Clinical Instructions for Using Silver Diamine Fluoride (SDF) in Dental Caries Management.

Manufacturer instructions for 38% silver diamine fluoride (SDF) are limited to current FDA clearance for tooth desensitization. There is a need for instructions to provide best-practice recommendations for off-label use of SDF for caries prevention and arrest. METHODS: The authors considered existing clinical approaches to the use of 38% SDF at pH 10 for the prevention and arrest of active dental caries, in light of the best current evidence. Application of SDF, with or without subsequent direct restoration, is included. The content was reviewed by stakeholders including but not limited to those listed on the consensus statement (Appendix A, below). RESULTS: 38% SDF for the prevention and arrest of active caries lesions, as well as compatibility with common direct restorative materials, such as glass-ionomer cement and resin composite, has a foundation in the scientific literature. A practical decision-flow diagram and accompanying best practices for treatment of caries lesions, based on clinical access and intention to restore, were developed based on available evidence and expert clinical observation when no evidence was available. CONCLUSIONS: Based on the best available evidence, a logical approach can be adopted regarding the practical use of 38% SDF for caries prevention and arrest. PRACTICAL IMPLICATIONS: SDF used as per these instructions for prevention on high-risk tooth surfaces and arrest of active caries lesions has a place in the practitioner's dental caries management armamentarium. When SDF is applied to active lesions, it can be used with or without subsequent restoration, depending on clinical context, expert judgment, and patient input.

Use of unbiased metagenomic and transcriptomic analyses to investigate the association between feline calicivirus and feline chronic gingivostomatitis in domestic cats.

OBJECTIVE: To identify associations between microbes and host genes in cats with feline chronic gingivostomatitis (FCGS), a debilitating inflammatory oral mucosal disease with no known cause, compared with healthy cats and cats with periodontitis (control cats). ANIMALS: 19 control cats and 23 cats with FCGS. PROCEDURES: At least 1 caudal oral mucosal swab specimen was obtained from each cat. Each specimen underwent unbiased metatranscriptomic next-generation RNA sequencing (mNGS). Filtered mNGS reads were aligned to all known genetic sequences from all organisms and to the cat transcriptome. The relative abundances of microbial and host gene read alignments were compared between FCGS-affected cats and control cats and between FCGS-affected cats that did and did not clinically respond to primary treatment. Assembled feline calicivirus (FCV) genomes were compared with reverse transcription PCR (RT-PCR) primers commonly used to identify FCV. RESULTS: The only microbe strongly associated with FCGS was FCV, which was detected in 21 of 23 FCGS-affected cats but no control cats. Problematic base pair mismatches were identified between the assembled FCV genomes and RT-PCR primers. Puma feline foamy virus was detected in 9 of 13 FCGS-affected cats that were refractory to treatment and 5 healthy cats but was not detected in FCGS-affected cats that responded to tooth extractions. The most differentially expressed genes in FCGS-affected cats were those associated with antiviral activity. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that FCGS pathogenesis has a viral component. Many FCV strains may yield false-negative results on RT-PCR-based assays. Coinfection of FCGS-affected cats with FCV and puma feline foamy virus may adversely affect response to treatment.

Managing dental caries against the backdrop of COVID-19: approaches to reduce aerosol generation.

The COVID-19 pandemic resulted in severe limitation and closure of dental practices in many countries. Outside of the acute (peak) phases of the disease, dentistry has begun to be practised again. However, there is emerging evidence that SARS-CoV-2 can be transmitted via airborne routes, carrying implications for dental procedures that produce aerosol. At the time of writing, additional precautions are required when a procedure considered to generate aerosol is undertaken.This paper aims to present evidence-based treatments that remove or reduce the generation of aerosols during the management of carious lesions. It maps aerosol generating procedures (AGPs), where possible, to alternative non-AGPs or low AGPs. This risk reduction approach overcomes the less favourable outcomes associated with temporary solutions or extraction-only approaches. Even if this risk reduction approach for aerosol generation becomes unnecessary in the future, these procedures are not only suitable but desirable for use as part of general dental care post-COVID-19.

Shear Bond Strength of Glass Ionomer Cement to Silver Diamine Fluoride-Treated Artificial Dentinal Caries.

