Guignon,RDH, MPH, CSP

Inﬂammation and the human microbiome - Stunning revelations about how microbes actually create disease

Anne N. Guignon,RDH, MPH, CSP

‣ in-depth health history ‣ tell the truth ‣ assess total needs ‣ provide all op+ons

What is a bioﬁlm?

A 3-Dimensional “community” of microbes a:ached to a surface

• ﬂuid interac=on • channels / pores • complex structure Illustration by Keith Kasnot,: Scientific American, July 2001

© 2019 Emerging Trends in Oral Care - Philips Oral Healthcare Symposium 2003 © 2019 Dr. Bill Costerton - The "Father" of Bioﬁms www.youtube.com/watch?v=M_DWNFFgHbE Dr. Bill Costerton - The "Father" of Bioﬁlms www.youtube.com/watch?v=M_DWNFFgHbE © 2019

Annu Rev Microbiol. 1995;49:711-45. millions of interac=ng microbes Microbial biofilms. Costerton JW, Lewandowski Z, Caldwell DE, Korber DR, Lappin-Scott HM. bacteria / spirochetes / protozoa / fungi / viruses Source Center for Biofilm Engineering, Montana State University, Bozeman 59717, USA. Abstract Direct observations have clearly shown that biofilm bacteria predominate, numerically and metabolically, in virtually all nutrient-sufficient ecosystems. Therefore, these sessile organisms predominate in most of the environmental, industrial, and medical problems and processes of interest to microbiologists. If biofilm bacteria were simply planktonic cells that had adhered to a surface, this revelation would be unimportant, but they are demonstrably and ‣ extracellular polysaccharide - slime profoundly different. We first noted that biofilm cells are at least 500 times more resistant to ‣ proper=es not seen in isolated micro-organisms antibacterial agents. Now we have discovered that adhesion triggers the expression of a sigma EPS - polysaccharides, proteins, nucleic acids factor that derepresses a large number of genes so that biofilm cells are clearly phenotypically ‣ distinct from their planktonic counterparts. Each biofilm bacterium lives in a customized ‣ 80% - infec=ous diseases microniche in a complex microbial community that has primitive homeostasis, a primitive ‣ 80% - EPS circulatory system, and metabolic cooperativity, and each of these sessile cells reacts to its ‣ 99% - bacteria in nature - stable, persistent special environment so that it differs fundamentally from a planktonic cell of the same The biofilm primer. J. W. Costerton 2007 species. Maddi A, Scannapieco FA. Oral biofilms, oral and periodontal infections, and systemic disease. Am J Dent. 2013 Oct;26(5):249-54. ‣ 20% - microbes encased in EPS matrix © 2019 © 2019 Biofilms in wounds: management strategies. Rhodes DD, Wolco: RD, Percival SL. J Wound Care. November 2008.17(11). © 2019 Wessel SW, Chen Y, et al. forces and composition of planktonic and adhering oral microbiomes. J Dent Res. 2014 Jan;93(1):84-8. Strength Where do biofilms form? Biofilm facts

- solid sub-stratums in ‣ mul=-species contact with moisture ‣ rapid regenera=on via signaling ‣ inter-species coopera=on ‣ tenacious - difficult to remove mechanically - so^ =ssue surfaces in living organisms intracellular matrix ‣ resistent - immune system / an=microbials ‣ - liquid-air interfaces - the surface of the ocean ‣ high adhesion to saliva-coated enamel ‣ FEP - func=onally equivalent pathogenicity The biofilm primer. J. W. Costerton 2007 Maddi A, Scannapieco FA. Oral biofilms, oral and periodontal infec=ons, and systemic disease. Am J Dent. 2013 Oct;26(5):249-54. Biofilms in wounds:management strategies. Rhodes DD, Wolcott RD, Percival SL. J Wound Care. November 2008.17(11). The biofilm primer. J. W. Costerton 2007 Wessel SW, Chen Y, et al. forces and composi=on of planktonic and adhering oral microbiomes. J Dent Res. 2014 Jan;93(1):84-8. Oliveira A, Cunha MLRS. Bacterial biofilms with emphasis on coagulase-negative staphylococci. J. Venom. Anim. Toxins incl. Trop. Dis. 2008.14(4): 572-596. http://www.scielo.br © 2019 © 2019 © 2019

Mixed bioﬁlm community - proportions Forming a bioﬁlm

‣ cri=cal density - release pheromones ‣ quorum sensing ‣ altered environment - phenotypic altera=ons in microbes

Bioﬁlms in wounds: management strategies. Rhodes DD, Wolco: RD, Percival SL. J Wound Care. November 2008.17(11). Bonnie Bassler: The secret, social lives of bacteria 4/8/2009 h:p://www.youtube.com/watch?v=TVfmUfr8VPA © 2019 © 2019 Costalonga M, Herzberg MC. The oral microbiome and the immunobiology of periodontal disease and caries. Immunology le6ers. 2014;162(2 0 0):22-38. = © 2019

Mixed biofilm community - Candida albicans (opportunis=c fungal pathogen) Liquid flowing through biofilm channels

‣ coadhesion ‣coaggrega=on ‣modiﬁed: ‣pH ‣nutrient supply ‣salivary factors

‣highly acidic pH The Great wave of Candida C. albicans, C. glabrata, C. parapsilosis © 2001 Stoodley, P. MSU Center for Biofilm Engineering Jenkinson HF and Douglas LJ. Interactions between Candida Species and Bacteria in Mixed Infections. www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=pmd&part=A2773#A Accessed 7/25/2009

Falsetta ML1, Klein MI, Symbiotic relationship between Streptococcus mutans and Candida albicans synergizes virulence of plaque bioﬁlms in vivo. Infect Immun. 2014 May;82(5):1968-81.© 2019 Cris=na Marcos © 2019 Emerging Trends in Oral Care - Philips Oral Healthcare Symposium 2003 © 2019

Moving biofilm “ripples” in Viscoelastic Properties of S. aureus Biofilm Growth and spontaneous mixed species biofilm detachment of cell cluster

Tap water bioﬁlm

20 hour sequence

P. aeruginosa P. fluorescens K. pneumoniae - common on skin S. maltophilia - natural flora - oral / nasal / vaginal - opportunis=c pathogen - nosocomial infec=ons © 2001 Stoodley, P. MSU Center for Biofilm Engineering © 2001 Stoodley, P. MSU Center for Biofilm Engineering Emerging Trends in Oral Care - Philips Oral Healthcare Symposium 2003 Emerging Trends in Oral Care - Philips Oral Healthcare Symposium 2003 © 2019 © 2019 © 2019

Shear induced creep and failure

http://www.erc.montana.edu/bioﬁlmbook/MODULE_01/Mod01_Blue/Mod01_S04_Blue.htm Accessed 7/25/09 © 2019 © 2019 http://www.erc.montana.edu/bioﬁlmbook/MODULE_01/Mod01_Blue/Mod01_S04_Blue.htm Accessed 7/25/09 © 2019

Understanding the trouble makers - microbial defenses that make it hard to treat disease

http://www.npuap.org/NPUAP%20Bioﬁlms%202009%20Schultz%20mod%202%20compressed-1.pdf

Bertesteanu S, Triaridis S, et al. Polymicrobial wound infec=ons: pathophysiology and current therapeu=c approaches. Int J Pharm. 2014 Mar 25;463(2):119-26. . © 2019 h:p://www.erc.montana.edu/bioﬁlmbook/MODULE_07/Mod07_S03_Blue.htm Accessed 7/25/09 © 2019 Layers pH and oxygen levels vary in bioﬁlms

surface microbes - most ac+ve

‣ most suscep=ble - an=microbials/host defenses

deeper layers - sheltered

‣ less metabolically ac=ve ‣ more resistant - an=microbial therapies ‣ can recons=tute bioﬁlm (persisters)

http://www.cbe.montana.edu/biofilmbook/MODULE_07/Mod07_S01-2_Blue.htm Biofilms in wounds:management strategies. Rhodes DD, Wolcott RD, Percival SL. J Wound Care. November 2008.17(11). http://www.erc.montana.edu/biofilmbook/MODULE_01/Mod01_Blue/Mod01_S04_Blue.htm Accessed 7/25/09 © 2019 © 2019 © 2019

Resistance Antibiotic resistance - 1,000x greater adapts to stresses resistant to ‣as compared planktonic ‣ ultraviolet light ‣to antibiotics and biocides ‣ biocides biofilm-related infections ‣ an=bio=cs can’t be treated with ‣ host defenses conventional antibiotic therapy

management - mul+ple, concurrent strategies

http://www.npr.org/sections/health-shots/2016/09/08/492965889/watch-bacteria-invade-antibiotics-and-transform-into-superbugs? Bioﬁlms in wounds:management strategies. Rhodes DD, Wolcott RD, Percival SL. J Wound Care. November 2008.17(11). Sauer K, Thatcher E. et al. Biofouling 2009;25(1):45-54. © 2019 © 2019 utm_campaign=storyshare&utm_source=twitter.com&utm_medium=social

