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Molecular Diagnosis and New Therapeutic Approach of Oral Diseases

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A special issue of Journal of Personalized Medicine (ISSN 2075-4426).

Deadline for manuscript submissions: closed (20 September 2021) | Viewed by 40987

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Special Issue Editors

Prof. Dr. Gaetano Isola

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Guest Editor

Department of General Surgery and Surgical-Medical Specialties, University of Catania, Catania, Italy
Interests: periodontitis; oral surgery; oral pathology; oral health-systemic health; bone biology
Special Issues, Collections and Topics in MDPI journals

Dr. Romeo Patini

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Guest Editor

Department of Head, Neck and Sense Organs, Catholic University of the Sacred Heart, 00168 Rome, Italy
Interests: systematic review; methodology; oral microbiome; dental diseases; regenerative medicine
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

In recent years, medical sciences have undergone exponential growth in terms of cutting-edge diagnostic tools and techniques and experimental therapeutic strategies. The most crucial and innovative factor in this panorama is the constant effort of the researchers in adapting diagnostic appropriateness and therapeutic expertise to single-patient needs in a completely personalized way. This is currently known as personalized medicine. In this field, which is still in a whirlwind of evolution, much remains to be investigated. Oral health is now recognized as an important sentinel element for the preservation and maintenance of general health, and the study of genomics, proteomics, and metabolomics (among the various involved sciences) seems to be one of the most important factors in gaining new knowledge in the field of personalized dental medicine. All the abovementioned elements, along with the effects of the microbiome and the importance of its balance, for the development of preventive and therapeutic strategies to face oral diseases, constitute the essential framework of this Special Issue.

The aim of this Special Issue is to provide insight into recent advances in the field of personalized dental medicine. Potential topics in this Special Issue include but are not limited to in-vitro and clinical research on genetics, oral health, oral pathology, oral surgery, and oral microbiology.

Studies incorporating a new approach or providing novel information are high-priority. Systematic and narrative reviews in the field of dentistry focusing mainly on infectious disease are also welcome. It is our pleasure to invite you to contribute to this Special Issue.

Prof. Dr. Gaetano Isola
Prof. Dr. Romeo Patini
Guest Editors

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Journal of Personalized Medicine is an international peer-reviewed open access monthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2600 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

periodontitis
caries
oral health
microbiota
oral microbiology
oral bacteria
oral diseases
periodontal diseases
peri-implant diseases
dental infections
oral surgery
oral pathology
dental biomaterials
in-vitro research
clinical research

Published Papers (11 papers)

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Editorial

Jump to: Research

3 pages, 179 KiB

Open AccessEditorial

Salivary Tests: A New Personalized Approach for the Early Diagnosis of Oral and Periodontal Diseases

by Gaetano Isola

J. Pers. Med. 2022, 12(10), 1636; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm12101636 - 02 Oct 2022

Cited by 2 | Viewed by 1486

Abstract

Periodontitis is a chronic bacterial infection of the periodontium that results in the destruction of bone and connective tissue [...] Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

4 pages, 198 KiB

Open AccessEditorial

Personalized Dentistry: Approaching a New Way for Diagnosis and Treatment of Oral Diseases

by Romeo Patini

J. Pers. Med. 2020, 10(2), 35; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm10020035 - 01 May 2020

Cited by 4 | Viewed by 4235

Abstract

For years, it has been thought that the field of dentistry was referring exclusively to some diseases that strictly affect the oral cavity. Dental caries, periodontal disease, and pathologies associated with their worsening were considered almost the only interest in scientific research in dentistry. Recent studies have begun to shed light on the effect of the oral microbiota on general health and on the crucial role of dentistry in its maintenance. In this way, we came to understand that the bacterial populations that make up the oral microbiota can vary profoundly between individuals and that contribute in a fundamental way to outlining the so-called “oral signature”. This characteristic is called into question to evaluate the susceptibility, or lack thereof, of the subject to the contraction of a wide range of pathologies, apparently not connected with oral health. From this evidence, it will also be possible to study therapeutic approaches aimed at the eradication of species considered at risk or colonization with species considered protective; thus, giving life to so-called “personalized dentistry”. Therefore, this Special Issue is aimed at spreading the scientific knowledge over the current limits in terms of new molecular and culturomic approaches towards the diagnosis of oral microbiota and the treatment techniques of eventually associated systemic diseases. In vivo studies and systematic literature reviews with quantitative analysis of results, when possible, will be given a high priority. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