Purpose: The purpose of this study was to measure the shear bond strength (SBS) of glass ionomer cement (GIC) to artificial carious dentin with and without silver diamine fluoride (SDF) treatment. Methods: Permanent molars were sectioned and demineralized to create artificial carious lesions. In five groups, the demineralization of dentin, application of SDF, use of conditioner, and elapsed time between the placement of SDF and restoration were tested for differences in SBS using an UltraTester machine. Statistical analysis was done using the Kruskal-Wallis test and Tukey-Kramer multiple comparison tests. Results: The highest bond strength was found when GIC was placed on conditioned and demineralized dentin treated with SDF one week earlier. Treatment with SDF and use of conditioner did not statistically affect the SBS of GIC to demineralized dentin. Statistically significant increases in bond strength were found when one week elapsed between SDF application and GIC placement. The lowest bond strength was found with immediate GIC application onto SDF-treated demineralized dentin. Conclusions: These in vitro findings suggest that silver diamine fluoride treatment does not significantly affect the bond strength of glass ionomer cement to dentin lesions, and improved retention is obtained by allowing SDF solution to set for one week prior to GIC placement.

Enhanced Tooth Structure Via Silver Microwires Following Treatment with 38 Percent Silver Diamine Fluoride.

Purpose: American Academy of Pediatric Dentistry guidelines recommend treatment of primary teeth with 38 percent silver diamine fluoride (SDF) as a noninvasive option to arrest active dental caries lesions. A significant outcome of SDF treatment are lesions that clinically harden and become more resistant to further decay. Many practicing dentists believe that this increased hardening is due to the reaction of silver and fluoride with carious dentin. The purpose of this study was to focus on the structural and chemical effects of silver diamine fluoride treatment on the native tooth. Methods: In SDF-treated cavitated dentin lesions in teeth subsequently extracted for orthodontic reasons, the authors observed continuous, filamentous silver densities formed in situ from 50 to 2,100 µm in length and 0.25 to 7.0 µm in diameter using high-resolution synchrotron X-ray microcomputer tomography and field emission scanning electron microscopy. These "microwires" fill voids in the lesion caused by disease and permeate through surrounding dentinal tubules. Results: Spectroscopy confirmed that the chemical composition of the observed microwires is predominantly silver. Conclusions: These observations suggest mechanistic explanations for the structural reinforcement of carious dentin in addition to remineralization. It is hypothesized that silver diamine fluoride may achieve its antimicrobial functions by biochemical interactions and through its inherent ability to integrate into the native tooth structure.

Prevention of Dental Caries by Silver Diamine Fluoride.

The use of silver diamine fluoride (SDF) for management of dental caries has gained considerable attention due to recent regulatory clearance in the United States. The primary focus of policies, presentations, and publications has been the arrest of caries lesions (cavities) because of the material's unique ability to non-invasively achieve this elusive and clinically important goal. However, SDF also has proven efficacy in prevention, ie, decreasing the incidence of new caries lesions. Analysis of nine clinical trials in children shows that SDF prevented 61% of new lesions compared to controls. To prevent one new caries lesion, clinicians need to treat four primary teeth (one patient) or 12.1 permanent molars (three patients) with SDF. The preventive effect appears to be immediate and maintains at the same fraction over time. Direct comparisons of SDF applied once per year with alternative treatments show that SDF is more effective than other topical fluorides placed two to four times per year and more cost-effective than dental sealants. Enamel lesions may be even more responsive than cavitated dentin lesions. Annual application of SDF to high-risk surfaces (eg, mesial surfaces of permanent first molars where the distal surface of the second primary molar is carious) in patients with any risk of new caries lesions appears to be the most cost-effective approach available to prevent dental caries. SDF is an underutilized evidence-based preventive agent for dental caries.

Fluorides and Other Preventive Strategies for Tooth Decay.

We focus on scalable public health interventions that prevent and delay the development of caries and enhance resistance to dental caries lesions. These interventions should occur throughout the life cycle, and need to be age appropriate. Mitigating disease transmission and enhancing resistance are achieved through use of various fluorides, sugar substitutes, mechanical barriers such as pit-and-fissure sealants, and antimicrobials. A key aspect is counseling and other behavioral interventions that are designed to promote use of disease transmission-inhibiting and tooth resistance-enhancing agents. Advocacy for public water fluoridation and sugar taxes is an appropriate dental public health activity.

Topical silver diamine fluoride for dental caries arrest in preschool children: A randomized controlled trial and microbiological analysis of caries associated microbes and resistance gene expression.