Chronic wounds Chronic wound infections Host impairment

‣ immune system can’t clear ‣ barrier to healing ‣ repe==ve trauma ‣ an=microbial resistant ‣ anaerobes - chronic wounds ‣ hyperglycemia ‣ suscep=ble to contamina=on ‣ poor perfusion ‣ systemic / topical ‣ molecular techniques ‣ presence of a foreign body ‣ available substrate ‣ malnutri=on ‣ frequent debridement cri+cal ‣ white blood cell disfunc=on ‣ diverse popula+ons ‣ surface for a:achment ‣ pressure

www.erc.montana.edu/biofilmbook/MODULE_07/Mod07_S04-2_Blue.htm Accessed 11/11/09 © 2019 Biofilms in wounds: management strategies. Rhodes DD, Wolco: RD, Percival SL. J Wound Care. November 2008.17(11). © 2019 www.erc.montana.edu/biofilmbook/MODULE_07/Mod07_S04-2_Blue.htm Accessed 11/11/09 © 2019 Chronic wound infections Oral biofilm survival - Ecological plaque hypothesis - inflammatory periodontal disease making inflammation a lifestyle

‣ polymicrobial bioﬁlms

‣ clinical aspects resemble other biofilm infec=ons Marsh PD, Mo:er A, Devine D. Dental plaque biofilms: Communi=es, conflict and control. Periodontology 2000, Vol. 55, 2011, 16–35. www.erc.montana.edu/biofilmbook/MODULE_07/Mod07_S04-2_Blue.htm Accessed 11/11/09 © 2019 © 2019

Inﬂammation Red complex microbes

‣ T. den=cola - long thin spirochete creates bridges

‣ P. gingivalis - grape-like ‣ originates - bacterial assault coccobacillus surface vesicles ‣ inﬂammatory response - most damage ‣ we used to think...... Only a handful of oral bacteria ‣ T. (Bactericides) forsythus were implicated in periodontal disease larger fusiform rod

Könönen E, Müller HP. Microbiology of aggressive periodon==s. Periodontol 2000. 2014 Jun;65(1):46-7. © 2019 Zhu Y, Dashper SG, Chen YY, Crawford S, Slakeski N, et al. (2013) Porphyromonas gingivalis and Treponema denticola Synergistic Polymicrobial Biofilm Development. PLoS ONE 8(8): e71727. doi:10.1371/journal.pone.0071727 © 2019 Periasamy S1, Kolenbrander PE. Mutualis=c bioﬁlm communi=es develop with Porphyromonas gingivalis and ini=al, early, and late colonizers of enamel. J Bacteriol. 2009 Nov;191(22):6804-11. http://journals.plos.org/plosone/article?id=info:doi/10.1371/journal.pone.0071727 © 2019

Inﬂammation Inﬂammation

Periodon==s ‣ requires suscep=ble host ‣ fosters dysbio=c growth ‣ dysbio+c microbial communi=es ‣ selects for certain pathogens ‣ inflammo-phillic ‣ dysbiosis and inflamma+on support each other ‣ inflamma=on - provides nutrients control of inflamma+on - cri+cal Hajishengallis G. Immunomicrobial pathogenesis of periodon==s: keystones, pathobionts, and host response. Immunol. 2014 Jan;35(1):3-11. Hajishengallis G. Immunomicrobial pathogenesis of periodon==s: keystones, pathobionts, and host response. Immunol. 2014 Jan;35(1):3-11. Hajishengallis G. The inflammophilic character of the periodon==s-associated microbiota.Mol Oral Microbiol. 2014 Jun 26. © 2019 Hajishengallis G. The inflammophilic character of the periodon==s-associated microbiota.Mol Oral Microbiol. 2014 Jun 26. © 2019 Hajishengallis G. Periodon==s: from microbial immune subversion to systemic inflamma=on. Nature reviews Immunology. 2015;15(1):30-44. © 2019 Polymicrobial synergy - dysbiosis (PSD) - Polymicrobial synergy - dysbiosis (PSD) - model of periodontal disease etiology model of periodontal disease etiology

Keystone pathogen Keystone pathogens P. gingivalis ‣ low abundance microbe ‣ =p the balance to dysbiosis ‣ modulates host response ‣ pro-inﬂammatory microbial community ‣ impairs host immune system ‣ elicit non-resolving, =ssue-destruc=ve host response P. gingivalis

Hajishengallis G., Lamont RJ. Beyond the red complex and into more complexity: the polymicrobial synergy and dysbiosis (PSD) model of periodontal disease e=ology. Hajishengallis G., Lamont RJ. Beyond the red complex and int o more complexity: the polymicrobial synergy and dysbiosis (PSD) model of periodontal disease e=ology.

Costalonga M, Herzberg MC. The oral microbiome and the immunobiology of periodontal disease and caries. Immunology le6ers. 2014;162(2 0 0):22-38. © 2019 Mol Oral Microbiol. 2012 Dec: 27(6);407-19. © 2019 Mol Oral Microbiol. 2012 Dec: 27(6);407-19. © 2018

Polymicrobial synergy - dysbiosis (PSD) - Inﬂammation model of periodontal disease etiology

Keystone pathogen

P. gingivalis (Pg) P. gingivalis creates pathobionts ‣ 4 hours a^er scaling ‣ elevates community virulence ‣ early, middle and late stage bioﬁlm

P. gingivalis ‣ sugar capsule Thurnheer T, Belibasakis GN, Bostanci N. Colonisa=on of gingival epithelia by subgingival biofilms in vitro: Role of "red complex" bacteria. Arch Oral Biol. 2014 Sep;59(9):977-86. Oral Biol. 2014 Sep;59(9):977-86. hides proteins from immune system Arora N, Mishra A, Chugh S. Microbial role in periodon==s: Have we reached the top? Some unsung bacteria other than red complex. J Indian Soc Periodontol. 2014 Jan;18(1):9-13. Hajishengallis G., Lamont RJ. Beyond the red complex and int o more complexity: the polymicrobial synergy and dysbiosis (PSD) model of periodontal disease e=ology. Mol Oral Microbiol. 2012 Dec: 27(6);407-19. Mahendra J, Mahendra L, Prevelance of periodontopathogenic bacteria in subgingival biofilm and atherosclero=c plaques of pa=ents undergoing coronary © 2019 Petersen C, Round JL. Defining dysbiosis and its influence on host immunity and disease. Cellular Microbiology. 2014;16(7):1024-1033. © 2019 revasculariza=on surgery. J Indian Soc Periodontol. 2013 Nov;17(6):719-24. © 2019

Polymicrobial synergy Synergy Fusobacterium nucleatum T. denticola (Td) (Fn)

P.gingivalis and T. den=cola T. denticola from a periodontal pocket P. gingivalis ‣ anaerobic oral commensal ‣ superficial layers - subgingival plaque ‣ mo=le ‣ perio and endo pathogen ‣ chronic periodon==s lesions ‣ creates pores - biofilm matrix ‣ increases with disease severity, ‣ increase biomass and thickness ‣ moves through highly viscous environment inflamma=on, pocket depth ‣ pores enhance nutrient flow ‣ higher in smokers, diabe=cs T. denticola Zhu Y, Dashper SG, et al. Porphyromonas gingivalis and Treponema den=cola synergis=c polymicrobial biofilm development. PLoS One. 2013 Aug 26;8(8):e71727.h:p:// bacmap.wishartlab.com/organisms/70#kegg-pathways - Accessed 1-31-16 Han YW. Fusobacterium nucleatum: a commensal-turned pathogen. Current opinion in microbiology. 2015;0:141-147. Zhu Y, Dashper SG, et al. Porphyromonas gingivalis and Treponema den=cola synergis=c polymicrobial biofilm development. PLoS One. 2013 Aug 26;8(8). © 2019 © 2019 © 2019 FINAL REPORT

FINAL REPORT First parasitic Ordering Provider Guignon, Anne Sample Information (43000133) Doe, John Ordering Provider Sample Information Parasitic outcomes? Date Of Birth: 08/20/1949 Terri Alani DDS Accession: 10244004 Received: 11/22/2010 11:52 Date Of Birth: 09/20/1980 (36 yrs) Ronald McGlennen MD Specimen#: 3022131004 Collected: 11/12/2016 10:30 Gender: Female 5636 WEstheimer Specimen: Oral Rinse Reported: 11/24/2010 07:56 Gender: Male 7400 Flying Cloud Drive Accession#: 201611-08469 Received: 11/13/2016 12:01 Houston, TX Collected: 11/18/2010 Printed: 11/24/2010 07:57 Patient Id: Eden Prairie, MN 55344 Specimen: Oral Rinse(P) Reported: 11/16/2016 12:30 77056 713-651-5141 Patient Location: 855-672-5362 MYPERIOPATH MOLECULAR ANALYSIS OF PERIODONTAL AND SYSTEMIC PATHOGENS relationship Result: NORMAL - NO PATHOGENS REPORTED ABOVE THRESHOLD Bacterial Risk: LOW - Some evidence of increased risk for attachment loss Result: PATHOGENIC BACTERIA DETECTED, 5 ABOVE THERAPEUTIC THRESHOLD Bacterial Risk: HIGH - Very strong evidence of increased risk for attachment loss Legend Result Interpretation: Periodontal disease is caused by specific, or groups of specific bacteria. Threshold Legend Result Interpretation: Periodontal disease is caused by specific, or groups of specific bacteria. = Pathogen Load Threshold* levels represent the concentration above which patients are generally at increased risk for attachment loss. Threshold levels represent the concentration above which patients are generally at increased risk for = Therapeutic Threshold* DL = Detection Limit Bacterial levels should be considered collectively and in context with clinical signs and other risk factors. DL = Detection Limit attachment loss. Bacterial levels should be considered collectively and in context with clinical signs and other risk factors. High Risk Pathogens Moderate Risk Pathogens Low Risk Pathogens High Risk Pathogens Moderate Risk Pathogens Low Risk Pathogens discovered