Research

Jump to: Editorial

10 pages, 783 KiB

Open AccessArticle

Real-Time Feedback of the Applied Light-Curing Technique and Its Impact on Degree of Conversion of Composite Restorations—A Study with Undergraduate Dental Students

by Phoebe Burrer, Matej Par, Thomas Attin and Tobias T. Tauböck

J. Pers. Med. 2021, 11(10), 1012; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm11101012 - 09 Oct 2021

Cited by 1 | Viewed by 1136

Abstract

The objective of this study was to investigate the effect of individual instructions and training of dental students on the amount of applied light irradiance before and after training using a patient simulator with integrated visual feedback. Furthermore, the effect on the degree of conversion of composite restorations placed by the dental students was assessed. Forty-two dental students, split into two groups, light-cured a simulated restoration in tooth 27 of a dental patient simulator for 20 s. The irradiance (mW/cm²) received by the detector was measured in real-time before and after individual instructions and training, and the energy delivered (J/cm²) was calculated for each student. The degree of conversion at the bottom of incrementally placed composite restorations prior to individual instructions (group 1) and after individual instructions (group 2) was assessed using Fourier-transform infrared (FTIR) spectroscopy. The irradiance and degree of conversion measurements were re-assessed after all students received individual instructions. Data were analyzed using Wilcoxon signed-rank test and Mann-Whitney U-test at an overall level of significance of α = 0.05. A significant increase (p < 0.001) in applied light irradiance could be observed after individual instructions for both groups, with notably reduced data scattering. However, no significant difference was detected for the degree of conversion of placed composite restorations before or after instruction and training. Neither gender nor age of the dental students affected the obtained results. Consistent light energy delivered by dental students could be achieved through individual instructions and training with a patient simulator, also leading to less scattered irradiance results. However, the improved light-curing performance after the training did not affect the degree of conversion of the placed class II composite restorations. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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11 pages, 1122 KiB

Open AccessArticle

Using Cone-Beam Computed Tomography to Assess Changes in Alveolar Bone Width around Dental Implants at Native and Reconstructed Bone Sites: A Retrospective Cohort Study

by Kai-Fang Hu, Szu-Wei Lin, Ying-Chu Lin, Jiiang-Huei Jeng, Yu-Ting Huang, Pei-Feng Liu, Ching-Jiunn Tseng and Yu-Hsiang Chou

J. Pers. Med. 2021, 11(10), 1011; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm11101011 - 08 Oct 2021

Cited by 3 | Viewed by 2285

Abstract

The aim of this study was to use a cone-beam computed tomography (CBCT) to assess changes in alveolar bone width around dental implants at native and reconstructed bone sites before and after implant surgery. A total of 99 implant sites from 54 patients with at least two CBCT scans before and after implant surgery during 2010–2019 were assessed in this study. Demographic data, dental treatments and CBCT scans were collected. Horizontal alveolar bone widths around implants at three levels (subcrestal width 1 mm (CW1), subcrestal width 4 mm (CW4), and subcrestal width 7 mm (CW7)) were measured. A p-value of < 0.05 indicated statistically significant differences. The initial bone widths (mean ± standard deviation (SD)) at CW1, CW4, and CW7 were 6.98 ± 2.24, 9.97 ± 2.64, and 11.33 ± 3.00 mm, respectively, and the postsurgery widths were 6.83 ± 2.02, 9.58 ± 2.55, and 11.19 ± 2.90 mm, respectively. The change in bone width was 0.15 ± 1.74 mm at CW1, 0.39 ± 1.12 mm at CW4 (p = 0.0008), and 0.14 ± 1.05 mm at CW7. A statistically significant change in bone width was observed at only the CW4 level. Compared with those at the native bone sites, the changes in bone width around implants at reconstructed sites did not differ significantly. A significant alveolar bone width resorption was found only at the middle third on CBCT scans. No significant changes in bone width around implants were detected between native and reconstructed bone sites. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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10 pages, 2381 KiB

Open AccessArticle

Non-Syndromic Dentinogenesis Imperfecta Caused by Mild Mutations in COL1A2

by Yejin Lee, Youn Jung Kim, Hong-Keun Hyun, Jae-Cheoun Lee, Zang Hee Lee and Jung-Wook Kim

