of two fluorescence-based methods in detecting occlusal caries lesions in primary teeth.

This in vivo study aimed to evaluate the performance of 2 fluorescence-based methods in detecting occlusal caries lesions in primary teeth, compared with the performance of visual inspection and radiographic methods, and to propose a mathematic correction of the diagnostic parameters due to the imperfect reference standard method used in the study. Two examiners assessed the occlusal surfaces of 407 primary teeth (62 children) using visual inspection (ICDAS), radiographic, DIAGNOdent pen (pen type laser fluorescence; LFpen), and fluorescence camera (FC) methods. At the noncavitated threshold (NC) the reference standard method was the results of ICDAS, and at the dentine caries threshold (D3) teeth diagnosed with dentine caries by ICDAS or radiographic methods were subjected to operative treatment to confirm the presence of lesion. Reproducibility, sensitivity, specificity, accuracy, and the area under the ROC curve were calculated for the methods at both thresholds. At the NC threshold, LFpen had a slightly better performance compared to the FC and radiographic methods. However, at the D3 threshold, both fluorescence-based methods performed similarly. Visual inspection and radiographic methods presented higher specificities but lower sensitivities than fluorescence methods. After corrections, there was a significant decrease in some parameters. In conclusion, both fluorescence-based methods presented similar performance in detecting occlusal dentine caries lesions in primary teeth, but they usually gave more false-positive results than did the visual and radiographic methods. The correction proposed shows that the performance of the methods can be overestimated, and the correction should be validated and considered in further studies that use an imprecise reference standard method.

Histatin 1 resists proteolytic degradation when adsorbed to hydroxyapatite.

Histatins are salivary proteins that exhibit a high affinity for hydroxyapatite and contribute to the acquired enamel pellicle. Previous studies have observed that, despite the high proteolytic activity in saliva, significant numbers of histatin molecules in acquired enamel pellicle are intact. Our working hypothesis was that histatins are less susceptible to proteinases present in saliva when adsorbed on the hydroxyapatite. To test this premise, we incubated histatin 1 with hydroxyapatite and human whole saliva. Proteolytic products of this incubation were then characterized by PAGE, HPLC, and mass spectrometry. This study shows for the first time that binding to hydroxyapatite confers intact histatin 1 with resistance to proteolytic degradation.

Evidence of intact histatins in the in vivo acquired enamel pellicle.

Understanding the composition and function of the acquired enamel pellicle (AEP) has been a major goal in oral biology. The aim of this study was to test the hypothesis that intact histatins are part of the in vivo AEP and that histatins after adsorption to HA have effects on in vitro enamel demineralization. This is the first study demonstrating the presence of intact histatins in vivo in the AEP. The in vitro experiments show that all naturally occurring histatins in the AEP have the potential to provide some level of protection against acid injury.

Multiple components contribute to ability of saliva to inhibit influenza viruses.

INTRODUCTION: Saliva is a potentially important barrier against respiratory viral infection but its mechanism of action is not well studied. METHODS: We tested the antiviral activities of whole saliva, specific salivary gland secretions, and purified salivary proteins against strains of influenza A virus (IAV) in vitro. RESULTS: Whole saliva or parotid or submandibular/sublingual secretions from healthy donors inhibited IAV based on hemagglutination inhibition and neutralization assays. This differs from human immunodeficiency virus (HIV), for which only submandibular/sublingual secretions are reported to be inhibitory. Among purified salivary proteins, MUC5B, scavenger receptor cysteine-rich glycoprotein 340 (salivary gp-340), histatins, and human neutrophil defensins (HNPs) inhibited IAV at the concentrations present in whole saliva. In contrast, some abundant salivary proteins (acidic proline-rich proteins and amylase) had no activity, nor did several other less abundant salivary proteins with known activity against HIV (e.g. thrombospondin or serum leukocyte protease inhibitor). Whole saliva and MUC5B did not inhibit neuraminidase activity of IAV and viral neutralizing and aggregating activity of MUC5B was potentiated by the neuraminidase inhibitor oseltamivir. Hence, MUC5B inhibits IAV by presenting a sialic acid ligand for the viral hemagglutinin. The mechanism of action of histatins requires further study. CONCLUSIONS: These findings indicate that saliva represents an important initial barrier to IAV infection and underline the complexity of host defense activity of oral secretions. Of interest, antiviral activity of saliva against IAV and HIV differs in terms of specific glandular secretions and proteins that are inhibitory.

Proteome of human minor salivary gland secretion.

Recent research efforts in oral biology have resulted in elucidation of the proteomes of major human salivary secretions and whole saliva. One might hypothesize that the proteome of minor gland secretions may show significantly different characteristics when compared with the proteomes of parotid or submandibular/sublingual secretions. To test this hypothesis, we conducted the first exploration into the proteome of minor salivary gland secretion. Minor gland secretion was obtained from healthy volunteers, and its components were subjected to liquid-chromatography-electrospray-ionization-tandem-mass-spectrometry. This led to the identification of 56 proteins, 12 of which had never been identified in any salivary secretion. The unique characteristics of the minor salivary gland secretion proteome are related to the types as well as the numbers of components present. The differences between salivary proteomes may be important with respect to specific oral functions.