107 107 107 107 107 107 6 6 6 106 106 106 10 10 10 5 5 5 105 105 105 10 10 10 4 4 4 104 104 104 10 10 10 DL DL DL DL DL DL

Pathogen Result Clinical Significance Pathogen Result Clinical Significance Aggregatibacter High Very strong association with PD: Transmittable, tissue invasive, and pathogenic at relatively low actinomycetemcomitans bacterial counts. Associated with aggressive forms of disease.

Porphyromonas gingivalis High Very strong association with PD: Transmittable, tissue invasive, and pathogenic at relatively low bacterial counts. Associated with aggressive forms of disease. Fusobacterium High Strong association with PD: adherence properties to several oral pathogens; often seen in refractory T. denticola from a periodontal pocket nucleatum/periodonticum disease. Tf Tannerella forsythia Low Very strong association with PD: common pathogen associated with refractory periodontitis. Strongly Prevotella intermedia High Strong association with PD: virulent properties similar to Pg; often seen in refractory disease. related to increasing pocket depths. ‣ Bdellovibrio - RNA fragment Capnocytophaga species High Some association with PD: Frequently found in gingivitis. Often found in association with other (gingavalis,ochracea,sputigena) periodontal pathogens. May increase temporarily following active therapy.

Cr Campylobacter rectus Low Moderate association with development of PD: usually found in combination with other suspected Pi Prevotella intermedia Low Strong association with PD: virulent properties similar to Pg; often seen in refractory disease. pathogens in refractory disease. ‣ A. odontoly=cus evades macrophages Pm Peptostreptococcus (Micromonas) Low Moderate association with PD: detected in higher numbers at sites of active disease. micros Ec Eikenella corrodens Low Moderate association with PD: Found more frequently in active sites of disease; often seen in ‣ 700 genes - can’t make amino acids refractory disease.

Not Detected: (Tf) Tannerella forsythia, (Td) Treponema denticola, (En) Eubacterium nodatum Not Detected: (Aa) Aggregatibacter actinomycetemcomitans, (Pg) Porphyromonas gingivalis, (Td) Treponema denticola, (En) Eubacterium nodatum, (Fn) Fusobacterium Additional information is available from OralDNA.com nucleatum/periodonticum, (Cr) Campylobacter rectus, (Pm) Peptostreptococcus Methodology: Genomic DNA is extracted from the submitted sample and tested for 10 species-specific bacteria and 1 genus of bacteria known to cause periodontal resistant to streptomycin (Micromonas) micros, (Ec) Eikenella corrodens, (Cs) Capnocytophaga species ‣ disease. The bacteria are assayed by real-time quantitative polymerase chain reaction (qPCR). Bacterial loads are reported in log copies per mL of sample (e.g. 1x10^3 (gingavalis,ochracea,sputigena) = 1000 bacteria copies per mL of collection). *Modified from: Microbiological goals of periodontal therapy; Periodontology 2000, Vol. 42, 2006, 180-218. This test was developed, and its performance characteristics determined by OralDNA Labs pursuant to CLIA requirements. This test has not been cleared or approved by the U.S. lives on AcKnomyces odontolyKcus (2,200 genes) Food and Drug Administration. The FDA has determined that such clearance or approval is not necessary. ‣ Additional information is available from MyOralDNA.com on Interpreting Results

Methodology: Genomic DNA is extracted from the submitted sample and tested for 13 bacteria associated with periodontal disease. The bacteria are tested by polymerase chain reaction (PCR) amplification followed by fluorescent endpoint detection of sample bacterial concentrations (e.g. 10^3 = 1000 bacteria copies per amplified reaction). *Modified from: Microbiological goals of periodontal therapy; Periodontology 2000, Vol. 42, 2006, 180-218. Disclaimer: 1. OralDNA is not liable for any outcomes arising from clinician's treatment protocols and decisions. Dentists should consult with a periodontist or patient's physician when infections are advanced or high concentraKon - periodonKKs and cysKc ﬁbrosis as indicated by patient's medical condition. 2. OralDNA is not responsible for inaccurate test results due to poor sample collection. 3. This test was developed and its performance characteristics determined by ‣ OralDNA Labs, Inc pursuant to CLIA requirements. It has not been cleared or approved by the U.S. Food and Drug Administration. The FDA has determined that such clearance or approval is not necessary. nutrients from host, kills host, holes in A.odontolyKcus OralDNA Labs, Inc., 214 Overlook Circle, Suite 120, Brentwood, TN 37027 6155779055; Fax: 6156272826 www.oraldna.com ‣ Medical Director: Ronald McGlennen, MD Chief Dental Officer: Thomas W. Nabors, DDS Page 1 of 2 OralDNA Labs, A Service of Access Genetics, LLC, 7400 Flying Cloud Drive , Eden Prairie, MN 55344 Phone: 855-672-5362; Fax: 952-942-0703 www.oraldna.com CLIA#: 24D1033809 CAP#: 7190878 1 of 2 Web enabled system provided by: Coghlan A. New life form discovered in saliva is linked to human disease. h:ps://www.newscien=st.com/ar=cle/2094902-new-life-form-discovered-in- © 2019 saliva-is-linked-to-human-disease/ - Accessed 6-23-16 Guignon, Anne Sample Information © 2019 © 2019 Coghlan A. New life form discovered in saliva is linked to human disease. h:ps://www.newscien=st.com/ar=cle/2094902-new-life-form-discovered-in-saliva-is-linked-to-human-disease/ Accessed 6-23-16

•Clean hydroxyapa=te surface Improving the patient’s outcome Telling the story…… •Pulsed with human saliva •8 hour =me lapse sequence •Early colonizers - Strep and having fun Rods and ﬁlamentous organisms • individual best •Nutrient and waste channels form clinical external expertise evidence EBM

patient values & expectations

Health history clues 2013

‣ medica=ons - poly pharmacy ‣ an=bio=cs - prolonged use compromise intes=nal flora ‣ acid reducers - alter intes=nal tract pH 2015 ‣ 500,000 infec=ons annually ‣ 29,000 die within one month ‣ most common microbial cause - health-care associated infec=ons Saving lives © 2019 © 2019 https://www.cdc.gov/media/releases/2015/p0225-clostridium-difficile.html © 2019 Links to systemic disease….. Characteristics of biofilm infections what can a clinician do?

Evaluate individually ‣ their risks ‣ willing to do ‣ mo=vated to change

Aging/inﬂammation….Gut dysbiosis? Gut dysbiosis……Chronic inﬂammation? Oral microbes ……impact

Buford TW. (Dis)Trust your gut: the gut microbiome in age-related inflamma=on, health, and disease. Microbiome. 2017;5:80. doi:10.1186/s40168-017-0296-0. © 2019 Buford TW. (Dis)Trust your gut: the gut microbiome in age-related inflamma=on, health, and disease. Microbiome. 2017;5:80. doi:10.1186/s40168-017-0296-0. © 2019 Hajishengallis G. Periodon==s: from microbial immune subversion to systemic inflamma=on. Nature reviews Immunology. 2015;15(1):30-44. © 2019

Oral microbes - systemic infections / inﬂammation ‣ perio disease - bacteremia ‣ greater microbial diversity ‣ extra-oral link - associa=on ‣ cardiovascular disease ‣ virulence varies ‣ adverse pregnancy outcomes ‣ respiratory tract infec=ons ‣ rheumatoid arthri=s ‣ meningi=s - brain accesses ‣ inﬂammatory bowl disease ‣ lung, liver, spleen accesses ‣ colorectal cancer ‣ appendici=s

Han YW, Wang X. Mobile Microbiome: Oral Bacteria in Extra-oral Infec=ons and Inﬂamma=on. Journal of Dental Research. 2013;92(6):485-491. © 2019 Winning L, Linden GL. Periodon==s and systemic disease. BJD Team. November 27, 2015 © 2019 Han YW, Wang X. Mobile Microbiome: Oral Bacteria in Extra-oral Infections and Inﬂammation. Journal of Dental Research. 2013;92(6):485-491. © 2019 Cardiovascular disease - atherosclerosis Bacterial signatures - MI thrombus