J. Pers. Med. 2021, 11(6), 526; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm11060526 - 08 Jun 2021

Cited by 5 | Viewed by 3179

Abstract

Hereditary dentin defects can be categorized as a syndromic form predominantly related to osteogenesis imperfecta (OI) or isolated forms without other non-oral phenotypes. Mutations in the gene encoding dentin sialophosphoprotein (DSPP) have been identified to cause dentinogenesis imperfecta (DGI) Types II and III and dentin dysplasia (DD) Type II. While DGI Type I is an OI-related syndromic phenotype caused mostly by monoallelic mutations in the genes encoding collagen type I alpha 1 chain (COL1A1) and collagen type I alpha 2 chain (COL1A2). In this study, we recruited families with non-syndromic dentin defects and performed candidate gene sequencing for DSPP exons and exon/intron boundaries. Three unrelated Korean families were further analyzed by whole-exome sequencing due to the lack of the DSPP mutation, and heterozygous COL1A2 mutations were identified: c.3233G>A, p.(Gly1078Asp) in Family 1 and c.1171G>A, p.(Gly391Ser) in Family 2 and 3. Haplotype analysis revealed different disease alleles in Families 2 and 3, suggesting a mutational hotspot. We suggest expanding the molecular genetic etiology to include COL1A2 for isolated dentin defects in addition to DSPP. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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18 pages, 3661 KiB

Open AccessArticle

Salivary Biomarkers for Dental Caries Detection and Personalized Monitoring

by Pune N. Paqué, Christopher Herz, Daniel B. Wiedemeier, Konstantinos Mitsakakis, Thomas Attin, Kai Bao, Georgios N. Belibasakis, John P. Hays, Joël S. Jenzer, Wendy E. Kaman, Michal Karpíšek, Philipp Körner, Johannes R. Peham, Patrick R. Schmidlin, Thomas Thurnheer, Florian J. Wegehaupt and Nagihan Bostanci

J. Pers. Med. 2021, 11(3), 235; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm11030235 - 23 Mar 2021

Cited by 20 | Viewed by 4913

Abstract

This study investigated the potential of salivary bacterial and protein markers for evaluating the disease status in healthy individuals or patients with gingivitis or caries. Saliva samples from caries- and gingivitis-free individuals (n = 18), patients with gingivitis (n = 17), or patients with deep caries lesions (n = 38) were collected and analyzed for 44 candidate biomarkers (cytokines, chemokines, growth factors, matrix metalloproteinases, a metallopeptidase inhibitor, proteolytic enzymes, and selected oral bacteria). The resulting data were subjected to principal component analysis and used as a training set for random forest (RF) modeling. This computational analysis revealed four biomarkers (IL-4, IL-13, IL-2-RA, and eotaxin/CCL11) to be of high importance for the correct depiction of caries in 37 of 38 patients. The RF model was then used to classify 10 subjects (five caries-/gingivitis-free and five with caries), who were followed over a period of six months. The results were compared to the clinical assessments of dental specialists, revealing a high correlation between the RF prediction and the clinical classification. Due to the superior sensitivity of the RF model, there was a divergence in the prediction of two caries and four caries-/gingivitis-free subjects. These findings suggest IL-4, IL-13, IL-2-RA, and eotaxin/CCL11 as potential salivary biomarkers for identifying noninvasive caries. Furthermore, we suggest a potential association between JAK/STAT signaling and dental caries onset and progression. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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11 pages, 1243 KiB

Open AccessArticle

Sagittal Relationship between the Maxillary Central Incisors and the Forehead in Digital Twins of Korean Adult Females

by Seoung-Won Cho, Soo-Hwan Byun, Sangmin Yi, Won-Seok Jang, Jong-Cheol Kim, In-Young Park and Byoung-Eun Yang

J. Pers. Med. 2021, 11(3), 203; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm11030203 - 13 Mar 2021

Cited by 15 | Viewed by 3163

Abstract

Objective: Digital twins of adult Korean females were created as a tool to evaluate and compare the sagittal relationship between the maxillary central incisors and the forehead before and after orthodontic treatment. Methods: Digital twins were reconstructed for a total of 50 adult female patients using facial scans and cone-beam computed tomography (CBCT) images. The anteroposterior position of the maxillary central incisor and the forehead inclination were measured. Results: The control group presented a mean of 6.7 mm for the sagittal position and 17.5° for forehead inclination. The study group showed a mean of 9.3 mm for the sagittal position and 13.6° for forehead inclination. Most Korean females seeking orthodontic treatment had their maxillary central incisor anterior to the glabella. In contrast, fewer Korean females who completed their orthodontic treatments had their maxillary central incisor anterior to the glabella. Furthermore, patients who had completed the orthodontic treatment were more likely to have the maxillary central incisor between the forehead facial axis and glabella. Conclusion: The use of digital twins for three-dimensional (3D) analysis of the profile implies a high clinical significance. In addition, as the facial profile of Koreans is different from that of Caucasians, careful consideration should be made when setting treatment goals for the anteroposterior position of the maxillary central incisors. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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9 pages, 21609 KiB