OBJECTIVES: The Stopping Cavities Trial investigated effectiveness and safety of 38% silver diamine fluoride in arresting caries lesions. MATERIALS AND METHODS: The study was a double-blind randomized placebo-controlled superiority trial with 2 parallel groups. The sites were Oregon preschools. Sixty-six preschool children with ≥1 lesion were enrolled. Silver diamine fluoride (38%) or placebo (blue-tinted water), applied topically to the lesion. The primary endpoint was caries arrest (lesion inactivity, Nyvad criteria) 14-21days post intervention. Dental plaque was collected from all children, and microbial composition was assessed by RNA sequencing from 2 lesions and 1 unaffected surface before treatment and at follow-up for 3 children from each group. RESULTS AND CONCLUSION: Average proportion of arrested caries lesions in the silver diamine fluoride group was higher (0.72; 95% CI; 0.55, 0.84) than in the placebo group (0.05; 95% CI; 0.00, 0.16). Confirmatory analysis using generalized estimating equation log-linear regression, based on the number of arrested lesions and accounting for the number of treated surfaces and length of follow-up, indicates the risk of arrested caries was significantly higher in the treatment group (relative risk, 17.3; 95% CI: 4.3 to 69.4). No harms were observed. RNA sequencing analysis identified no consistent changes in relative abundance of caries-associated microbes, nor emergence of antibiotic or metal resistance gene expression. Topical 38% silver diamine fluoride is effective and safe in arresting cavities in preschool children. CLINICAL SIGNIFICANCE: The treatment is applicable to primary care practice and may reduce the burden of untreated tooth decay in the population.

The Effect of New Oral Care Technologies on the Need for Dentists in 2040.

This article addresses changes in technology of oral self-care or professional care that may increase or decrease the demand for dentists by 2040. The focus is on dental caries, periodontitis, and temporomandibular joint disorders (TMD), as the first two are the main areas of current practice and because TMD is an area for growth. To address this question, the authors examined the scientific literature and government registries to identify recent or soon-to-be-available technologies. They also examined the state of translational efficiency, dissemination, and adoption of advances into dental practice. The pipeline of applicable technology is limited. Nevertheless, between now and 2040, emerging technologies will continue to reduce the need for training more dentists, while no technologies are emerging that will significantly increase the need. Technology in dentistry is adopted slowly as is true in other medical specialties. If a breakthrough product did appear, the results of industry-sponsored trials would be viewed skeptically by the profession, and considerable time would be required to establish the applicability of the findings to the broader population. Greater integration of dentistry into preventive medicine, with dentists offering point-of-service medical testing for systemic disease as suggested by the American Dental Association (ADA), would require a paradigm shift, can occur only over a lengthy period, and is unlikely to impact this assessment. This article was written as part of the project "Advancing Dental Education in the 21st Century."

Plasmid-free CRISPR/Cas9 genome editing in Plasmodium falciparum confirms mutations conferring resistance to the dihydroisoquinolone clinical candidate SJ733.

Genetic manipulation of the deadly malaria parasite Plasmodium falciparum remains challenging, but the rise of CRISPR/Cas9-based genome editing tools is increasing the feasibility of altering this parasite's genome in order to study its biology. Of particular interest is the investigation of drug targets and drug resistance mechanisms, which have major implications for fighting malaria. We present a new method for introducing drug resistance mutations in P. falciparum without the use of plasmids or the need for cloning homologous recombination templates. We demonstrate this method by introducing edits into the sodium efflux channel PfATP4 by transfection of a purified CRISPR/Cas9-guide RNA ribonucleoprotein complex and a 200-nucleotide single-stranded oligodeoxynucleotide (ssODN) repair template. Analysis of whole genome sequencing data with the variant-finding program MinorityReport confirmed that only the intended edits were made, and growth inhibition assays confirmed that these mutations confer resistance to the antimalarial SJ733. The method described here is ideally suited for the introduction of mutations that confer a fitness advantage under selection conditions, and the novel finding that an ssODN can function as a repair template in P. falciparum could greatly simplify future editing attempts regardless of the nuclease used or the delivery method.

MinorityReport, software for generalized analysis of causal genetic variants.

BACKGROUND: The widespread availability of next generation genome sequencing technologies has enabled a wide range of variant detection applications, especially in cancer and inborn genetic disorders. For model systems and microorganisms, the same technology may be used to discover the causative mutations for any phenotype, including those generated in response to chemical perturbation. In the case of pathogenic organisms, these approaches have allowed the determination of drug targets by means of resistance selection followed by genome sequencing. RESULTS: MinorityReport is open source software written in python that facilitates the comparison of any two sets of genome alignments for the purpose of rapidly identifying the spectrum of nonsynonymous changes, insertions or deletions, and copy number variations in a presumed mutant relative to its parent. Specifically, MinorityReport relates mapped sequence reads in SAM format output from any alignment tool for both the mutant and parent genome, relative to a reference genome, and produces the set of variants that distinguishes the mutant from the parent, all presented in an intuitive, straightforward report format. MinorityReport features tunable parameters for evaluating evidence and a scoring system that prioritizes reported variants based on relative proportions of read counts supporting the variant in the mutant versus parent data sets. The utility of MinorityReport is demonstrated using previously published publicly available data sets to find the determinants of resistance for novel anti-malarial drugs. CONCLUSIONS: MinorityReport is readily available (github: JeremyHorst/MinorityReport) to identify the genetic mechanisms of drug resistance in Plasmodium, genotype-phenotype relationships in human diads, or genomic variations between any two related organisms.