‣ systemic spread of oral microbes - 1954 DNA evidence ‣ perio pathogens - Aa, Pg, Tf, Td and Fn ‣ 78 % - S. Viridans (endodon=c) ‣ acute and chronic inﬂamma=on ‣ 35% - periodontal pathogens ‣ periodontal instrumenta=on (40%) ‣ extrac=ons (35%) 9 thrombi - bacterial components ‣ brushing (24%) ‣ thrombus - load 16x higher ‣ mas=ca=on (17%) ‣ whole bacteria - 3 of 9 thrombi

Bale BR, Donen AL, Vigerust DJ. High risk periodontal pathogens contribute ot the pathogenesis of athrerosclerosis. Postgrad Med J. First published November 29, 2016. Pessi T1, Karhunen V, et al. . Bacterial signatures in thrombus aspirates of pa=ents with myocardial infarc=on.Circula=on. 2013 Mar 19;127(11):1219-28, e1-6. © 2019 © 2019 Cleveland Heart Lab 2017. © 2019

Cardiovascular disease medical condi+on with a dental solu+on Inﬂammation - Pre-term birth Pregnancy - Low birth weight - Association

‣ high risk PD pathogens ‣ bacteremia ‣ pathogen burden ‣ inﬂammatory mediators ‣ causal - clinical grounds ‣ vagina to placental microbiome ‣ contribute to ASVD ‣ therapy - reduce risk

‣ Fusobacterium nucleatum ‣ P. Gingivalis ‣ independent risk factor ‣ young ‣ preeclampsia invades host cells amnio=c ﬂuid - premature delivery ‣ severe or generalized - ‣ HIV-infected ‣ pre-pregnancy obesity intrauterine infec=on placenta - preeclampsia promotes PTB ‣ gene=c predisposi=on Bale BR, Donen AL, Vigerust DJ. High risk periodontal pathogens contribute ot the pathogenesis of athrerosclerosis. Postgrad Med J. First published November 29, 2017. Cobb CM, Kelly PJ, et al. The oral microbiome and adverse pregnancy outcomes. InternaKonal Journal of Women’s Health. 2017;9:551-559. © 2019 .Ren H, Du M. Role of Maternal Periodon==s in Preterm Birth. FronKers in Immunology. 2017;8:139. © 2019 © 2019

Erectile dysfunction - positive association Macular Degeneration - Association Pancreatic cancer risk

‣ common risk factors - ‣ oral dysbiosis precedes cancer diabetes, smoking, alcohol, heart disease ‣ endothelial dysfunc=on - inﬂamma=on ‣ P. gingivalis - 59% higher ‣ perio - possible risk factor for ED ‣ A. ac=nomycetemcomitans - 50% higher

Perio treatment - Improvement ‣ findings are not causal ‣ plaque index ‣ BOP ‣ suscep=bility to inflamma=on? ‣ Pg - can invade human re=nal- ‣ probing depths pigment epithelial cells ‣ microbes causing inflamma=on? ‣ clinical a:achment ‣ Pg - elevate AMD-related genes Jacob JA. Study links periodontal disease bacteria to pancrea=c cancer risk. JAMA. 2016;315(24):2653-2654. Eltas A, Oguz F, Uslu MO, Akdemir E. The effect of periodontal treatment in improving erec=le dysfunc=on: a randomized controlled trial. J Clin Periodontol. 2013 Feb;40(2):148-54. Interna=onal & American Associa=ons for Dental Research. "Inves=ga=ng the enigma=c link between periodontal inflamma=on and re=nal degenera=on." P. Maisonneuve S. Amar A. B. Lowenfels Periodontal disease, edentulism, and pancrea=c cancer: a meta-analysis Annals of Oncology, Singh VP, Ne:emu SK, Ne:em S, Hosadurga R, Nayak SU. Oral Health and Erec=le Dysfunc=on. J Hum Reprod Sci. 2017 Jul-Sep;10(3):162-166. © 2019 ScienceDaily. March 23, 2018. © 2019 Volume 28, Issue 5, 1 May 2017, Pages 985–995, © 2019

Esophageal squamous cell carcinoma risk Exposure to oral pathogens is associated with Alzheimers disease systemic inﬂammaKon, cogniKve decline, and AD.

In the cascade of events causing AD, oral ‣ P. gingivalis - 61% cancerous =ssues microorganisms may play a role, parKcularly anaerobic bacteria such as treponemes, P. gingivalis, ‣ inﬂamma=on ‣ 12% surrounding =ssues Prevotella spp., Fusobacterium and AcKnomyces, but also facultaKve anaerobic Candida species. ‣ microbes pass through ‣ associated with progression on ESCC It is important to recognize that infecKon can occur compromised blood vessels? decades before the manifestaKon of demenKa.

‣ ﬁndings are not causal ‣ diﬀerent bacterial popula=ons ‣ PG a disease biomarker? ‣ Alzheimer’s disease ‣ Parkinson’s, Alcoholic demen=a ‣ increased bacterial load ‣ Vascular demen=a ‣ Lewey body disease ‣ oral perio microbes

Gao S, Li S, Ma Z, et al. Presence of Porphyromonas gingivalis in esophagus and its association with the clinicopathological characteristics and survival in Harris SA, Harris EA. Herpes Simplex Virus Type 1 and Other Pathogens are Key Causa=ve Factors in Sporadic Alzheimer’s Disease. Mancuso R, ed. Journal of Alzheimer’s Disease. 2015;48(2):319-353. © 2019 patients with esophageal cancer. Infect Agent Cancer. 2016 Jan 19;11:3. Olsen I, Singhrao SK. Can oral infec=on be a risk factor for Alzheimer’s disease? Journal of Oral Microbiology. 2015;7:10.3402/jom.v7.29143.© 2019 © 2019 Emery DC, Shoemark DK, et al. (2017) 16S rRNA Next GeneraKon Sequencing Analysis Shows Bacteria in Alzheimer’s Post-Mortem Brain. Front. Aging Neurosci. 9:195.

Health implications - Health implications aspirated bioﬁlm Aspirated bioﬁlm

‣ cuﬀ on endo tube - bioﬁlm

‣ oral gel reduced contamina=on

‣ small scratches - hard to clean

‣ retained microbes - proliferate - appliance reinserted

http://www.bioﬁlm.montana.edu/Res-Lib99-SW/Image_Library/Medical%20&%20Health/default.htm Accessed 7/25/09 © 2019 Takeyasu Y, Yamane GY, et al. Ven=lator-associated Pneumonia Risk Decreased by Use of Oral Moisture Gel in Oral Health Care. Bull Tokyo Dent Coll. 2014;55(2):95-102. Winning L, Linden GL. Periodon==s and systemic disease. BJD Team. November 27, 2015 © 2019

! Nuts Links to systemic disease….. ! Avocados what can a clinician do? ! Spinach ! Tart cherries ! Olive oil Increase ! Orange vegetables and fruits Evaluate individually inflammation - ! Pineapple ! Tumeric, ginger onions, garlic ‣ their risks ‣ willing to do ‣ mo=vated to change ! processed meats ! gluten Individualize your recommenda:ons ! fried food ! casein ‣ establish a dialogue with other healthcare providers ! so^ drinks, high fructose corn syrup ! oils - safflower, sunflower, corn

‣ 28% - age 20-34 ‣ 26% - age 35-59 ‣ 28% age 2-5 ‣ 22% - age 50-64 ‣ 50% by age 11 28% men, 23% women ‣ 68% at age 19 ‣ 18% - age 65+

20% men, 16% women New ideas - about caries bioﬁlm nidcr.nih.gov/DataSta=s=cs/FindDataByTopic/DentalCaries/DentalCariesAdults20to64.htm nidcr.nih.gov/DataSta=s=cs/FindDataByTopic/DentalCaries/DentalCariesSeniors65older.htm Accessed 7/28/2015 © 2019 h:p://www.cdc.gov/OralHealth/publica=ons/factsheets/dental_caries.htm 7/28/2015 © 2019

Ecological plaque hypothesis - caries Predisposing risk factors Predisposing risk factors

! medium/high levels of cariogenic microbes ! new research – caries bioﬁlm more complex ! heavy / visible plaque accumula=ons ! bacteria on diﬀerent surfaces ! snacking - 3+ =mes / day

Caries risk assessment appropriate for the age 1 visit. J Calif Dent Assoc. 2007 ! mutans streptococci dominate fissures Caries risk assessment in prac=ce from age 6 through adult. J Calif Dent Assoc. 2007 Bacteria of dental caries on primary and permanent teeth in children and young adults. Aas JA, et al.J clin Microbiol. 2008 Apr;46(4):1407-17. Bacteria of dental caries on primary and permanent teeth in children and young adults. Aas JA, et al.J clin Microbiol. 2008 Apr;46(4):1407-17. Marsh PD. Dental plaque as a biofilm and a microbial community - implica=ons for health and disease. BMC Oral Health 2006, 6(Suppl 1):S14. profiles of root caries in elderly pa=ents. Preza D, et al. J Clin Microbiol. 2008 Jun;46(6):2015-21. Bacterial profiles of root caries in elderly pa=ents. Preza D, et al. J Clin Microbiol. 2008 Jun;46(6):2015-21. © 2019 Researchers: S. Mutans bacteria not the problem. Harrison L. drbicuspic.com Accessed 9/1/09 © 2019 S. Mutans bacteria not the problem. Harrison L. drbicuspic.com Accessed 9/1/09 © 2019