Open AccessArticle

Comparison between Additive and Subtractive CAD-CAM Technique to Produce Orthognathic Surgical Splints: A Personalized Approach

by Giuseppe Palazzo, Vincenzo Ronsivalle, Giacomo Oteri, Antonino Lo Giudice, Corrado Toro, Paola Campagna, Romeo Patini, Salvatore Bocchieri, Alberto Bianchi and Gaetano Isola

J. Pers. Med. 2020, 10(4), 273; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm10040273 - 11 Dec 2020

Cited by 7 | Viewed by 3860

Abstract

The present study aimed to evaluate the accuracy of digitally designed surgical splints generated with milling technology (material subtractive procedure) and with 3D printing technology (material additive procedure) through a customized approach in the planning of surgical orthognathic splints. Cone beam computed tomography (CBCT) examinations and scanned dental models of 10 subjects who had required surgical treatment of skeletal malocclusion were included. Simulation of the orthognathic surgery was performed according to dento-skeletal and aesthetic characteristics of the subjects and the visual treatment objective (VTO), using Dolphin3D software (Dolphin Imaging, version 11.0, Chatsworth, CA, USA). Afterward, the Appliance Designer software (3Shape A/S, Copenhagen, Denmark) was used to digitally design the surgical splints that were generated twice using laser stereolithography technology (DWS 0.29D, DWS, Vicenza, Italy) and milling technology (Sirona inLab MC X5). Finally, each physical splint was digitalized using a desktop scanner (D500 3D, 3Shape A/S, Copenhagen, Denmark) in order to perform deviation analysis using the original project as a reference. The relative percentage of matching (trueness) was calculated (Geomagic Control X software (3D Systems, version 2018.1.1, 3D Systems, Rock Hill, SC, USA). An Independent Student’s t-test was used to statistically analyze the data. The milled splints showed a lower value of root to mean square (RMS) relative to the original project (0.20 mm ± 0.018) compared to the prototyped splints (0.31 ± 0.021) (p < 0.001). According to the present findings, surgical splints generated with milling technology present higher trueness compared with 3D printing technology. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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Graphical abstract

8 pages, 263 KiB

Open AccessArticle

Bruxism Throughout the Lifespan and Variants in MMP2, MMP9 and COMT

by Alexandre R. Vieira, Rafaela Scariot, Jennifer T. Gerber, Juliana Arid, Erika C. Küchler, Aline M. Sebastiani, Marcelo Palinkas, Kranya V. Díaz-Serrano, Carolina P. Torres, Simone C. H. Regalo, Paulo Nelson-Filho, Diego G. Bussaneli, Kathleen Deeley and Adriana Modesto

J. Pers. Med. 2020, 10(2), 44; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm10020044 - 27 May 2020

Cited by 6 | Viewed by 3820

Abstract

Bruxism is a masticatory muscle activity characterized by grinding of the teeth and clenching of the jaw that causes tooth wear and breakage, temporomandibular joint disorders, muscle pain, and headache. Bruxism occurs in both adults and children. Clinical characteristics and habits were evaluated in an adult sample. Moreover, we used DNA samples from 349 adults and 151 children to determine the presence of association with specific genes. Genomic DNA was obtained from saliva. The markers rs2241145 and rs243832 (metalloproteinase 2 (MMP2)), rs13925 and rs2236416 (metalloproteinase 9 (MMP9)), and rs6269 (cathecol-o-methyltransferase (COMT)) were genotyped. Data were submitted to statistical analysis with a significance level of 0.05. In adults, in univariate logistic regression, presence of caries, attrition, and use of alcohol were increased in bruxism individuals (p < 0.05). In addition, in adults, there was an association between bruxism and MMP9 (rs13925, p = 0.0001) and bruxism and COMT (rs6269, p = 0.003). In children, a borderline association was observed for MMP9 (rs2236416, p = 0.08). When we performed multivariate logistic regression analyses in adults, the same clinical characteristics remained associated with bruxism, and orthodontic treatment was also associated, besides rs13925, in the AG genotype (p = 0.015, OR_a: 3.40 (1.27–9.07)). For the first time, we provide statistical evidence that these genes are associate with bruxism. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

9 pages, 2875 KiB

Open AccessArticle

Experimental Biointegration of a Titanium Implant in Delayed Mandibular Reconstruction

by Alexander Dolgolev, Igor Reshetov, Dmitry Svyatoslavov, Mikhail Sinelnikov, Konstantin Kudrin, Vladimir Dub, Vladimir Put and Vladimir Anikin