UCSF Protocol for Caries Arrest Using Silver Diamine Fluoride: Rationale, Indications and Consent.

The Food and Drug Administration recently cleared silver diamine fluoride for reducing tooth sensitivity. Clinical trials document arrest and prevention of dental caries by silver diamine fluoride. This off-label use is now permissible and appropriate under U.S. law. A CDT code was approved for caries arresting medicaments for 2016 to facilitate documentation and billing. We present a systematic review, clinical indications, clinical protocol and consent procedure to guide application for caries arrest treatment.

Conservation of the glycoprotein B homologs of the Kaposi׳s sarcoma-associated herpesvirus (KSHV/HHV8) and old world primate rhadinoviruses of chimpanzees and macaques.

The envelope-associated glycoprotein B (gB) is highly conserved within the Herpesviridae and plays a critical role in viral entry. We analyzed the evolutionary conservation of sequence and structural motifs within the Kaposi׳s sarcoma-associated herpesvirus (KSHV) gB and homologs of Old World primate rhadinoviruses belonging to the distinct RV1 and RV2 rhadinovirus lineages. In addition to gB homologs of rhadinoviruses infecting the pig-tailed and rhesus macaques, we cloned and sequenced gB homologs of RV1 and RV2 rhadinoviruses infecting chimpanzees. A structural model of the KSHV gB was determined, and functional motifs and sequence variants were mapped to the model structure. Conserved domains and motifs were identified, including an "RGD" motif that plays a critical role in KSHV binding and entry through the cellular integrin αVß3. The RGD motif was only detected in RV1 rhadinoviruses suggesting an important difference in cell tropism between the two rhadinovirus lineages.

Amyloid-like ribbons of amelogenins in enamel mineralization.

Enamel, the outermost layer of teeth, is an acellular mineralized tissue that cannot regenerate; the mature tissue is composed of high aspect ratio apatite nanocrystals organized into rods and inter-rod regions. Amelogenin constitutes 90% of the protein matrix in developing enamel and plays a central role in guiding the hierarchical organization of apatite crystals observed in mature enamel. To date, a convincing link between amelogenin supramolecular structures and mature enamel has yet to be described, in part because the protein matrix is degraded during tissue maturation. Here we show compelling evidence that amelogenin self-assembles into an amyloid-like structure in vitro and in vivo. We show that enamel matrices stain positive for amyloids and we identify a specific region within amelogenin that self-assembles into ß-sheets. We propose that amelogenin nanoribbons template the growth of apatite mineral in human enamel. This is a paradigm shift from the current model of enamel development.

UCSF Protocol for Caries Arrest Using Silver Diamine Fluoride: Rationale, Indications and Consent.

The Food and Drug Administration recently cleared silver diamine fluoride for reducing tooth sensitivity. Clinical trials document arrest and prevention of dental caries by silver diamine fluoride. This off-label use is now permissible and appropriate under U.S. law. A CDT code was approved for caries arresting medicaments for 2016 to facilitate documentation and billing. We present a systematic review, clinical indications, clinical protocol and consent procedure to guide application for caries arrest treatment.

(+)-SJ733, a clinical candidate for malaria that acts through ATP4 to induce rapid host-mediated clearance of Plasmodium.