New news about caries biofilm Ecological plaque Ecological plaque Strep mutans not the only organism - hypothesis hypothesis Different microbes colonize ! white spots ! den=n lesions ! health = homeostasis ! increase mutans streptococci (gateway) ! root caries ! disease sites species differ from health ! den=n lesions = lactobacillus + proteoly=c microbes ! primary and secondary den==on ! caries - shi^ - acidogenic / aciduric ! specific tooth surfaces ! sustained low pH - select for Ms and Lb Asa JA, Griffen AL, et al. Bacteria of dental caries in primary and permanent teeth in children and young adults. J of Clin Microbiol. April 2008. 46(4):1407-17. ! imbalance in resident microflora Preza D, Olden I, et al. Microarray analysis of the microflora of root caries in elderly. Eur J Clin Infec iDs. May 2009. 28(5). Cavalcan= YW, Bertolini MM, et al. A three-species biofilm model for the evalua=on of enamel and den=n demineraliza=on. Biofouling. 2014;30(5)579-88. Marsh PD. Dental plaque as a biofilm and a microbial community - implica=ons for health and disease. BMC Oral Health 2006, 6(Suppl 1):S14. Marsh PD. Dental plaque as a biofilm and a microbial community - implica=ons for health and disease. BMC Oral Health 2006, 6(Suppl 1):S14. © 2018 Asa JA, Griffen AL, et al. Bacteria of dental caries in primary and permanent teeth in children and young adults. J of Clin Microbiol. April 2008. 46(4):1407-17. © 2018 © 2018 Costalonga M, Herzberg MC. The oral microbiome and the immunobiology of periodontal disease and caries. Immunology letters. 2014;162(2 0 0):22-38. doi: Asa JA, Griffen AL, et al. Bacteria of dental caries in primary and permanent teeth in children and young adults. J of Clin Microbiol. April 2008. 46(4):1407-17.

More news - Microbial adaptation caries bioﬁlm

! S. Mutans - early colonizer ! “good” bacteria mutate / adapt ! 30% rampant caries - do not test posi=ve for S.Mutans ‣ low income children - 2 X decay ! become acid producers to survive-pathobionts ‣ 4 X more common than asthma (42% versus 9.5%) ! adapt in 30 minutes to survive low pH ! children with ECC ‣ 52,000,000 lost school hours/year - dental disease high levels of S. mutans and C. albicans Takahashi N, Nevada B. Caries ecology revisited: Microbial dynamics and the caries process. Caries Res. 2008;42(6):409-18. False:a ML, Klein MI, Symbio=c rela=onship between Streptococcus mutans and Candida albicans synergizes virulence of plaque bioﬁlms in vivo. Infect Immun. 2014 May;82(5):1968-81. Takahashi N, Nevada B. Caries ecology revisited: Microbial dynamics and the caries process. Caries Res. 2008;42(6):409-18. © 2018 h:p://www.cdc.gov/OralHealth/publica=ons/factsheets/dental_caries.htm 7/28/2015 False:a ML, Klein MI, Symbio=c rela=onship between Streptococcus mutans and Candida albicans synergizes virulence of plaque bioﬁlms in vivo. Infect Immun. 2014 May;82(5):1968-81. © 2018

Oral appliances Cigarettes Micro Ultrasonics….using today’s technology ‣ dentures / par=als ➡increase C. albicans adhesion/growth/bioﬁlm forma=on ‣ metal bands, brackets, acrylic resin, elas=c rings ‣ bite guards cigare:e smoke condensate ‣ orthodon=c aligners ‣ increased adhesion - S. mutans - bands, acrylic and brackets ‣ sports mouth guards C. albicans - bands, acrylic, brackets, elas=cs ‣ increased biomass - S. mutans - only brackets ‣ whitening trays C. albicans - on brackets and acrylic

‣ surface roughness ‣ nico=ne-S. mutans - increase growth and ESP

Baboni FB1, Guariza Filho O, Moreno AN, Rosa EA. Inﬂuence of cigare:e smoke condensate on cariogenic and candidal bioﬁlm forma=on on orthodon=c ‣ Strep and Candida adhered materials. Am J Orthod Dentofacial Orthop. 2010 Oct;138(4):427-34.

‣ reinsert - microbes proliferate Semlali A, Killer K,et. al. Cigare:e smoke condensate increases C. albicans adhesion, growth, biofilm forma=on, and EAP1, HWP1 and SAP2 gene expression. BMC Microbiol. 2014 Mar 12;14:61. Verran J, Jackson S, The effect of den=frice abrasion on denture topography and the subsequent reten=on of microorganisms on abraded surfaces. J Prosthet Dent. 2014 Jul 3. Bains VK, Mohan R, Bains R. Applica=on of ultrasound in periodon=cs: Part I.J Indian Soc Periodontol. 2008 May-Aug; 12(2): 29–33. Przybyłowska D1, Rubinsztajn R,et al. Prevalence of Oral Inflamma=on Among Denture Wearing Pa=ents with Chronic Obstruc=ve Pulmonary Disease. Adv Exp Med Biol. 2015;858:87-91 © 2018 Li M, Huang R,et al. Effect of nico=ne on dual-species biofilms of Streptococcus mutans and Streptococcus sanguinis. FEMS Microbiol Le:. Jan;350(2):125-32. Bains VK, Mohan R, Bains R. Applica=on of ultrasound in periodon=cs: Part II. J Indian Soc Periodontol. 2008 Sep;12(3):55-61.

Clinical beneﬁts - Structural differences Other reasons to use contemporary sensitive and non-sensitive dentin a polishing handpiece ultrasonic scalers

• applying desensi=zing medicaments • disrupt plaque biofilm and calculus • pre-procedural or site specific • reduce bacterial level below immune system threshold Non-sensi=ve Sensitive No of open tubules x 8 x applica=on • pocket disinfec=on Diameter of tubules 0.43 0.83 Fluid Flow (Poisseuille’s law) y 16 y • pre-scaling biofilm reduc=on • improved access – complex anatomy, deep, narrow pockets *Absi et al, J Clin Periodont 1987; pictures from h:p://www.thejcdp.com, Sept 2006 Bains VK, Mohan R, Bains R. Applica=on of ultrasound in periodon=cs: Part I.J Indian Soc Periodontol. 2008 May-Aug; 12(2): 29–33. • reduce ultrasonic aerosol bioburden Bains VK, Mohan R, Bains R. Applica=on of ultrasound in periodon=cs: Part II. J Indian Soc Periodontol. 2008 Sep;12(3):55-61. Acoustic microstreaming Cavitation / Bubbles How cavitation works Amplitude tip displacement

cavita=on = ultrasonic energy + water ! 10.5 microns ! collapsing into small fragments crea=on of micro-bubbles ! shock waves

acous=c micro-streaming !bubble on solid surface !deforming high velocity jet liquid implosion of micro-Bubbles ! !pierces and damages ! 47.5 microns destruc=on of bioﬁlm and calculus Laird W.R.E. and Walmsley A.D,1991). Laird W.R.E. and Walmsley A.D,1991). www.shockwavetherapy.ca Images accessed 4-4-2011 K. Bains,1, et al. Applica=on of ultrasound in periodon=cs: Part I.J Indian Soc Periodontol. 2008 May-Aug; 12(2): 29–33. K. Bains,1, et al. Applica=on of ultrasound in periodon=cs: Part I.J Indian Soc Periodontol. 2008 May-Aug; 12(2): 29–33.