J. Pers. Med. 2020, 10(1), 6; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm10010006 - 03 Feb 2020

Cited by 6 | Viewed by 4905

Abstract

Background: Mandibular reconstruction, after extensive resection of the mandible for the treatment of oral cancer, is a well-known procedure, however, relatively little is known about bone integration into the titanium implant after reconstruction with a temporary plastic implant. The main goal of this experimental study was to study the process of osseous integration into the titanium implant in an in vivo experiment following prior mandibular reconstruction with a temporary plastic implant. Materials and Methods: Four ewes initially underwent a partial one-sided resection of the mandible, with the formation of an approximately 3 × 1 cm defect. All of the subjects received reconstruction with an implantation of a plastic plate (3 cm). The plastic plate was removed and replaced by a titanium implant at 1, 3, 6, and 12 months, accordingly. Both plastic and titanium implants were made via 3D-printing technology and personalized modeling. A total of 6 months after titanium implantation, a histological evaluation of biointegration was performed. Results: All surgeries were uncomplicated. The integration of osseous tissue into the titanium implant was seen in all cases. Histologically, each case showed variable integration of dense fibrotic tissue with fibroblasts and non-mature bone tissue with a definitive layer of bone matrix with many osteoblasts on the periphery. The prior implantation of the plastic plate did not interfere with bone integration into the titanium implant. Conclusion: Preliminary results demonstrated that a temporary plastic implant for mandibular reconstruction does not interfere with the consequent osseous biointegration of a permanent titanium implant. This shows that temporary reconstruction is a safe solution when delayed mandibular reconstruction is required due to disease severity. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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17 pages, 2823 KiB

Open AccessArticle

Measuring the Microscopic Structures of Human Dental Enamel Can Predict Caries Experience

by Ariana M. Kelly, Anna Kallistova, Erika C. Küchler, Helena F. Romanos, Andrea Lips, Marcelo C. Costa, Adriana Modesto and Alexandre R. Vieira

J. Pers. Med. 2020, 10(1), 5; https://0-doi-org.brum.beds.ac.uk/10.3390/jpm10010005 - 02 Feb 2020

Cited by 13 | Viewed by 6098

Abstract

Objectives: The hierarchical structure of enamel gives insight on the properties of enamel and can influence its strength and ultimately caries experience. Currently, past caries experience is quantified using the decayed, missing, filled teeth/decayed, missing, filled surface (DMFT/DMFS for permanent teeth; dmft/dmfs for primary teeth), or international caries detection and assessment system (ICDAS) scores. By analyzing the structure of enamel, a new measurement can be utilized clinically to predict susceptibility to future caries experience based on a patient’s individual’s biomarkers. The purpose of this study was to test the hypothesis that number of prisms by square millimeter in enamel and average gap distance between prisms and interprismatic areas, influence caries experience through genetic variation of the genes involved in enamel formation. Materials and Methods: Scanning electron microscopy (SEM) images of enamel from primary teeth were used to measure (i) number of prisms by square millimeter and interprismatic spaces, (ii) prism density, and (iii) gap distances between prisms in the enamel samples. The measurements were tested to explore a genetic association with variants of selected genes and correlations with caries experience based on the individual’s DMFT+ dmft score and enamel microhardness at baseline, after an artificial lesion was created and after the artificial lesion was treated with fluoride. Results: Associations were found between variants of genes including ameloblastin, amelogenin, enamelin, tuftelin, tuftelin interactive protein 11, beta defensin 1, matrix metallopeptidase 20 and enamel structure variables measured (number of prisms by square millimeter in enamel and average gap distance between prisms and interprismatic areas). Significant correlations were found between caries experience and microhardness and enamel structure. Negative correlations were found between number of prisms by square millimeter and high caries experience (r value= −0.71), gap distance between prisms and the enamel microhardness after an artificial lesion was created (r value= −0.70), and gap distance between prisms and the enamel microhardness after an artificial lesion was created and then treated with fluoride (r value= −0.81). There was a positive correlation between number of prisms by square millimeter and prism density of the enamel (r value = 0.82). Conclusions: Our data support that genetic variation may impact enamel formation, and therefore influence susceptibility to dental caries and future caries experience. Clinical Relevance: The evaluation of enamel structure that may impact caries experience allows for hypothesizing that the identification of individuals at higher risk for dental caries and implementation of personalized preventative treatments may one day become a reality. Full article

(This article belongs to the Special Issue Molecular Diagnosis and New Therapeutic Approach of Oral Diseases)

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