Drug discovery for malaria has been transformed in the last 5 years by the discovery of many new lead compounds identified by phenotypic screening. The process of developing these compounds as drug leads and studying the cellular responses they induce is revealing new targets that regulate key processes in the Plasmodium parasites that cause malaria. We disclose herein that the clinical candidate (+)-SJ733 acts upon one of these targets, ATP4. ATP4 is thought to be a cation-transporting ATPase responsible for maintaining low intracellular Na(+) levels in the parasite. Treatment of parasitized erythrocytes with (+)-SJ733 in vitro caused a rapid perturbation of Na(+) homeostasis in the parasite. This perturbation was followed by profound physical changes in the infected cells, including increased membrane rigidity and externalization of phosphatidylserine, consistent with eryptosis (erythrocyte suicide) or senescence. These changes are proposed to underpin the rapid (+)-SJ733-induced clearance of parasites seen in vivo. Plasmodium falciparum ATPase 4 (pfatp4) mutations that confer resistance to (+)-SJ733 carry a high fitness cost. The speed with which (+)-SJ733 kills parasites and the high fitness cost associated with resistance-conferring mutations appear to slow and suppress the selection of highly drug-resistant mutants in vivo. Together, our data suggest that inhibitors of PfATP4 have highly attractive features for fast-acting antimalarials to be used in the global eradication campaign.

Pharmacokinetics of fluoride in toddlers after application of 5% sodium fluoride dental varnish.

The prevalence of dental caries (tooth decay) among preschool children is increasing, driven partially by an earlier age of onset of carious lesions. The American Academy of Pediatrics recommends application of 5% sodium fluoride varnish at intervals increasing with caries risk status, as soon as teeth are present. However, the varnishes are marketed for treatment of tooth sensitivity and are regulated as medical devices rather than approved by the US Food and Drug Administration for prevention of dental caries (tooth decay). The objective of this research is to examine the safety of use in toddlers by characterizing the absorption and distribution profile of a currently marketed fluoride varnish. We measured urinary fluoride for 5 hours after application of fluoride varnish to teeth in 6 toddlers aged 12 to 15 months. Baseline levels were measured on a separate day. The urine was extracted from disposable diapers, measured by rapid diffusion, and extrapolated to plasma levels. The mean estimated plasma fluoride concentration was 13 µg/L (SD, 9 µg/L) during the baseline visit and 21 µg/L (SD, 8 µg/L) during the 5 hours after treatment. Mean estimated peak plasma fluoride after treatment was 57 µg/L (SD, 22 µg/L), and 20 µg/kg (SD, 4 µg/L) was retained on average. Retained fluoride was 253 times lower than the acute toxic dose of 5 mg/kg. Mean plasma fluoride after placement of varnish was within an SD of control levels. Occasional application of fluoride varnish following American Academy of Pediatrics guidance is safe for toddlers.

Exome sequencing identifies a recurrent de novo ZSWIM6 mutation associated with acromelic frontonasal dysostosis.

Acromelic frontonasal dysostosis (AFND) is a rare disorder characterized by distinct craniofacial, brain, and limb malformations, including frontonasal dysplasia, interhemispheric lipoma, agenesis of the corpus callosum, tibial hemimelia, preaxial polydactyly of the feet, and intellectual disability. Exome sequencing of one trio and two unrelated probands revealed the same heterozygous variant (c.3487C>T [p. Arg1163Trp]) in a highly conserved protein domain of ZSWIM6; this variant has not been seen in the 1000 Genomes data, dbSNP, or the Exome Sequencing Project. Sanger validation of the three trios confirmed that the variant was de novo and was also present in a fourth isolated proband. In situ hybridization of early zebrafish embryos at 24 hr postfertilization (hpf) demonstrated telencephalic expression of zswim6 and onset of midbrain, hindbrain, and retinal expression at 48 hpf. Immunohistochemistry of later-stage mouse embryos demonstrated tissue-specific expression in the derivatives of all three germ layers. qRT-PCR expression analysis of osteoblast and fibroblast cell lines available from two probands was suggestive of Hedgehog pathway activation, indicating that the ZSWIM6 mutation associated with AFND may lead to the craniofacial, brain and limb malformations through the disruption of Hedgehog signaling.

CANDO and the infinite drug discovery frontier.

The Computational Analysis of Novel Drug Opportunities (CANDO) platform (http://protinfo.org/cando) uses similarity of compound-proteome interaction signatures to infer homology of compound/drug behavior. We constructed interaction signatures for 3733 human ingestible compounds covering 48,278 protein structures mapping to 2030 indications based on basic science methodologies to predict and analyze protein structure, function, and interactions developed by us and others. Our signature comparison and ranking approach yielded benchmarking accuracies of 12-25% for 1439 indications with at least two approved compounds. We prospectively validated 49/82 'high value' predictions from nine studies covering seven indications, with comparable or better activity to existing drugs, which serve as novel repurposed therapeutics. Our approach may be generalized to compounds beyond those approved by the FDA, and can also consider mutations in protein structures to enable personalization. Our platform provides a holistic multiscale modeling framework of complex atomic, molecular, and physiological systems with broader applications in medicine and engineering.