When are we done? Blending scaling techniques Subgingival bioﬁlm disruption - glycine

‣ power and hand or hand and power • glycine powder • all root surfaces - contact with ac=vated =p • so^, small par=cle size factors include: type of power scaler, condi=on of =ps, • no visible debris ‣ • reduced surface damage clinical challenge, type of deposit, pa=ent comfort • lowers bioﬁlm adhesion • no clinically detec=ble deposits

skill level / familiarity with power scaling techniques Moëne R1, Décaillet F, Andersen E, Mombelli A. Subgingival plaque removal using a new air-polishing device.J Periodontol. 2010 Jan;81(1):79-88.: 10.1902. ‣ Giacomelli L, Salerno M, Derchi G, Genovesi A, Paganin PP, Covani U. Eﬀect of air polishing with glycine and bicarbonate powders on a nanocomposite used in dental restora=ons: an in vitro study. Int J Periodon=cs Restora=ve Dent. 2011; 31(5):51-56. © 2018

Subgingival bioﬁlm disruption - glycine Silver diamine ﬂuoride 38%

‣ natural an=bacterial • comfortable ‣ hypersensi=vity relief ‣ carious den=n lesion turns black 5-9 mm pockets • ‣ no anesthesia

• lower P. gingivalis counts

more eﬀec=ve in removing subgingival bioﬁlm than SCP alone

Flemmig TF, Arushanov D, et al. Randomized controlled trial assessing efficacy and safety of glycine powder air polishing in moderate-to-deep periodontal pockets.J Periodontol. 2012 Apr;83(4):444-52. Sculean A, Bastendorf KD, et al. A paradigm shi^ in mechanical biofilm management? Subgingival air polishing: a new way to improve mechanical biofilm management in the dental prac=ce. © 2018 Healthy biofilm - getting rid of pathogens Rosenbla: A, Stamford TC, Niederman R. Silver diamine fluoride: a caries "silver-fluoride bullet”.J Dent Res. 2009 Feb;88(2):116-25. © 2018 Quintessence Int. 2013 Jul;44(7):475-7. © 2018 Evidence-Based Clinical Practice Guideline on Nonrestorative Treatments for Carious Lesions: Evidence-Based Clinical Practice Guideline on Nonrestorative Treatments for Carious Lesions: A Report from the American Dental Association A Report from the American Dental Association Summary of clinical recommendations for the nonrestorative treatment of caries on primary teeth Silver diamine fluoride 38% Clinical Pathway for the Nonrestorative Treatment of Carious Lesions on Primary Teeth GRADE Certainty in the Evidence GRADE Interpretation of Strength of Recommendations Primary Teeth We are very confident that the true Implications Strong Recommendations Conditional Recommendations High effect lies close to that of the estimate of the effect. For Patients Most individuals in this situation would want the The majority of individuals in this situation would want the suggested Coronal Surface recommended course of action and only a small course of action, but many would not. We are moderately confident in the Moderate effect estimate. The true effect is likely proportion would not. to be close to the estimate of the effect. For Clinicians Most individuals should receive the intervention. Recognize that different choices will be appropriate for individual patients Our confidence in the effect estimate and that you must help each patient arrive at a management decision Occlusal Facial or Lingual Approximal Low is limited. consistent with his or her values and preferences. For Policy The recommendation can be adapted as policy in Policy making will require substantial debate and involvement of various Very Low We have very little confidence in the effect estimate. Makers most situations. stakeholders. Noncavitated* Cavitated† Noncavitated* Cavitated† Noncavitated* Cavitated† Certainty in Strength of Expert Panel Recommendation the Evidence Recommendation • Sealants + 5% NaF • 1.23% APF Gel‡, §, • 5% NaF Varnish‡, § To arrest advanced cavitated carious lesions on any coronal surface of primary teeth, the expert panel Moderate Strong Varnish‡, §, or or Alone, or recommends clinicians* prioritize the use of 38% silver diamine fluoride (SDF) solution (biannual application) • Sealants Alone • 5% NaF Varnish‡ over 5% sodium fluoride varnish (application once per week for 3 weeks).† • Resin Infiltration Before SDF Application Alone, or To arrest or reverse noncavitated carious lesions on occlusal surfaces of primary teeth, the expert panel Moderate Strong § • Resin Infiltration + recommends clinicians* prioritize the use of sealants + 5% sodium fluoride varnish (application every 3-6 months) If not feasible ‡ or sealants alone over 5% sodium fluoride varnish alone (application every 3-6 months), 1.23% acidulated phosphate 5% NaF Varnish , or fluoride gel (application every 3-6 months), resin infiltration + 5% sodium fluoride varnish (application every 3-6 • Sealants Alone months), or 0.2% sodium fluoride mouthrinse (once per week).‡ • 5% NaF Varnish‡ To arrest or reverse noncavitated carious lesions on facial or lingual surfaces of primary teeth, the expert panel Moderate Conditional Alone, or ‣ arrests caries suggests clinicians* use 1.23% acidulated phosphate fluoride gel (application every 3-6 months) or 5% sodium to Low • 1.23% APF Gel‡, or After SDF Application fluoride varnish (application every 3-6 months).‡ • Resin Infiltration + To arrest or reverse noncavitated carious lesions on approximal surfaces of primary teeth, the expert panel Low to Conditional ‡ 5% NaF Varnish , or #, * * suggests clinicians* use 5% sodium fluoride varnish (application every 3-6 months), resin infiltration alone, Very Low 38% SDF Solution less than $1 / 1-2 teeth ‡ ‣ resin infiltration + 5% sodium fluoride varnish (application every 3-6 months), or sealants alone. • 0.2% NaF Mouthrinse¶ To arrest or reverse noncavitated carious lesions on coronal surfaces of primary teeth, the expert panel suggests Low Conditional Lesion(s) should be monitored (e.g., hardness/texture, color, clinicians* do not use 10% casein phosphopeptide-amorphous calcium phosphate paste if other fluoride interventions, radiographs) periodically throughout the course of treatment ‣ 8ml bo:le sealants, or resin infiltration is accessible. SDF = silver diamine fluoride NaF = sodium fluoride * Defined as International Caries Detection and Assessment System (ICDAS) 1 and 2 lesions. ¶At-home use once per week. * “Clinicians” refers to the target audience for this guideline, but only those authorized/trained to perform the specified interventions Copyright ©2018 American Dental Association. All rights reserved. Adapted with permission. APF = acidulated phosphate fluoride † Defined as ICDAS 5 and 6 lesions. #Biannual application. SDF = silver diamine fluoride ‡ Application every 3-6 months. ** In keeping with the concept of informed consent, all nonrestorative and restorative treatment should do so. Photos courtesy of the University of Washington’s Travis Nelson, D.D.S., M.S.D., M.P.H. To see § The order of treatments included in this recommendation represents a ranking of priority defined by options and their potential side effects (such as blackened tooth surfaces treated with SDF) † In keeping with the concept of informed consent, all nonrestorative and restorative treatment options and their potential side effects full text of this article, please go to JADA.ADA.org/article/S0002-8177(18)30469-0/fulltext . the panel when accounting for treatment effectiveness, feasibility, patients’ values and preferences, should be offered and explained to all patients. anyone who can apply fluoride (such as blackened tooth surfaces treated with silver diamine fluoride) should be offered and explained to all patients. This page may be used, copied, and distributed for non-commercial purposes without obtaining and resource utilization. Considerations such as a particular patient’s values and preferences, special ‣ ‡ The order of treatments included in this recommendation represents a ranking of priority defined by the panel when accounting for prior approval from the ADA. Any other use, copying, or distribution, whether in printed or elec- needs, or insurance status should inform clinical decision making. treatment effectiveness, feasibility, patients’ values and preferences, and resource utilization. Considerations such as a particular tronic format, is strictly prohibited without the prior written consent of the ADA. patient’s values and preferences, special needs, or insurance status should inform clinical decision making.

ebd.ada.org/en/evidence/guidelines/nonrestorative-treatments-for-caries-lesions. Accessed 9/25/18 Rosenbla: A, Stamford TC, Niederman R. Silver diamine ﬂuoride: a caries "silver-ﬂuoride bullet”.J Dent Res. 2009 Feb;88(2):116-25. ebd.ada.org/en/evidence/guidelines/nonrestorative-treatments-for-caries-lesions. Accessed 9/25/18

When faced with this……. Oral microﬂora Survival strategies How fast does it grow?

mature bioﬁlm - 24 hours ‣ ‣ 1 mg plaque = 100,000,000 microbes ‣ mechanical disrup=on - rapid recovery ‣ en=re mouth = 20,000,000,000 microbes - at any given =me ‣ 2-4 days - fully mature ‣ 1 liter saliva swallowed daily - 100 billion microbes ‣ high biocide resistance ‣ oral bacteria - double every 4.8 hours!

‣ shedding planktonic bacteria Loesche, WJ. Dental caries: a treatable infec=on. Springﬁeld, IL: Charles C. Thomas; 1982. p. 558 Phillips PL, Wolco: RD, Fletcher j, Schul GS. Bioﬁlms made easy. May 2010. Vol 1. Issue 3. woundsinterna=onal.com

HydrogenHydrogen Peroxide Peroxide concentrations -≤3%

‣ antiseptic ‣ debridement Approved for oral use by FDA ‣ reduces inflammation Effect on biofilm ‣ degrades EPS - slime ‣ breaks down protein chains naturally occurs - breast milk, saliva, liver ‣ ‣ lyses bacterial cell walls ‣ softens hard deposits ‣ no allergic reactions ‣ oxygen - death to anaerobes ‣ no known bacterial resistance Marshall MV, Cancro LP, Fischman SL. Hydrogen peroxide: a review of its use in dentistry. J Periodontol. 1995 Sep;66(9):786-96. Herrero ER, Slomka V, Boon N, et al. Dysbiosis by neutralizing commensal mediated inhibition of pathobionts. Scientiﬁc Reports. 2016;6:38179. © 2018 Hydrogen peroxide - comparison Prescription trays 1.7% peroxide gel - S. mutans biofilm

Problem: “Therapeutic delivery of H2O2 to prevent periodontal disease required mechanical access to subgingival pockets.”

‣ trays create hydraulic seal

‣ eliminates dilution Untreated control 5 min contact 1.7% H2O2 10 min contact 1.7% H2O2 Placebo gel - no H2O2 up to 9mm ‣ ‣ S. mutans aerobic - will not die in oxygen ‣ 3% liquid ‣ 1.7% gel ‣ thick biofilm mass ‣ 10 min therapeutic ‣ pH 3 ‣ pH 5.5 - 5.8 ‣ live/dead dye: intact cell walls - green 15 min full O2 release ‣ degraded, lysed wall - red ‣ rapid spike - bubbling ‣ 15 min active time Allegheny Singer Study

© 2018 Dunlap T et al. Subgingival Delivery of Oral Debriding Agents: A Proof of Concept. The Journal of Clinical Den=stry. 2011, Nov;XXII(5):149-158. Dunlap T et al. Subgingival Delivery of Oral Debriding Agents: A Proof of Concept. The Journal of Clinical Den=stry. 2011, Nov;XXII(5):149-158.

Prescription tray therapy

January 2015 July 2016 June 8, 2016 July 7, 2016 tray delivery - 4/2016 ‣ ‣ 3 times a day ‣ 101 bleeding points to 7 only PerioGel ‣ ‣ 15 min per session ‣ 1-2mm probing reduction twice daily - 15 min ‣ ‣ PerioGel only Photos courtesy of Bruce Cochrane, DDS - Fort Dodge IA Photos courtesy of Bruce Cochrane, DDS - Fort Dodge IA Photos courtesy of Perfect Teeth - Denver, Colorado

Making it hard on the Ecological plaque hypothesis - caries bad bugs…… encouraging healthier microﬂora Prebiotics - prebiotics stimulate growth or activity of certain healthy bacteria

Probiotics -a microorganism introduced into the body for its beneﬁcial qualities

Marsh PD. Dental plaque as a bioﬁlm and a microbial community - implica=ons for health and disease. BMC Oral Health 2006, 6(Suppl 1):S14. McLean JS. Advancements toward a systems level understanding of the human oral microbiome. Fron=ers in Cellular and Infec=on Microbiology. 2014;4:98. Hutkins RW, Krumbeck JA, Bindels LB, et al. Prebio=cs: why deﬁni=ons ma:er. Current opinion in biotechnology. 2016;37:1-7. doi:10.1016/j.copbio.2015.09.001. Interesting herbal approach What is arginine? Arginine - mode of action

★ licorice root extract

★ 2x per day

★ 10 days ! natural amino acid ★ 2-4 times annually ! naturally found in saliva ★ stimulates saliva ! bipolar molecule - + and - groups ★ reduces Lactobacillus Urea%!"few"bacteria"" Arginine"!"many"bacteria" ! net posi=ve charge "saliva"&"crevicular"fluid" "low"in"saliva/abundant"in"pep:des" and S. mutans "broken"down"by"urea" "ADS"!"3"enzyme"system"" Ahn SJ, Park SN, Lee YJ, et al. .In vitro an=microbial ac=vi=es of 1-methoxyficifolinol, licorisoflavan A, and 6,8-diprenylgenistein against Streptococcus mutans.Caries Res. 2015;49(1):78-89. "byproduct!ammonia"" "byproduct"!"ammonia"" Ahn SJ, Cho EJ, et al.The an=microbial effects of deglycyrrhizinated licorice root extract on Streptococcus mutans UA159 in both planktonic and biofilm cultures. Anaerobe. "ac:on"produces"ATP Panagakis F, Schiff T, Guignon A. Den=n hypersensi=vity: Effec=ve treatment with an in-office desensi=zing paste containing 8% arginine and calcium carbonate. Amer J Dent. 2009. 22:March. © 2018 2012 Dec;18(6):590-6. Burne"RA,"Zeng"L,"Ahn"SJ,"et"al."Progress"Dissec:ng"the"Oral"Microbiome"in"Caries"and"Health."Advances"in"Dental"Research."2012;24(2):77!80."

Arginine - mode of action Arginine bicarbonate Emerging research - Arginine calcium carbonate calcium carbonate tooth paste - lower DMFS ! inhibits bacterial adhesion - tooth surfaces ! reduces bioﬁlm thickness

! exogenous source - toothpaste ! 6,000 children - low to moderate risk - 6-12 years old ! reduces EPS matrix density

! enhances alkaline pH - saliva and plaque ! double blind, randomized - 3 groups - 2 year study ! arginine + ﬂuoride - suppresses S. mutans and P. gingivalis

! 4 weeks ! 1,450 ppm Fl paste ! suppresses C. albicans growth

! higher alkali production - plaque samples - caries active subjects ! 1,450 Fl + 1.5% arginine/calcium carbonate or dicalcium phosphate ! facilitates microbial resistance - acidic environment

Fu D, Pei D, Huang C, Liu Y, Du X, Sun H. Effect of desensi=zing paste containing 8% arginine and calcium carbonate on biofilm forma=on of Streptococcus mutans in vitro. J Dent. 2013 Jul;41(7):619-27. Sharma S, Lavender S, Woo J, Guo L, Shi W, Kilpatrick-Liverman L, Gimzewski JK. Nanoscale characteriza=on of effect of L-arginine on Streptococcus mutans biofilm adhesion by atomic force microscopy. ! CA subjects - bacterial shift - healthier ! 16.5% lower DMFS - arginine/calcium/fluoride groups than FL paste alone Microbiology. 2014 Jul;160(Pt 7):1466-73. Zheng X, Cheng X, Wang L, Qiu W, Wang S, Zhou Y, Li M, Li Y, Cheng L, Li J, Zhou X, Xu X. Combinatorial effects of arginine and fluoride on oral bacteria. J Dent Res. 2015 Feb;94(2):344-53. Burne RA, Zeng L, Ahn SJ, Palmer SR, Liu Y, Lefebure T, Stanhope MJ, Nascimento MM. Progress dissec=ng the oral microbiome in caries and health. Adv Dent Res. 2012 Sep;24(2):77-80. Kraivaphan P, Amornchat C, Triratana T, Mateo LR, Ellwood R, Cummins D, DeVizio W, Zhang YP. Two-year caries clinical study of the efficacy of novel den=frices containing Nascimento MM, Browngardt C, et al. The effect of arginine on oral biofilm communi=es. Mol Oral Microbiol. 2014 Feb;29(1):45-54.1. Koopman JE, Röling WF, Buijs MJ, Sissons CH, ten Cate JM, Keijser BJ, Crielaard W, Zaura E. Stability and resilience of oral microcosms toward acidifica=on and Candida outgrowth by arginine Huang X, Schulte RM, Burne RA, Nascimento MM. Characteriza=on of the arginoly=c microflora provides insights into pH homeostasis in human oral biofilms. Caries Res. 2015;49(2):165-76. © 2018 1.5% arginine, an insoluble calcium compound and 1,450 ppm fluoride. Caries Res. 2013;47(6):582-90. supplementa=on. Microb Ecol. 2015 Feb;69(2):422-33.

Arginine-based products Arginine bicarbonate calcium carbonate - Remineralization strategies keeping saliva neutral

Discon=nued!

High ﬂuoride ! varnish, gels Beneﬁts Precipita=ng salts ! calcium phosphate ! less =me arginine bicarbonate less pa=ent discomfort ! Applica=on ! ! pa=ent acceptance ! 2+ =mes a year ! adhesives ! silicates ! preschool / adolescents / geriatrics Restora=ve materials Home ! caries preven=on ! resins ! hydroxyapa=te ! high risk popula=ons Professional

!so^laser Petersson LG. The role of fluoride in the preven=ve management of den=n hypersensi=vity and root caries. Clin Oral Inves=g. 2013 Mar;17 Suppl 1:S63-71. Murakami C, Bönecker M,et al. Effect of fluoride varnish and gel on dental erosion in primary and permanent teeth. Oral Biol. 2009 Nov;54(11):997-1001. Laser Ri:er AV, de L Dias W, Miguez P, Caplan DJ, Swi^ EJ Jr. cervical den=n hypersensi=vity with fluoride varnish: a randomized clinical study. J Am Dent Assoc. 2006 Jul;137(7). © 2017 Zero DT, Lussi A. Erosion-chemical and biological factors of importance to the dental prac==oner. Int Dent J. 2005;55(4 Suppl 1):285-90. Evidence-based Clinical Recommenda=ons: Professionally Applied Topical Fluoride Council on Scien=fic Affairs. American Dental AssociaKon May, 2006 © 2018

Patient-centered Chemotherapeutic strategies choices Mouth rinses – non-selec=ve "Essen=al oils "Chlorhexidine ! dry teeth / mix product ! paint all suscep=ble surfaces Toothpaste – non-selec=ve ! no wait =me - ea=ng/drinking "Stannous ﬂuoride ! 24 hour uptake =me ! applica=on - moist or in plaque ! horizontal swipe Toothpaste – gels

García-Godoy F, Hicks MJ. Maintaining the integrity of the enamel surface: the role of dental bioﬁlm, saliva and preven=ve agents in enamel demineraliza=on and remineraliza=on. ! © 2018 surface migra=on More solutions J Am Dent Assoc. 2008 May;139 Suppl:25S-34S.

Mechanical disruption Mechanical disruption ! 60 day trial ! plaque reduc=on - p< .001 ! reduc=on in hsCRP - p< .04

! biofilm turns green ! mint / berry bubble gum Feel a new ! .24% sodium fluoride level of clean Rmaile A, Carugo D, Removal of interproximal dental biofilms by high-velocity water microdrops. J Dent Res. 2014 Jan;93(1):68-73. Stoodley, P. MSU Center for Biofilm Engineering Ren Y, Jongsma MA,et al. Orthodon=c treatment with fixed appliances and biofilm forma=on-a poten=al public health threat? Clin Oral Inves=g. 2014 Apr 13. © 2018 Fasula K, Evans CA, et al. Randomized trial of plaque identifying toothpaste: Dental plaque and inflammation, The American Journal of Medicine (2016), © 2017 3 seconds exposure 99.9% removal

“The results were almost impossible for me to believe the first time through,” commented . “One of the difficulties with plaque biofilm is that you really can’t see it, it’s clear. So we didn’t have visual evidence of complete removal. But now with these direct methods, the scanning electron microscopy, you apply the Waterpik to plaque on a surface of a tooth and you look with a scanning scope and it’s gone. It’s simply gone. And that’s unequivocal and unarguable.”

Tongue Getting Tongue cleaners national press

! daily ! effec=ve ! manual Slots J, Jorgensen MG. Efficient an=microbial treatment in periodontal maintenance care. JADA, Vol.131, September 2000 ! mechanical Rupesh S, Winnier J, et al. The compara=ve evalua=on of the effects of tongue cleaning on salivary levels of mutans streptococci in children. Int J Dent Hyg. 2011 Jul 29. Oral malodor. JADA, Vol.134, February 2003 Bordas A, McNab R, et al. Impact of different tongue cleaning methods on the bacterial load of the tongue dorsum. Arch Oral Biol. 2008 Apr;53 Suppl 1:S13-8.

Nocturnal storage - Ecological plaque hypothesis Removable appliances acrylic appliances

! N=51 3 randomized groups ! contaminated with MRSA ! dry / water /water H2O2 tablet target putaKve pathogens (difficult) ! 4 groups - 1% bleach or 2% CHX - 10 min soak or 3 min in microwave ! biofilm samples - day 7 & 14 ! CHx and microwave - complete disinfec=on ! s=mulate saliva ! raise pH ! 3 min. daily soaking - denture cleanser ! tablets decrease - developing, maturing biofilms ! reduced bacterial counts / increased C. albicans levels ! tablets decrease biofilm mass / specific bacteria / C. Albicans ! limit sugar intake

Al=eri KT, Sanitá PV,et a. Eﬀec=veness of two disinfectant solu=ons and microwave irradia=on in disinfec=ng complete dentures contaminated with methicillin-resistant Staphylococcus poten=al eﬀects - stoma==s / systemic health aureus.J Am Dent Assoc. 2012 Mar;143(3):270-7. !

Lucena-Ferreira SC, Ricomini-Filho AP, et al. Influence of daily immersion in denture cleanser on mul=species biofilm. Clin Oral Inves=g. 2014 Mar 4. Marsh PD. Dental plaque as a biofilm and a microbial community - implica=ons for health and disease. BMC Oral Health 2006, 6(Suppl 1):S14. Duyck J, Vandamme K,et al. Overnight storage of removable dentures in alkaline peroxide-based tablets affects biofilm mass and composi=on. J Dent. 2013 Dec;41(12):1281-9. © 2018 García-Godoy F, Hicks MJ. Maintaining the integrity of the enamel surface: the role of dental biofilm, saliva and preven=ve agents in enamel and remineraliza=on. J Am Dent Assoc. 2008. © 2019 ! non-fermentable sweeteners Medical biofilm-based ! metabolic inhibitors - fluoride Understanding biofilm - impacting lives wound care

• debridement- frequent and aggressive ! an=-adhesion - xylitol • selec=ve biocides ! arginine products silver, Iodosorb, Hydrofera Blue • an=bioﬁlm agents lactoferrin, xylitol, farnasol Mul+ple concurrent strategies plant products, fa:y acid gel

•an=bio=cs (25-32% eﬀec=ve) adjunct strong and long

Marsh PD. Dental plaque as a biofilm and a microbial community - implica=ons for health and disease. BMC Oral Health 2006, 6(Suppl 1):S14. Biofilms in wounds:management strategies. Rhodes DD, Wolcott RD, Percival SL. J Wound Care. November 2008.17(11). García-Godoy F, Hicks MJ. Maintaining the integrity of the enamel surface: the role of dental biofilm, saliva and preven=ve agents in enamel and remineraliza=on. J Am Dent Assoc. 2008. © 2018

Wound healing Wound healing Wound healing - anti-bioﬁlm strategies anti-bioﬁlm agents antimicrobial agents ‣ lactoferrin - bacteriocidal / block surface a:achment / works with PMNs /binds iron even at a low pH

xylitol - interferes with biofilm forma=on 7-26-04 ‣ • ionic silver - minute ‣ enzymes - Dispersin B - causes biofilm detachment concentra=ons / broad spectrum / used in dressings ‣ gallium, EDTA, hyperbaric oxygen, bacteriophages, glucose oxidase, pulsed electric fields iodine - prevents wound infec=on / aids healing / 1-13-05 • povodone iodine preferred ‣ ultrasound / ultrasonics

5-25-05 • medicinal honey - an=bacterial ‣ no more amputa+ons!! Rhodes DD, Wolco: RD, Percival SL. J Bioﬁlms in wounds:management strategies. Wound Care. November 2008.17(11). Rhodes DD, Wolco: RD, Percival SL. J Bioﬁlms in wounds:management strategies. Wound Care. November 2008.17(11).

Biofilms in wounds:management strategies. Rhodes DD, Wolcott RD, Percival SL. J Wound Care. November 2008.17(11). Percival SL, Cuâng KF, Williams D(2010) Biofilms: possible strategies for suppression in chronic wounds. Nursing Standard Supplement. Percival SL, Cuâng KF, Williams D(2010) Biofilms: possible strategies for suppression in chronic wounds. Nursing Standard Supplement.

Honey

broad spectrum an=microbial an=-inﬂammatory ﬁrst recorded use over 4,500 years ago

‣ eﬀec=ve against P. gingivalis ‣ domes=c / Manuka (tea tree plant) honey ‣ both planktonic and bioﬁlm organisms ‣ inhibited forma=on / reduced viable numbers - both groups

Pieper B. Honey-based dressings and wound care: an op=on for care in the United States. J Wound Ostomy Con=nence Nurs. 2009 Jan-Feb;36(1):60-6. Eick S, Schäfer G, Kwieciński J, et al. Honey - a poten=al agent against Porphyromonas gingivalis: an in vitro study. BMC Oral Health. 2014 Mar 25;14(1):24. Dr. Bill Costerton - The "Father" of Bioﬁlms h:p://www.youtube.com/watch?v=lbLFOUHvAJg Ancient ideas and emerging technologies Molan PC. The evidence suppor=ng the use of honey as a wound dressing. Int J Low Extrem Wounds. 2006 Mar;5(1):40-54. Red Wine Silver - natural antimicrobial Red wine study What do we • 5-species supragingival bioﬁlm ‣ natural an=bacterial • regular red wine / de-alcoholized / extract owe our patients? ‣ bacteriocidal • regular solu=ons / solu=ons spiked with grape ‣ an=-fungal seed extract

Tannins - 72 hour study Products ‣ bandages • Pseudomonas aeruginosa - opportunis=c pathogen ‣ paper ‣ catheters ‣ current scien=fic informa=on • bacteriosta=c ‣ paint ‣ dental implants • bacterial membrane damage ‣ plas=cs ‣ endotrachel tubes ‣ pa=ents must make the final choice inhibited biofilm forma=on 6- 24 hours ‣ tex=les ‣ toothbrushes • How would you treat your Mom? • impaired EPS matrix produc=on ‣ orthodon=c wires Muñoz-González, Thurnheer T, et al. Red wine and oenological extracts display an=microbial effects in an oral bacteria biofilm model. J Agric Food Chem. 2014 May 21;62(20):4731-7. Maillard JY, Hartemann P. Silver as an antimicrobial: facts and gaps in knowledge. Crit Rev Microbiol. 2013 Nov;39(4):373-83. Tren=n DS, Silva DB, et al. Tannins possessing bacteriosta=c effect impair Pseudomonas aeruginosa adhesion and biofilm forma=on.PLoS One. 2013 Jun 11;8(6):e66257. © 2018

What’s the take home message? Understand......

‣ bioﬁlms are complex ‣ Mother Nature rules ‣ be:er health is possible ‣ take responsibility for your role ‣ new informa:on emerges every day ‣ share with your colleagues [email protected] 832-971-4540