Bio fi lms được công nhận là một trong những hệ sinh thái sớm nhất trên trái đất. Chúng bao gồm các tập hợp các tế bào vi sinh vật được bao bọc trong một chất nền tự sản sinh bám dính vào một bề mặt. Sinh học ống tủy là những cấu trúc đa vi khuẩn phức tạp bám vào bề mặt ống tủy được hình thành do vi sinh vật xâm nhập vào khoảng trống của răng. Các nghiên cứu mô bệnh học quan trọng được công bố cách đây vài thập kỷ lần đầu tiên ghi nhận sự hiện diện của các tế bào bám dính trên bề mặt ống tủy. Tuy nhiên, phải đến khi các kỹ thuật hiển vi và sinh học phân tử ra đời, chúng mới được công nhận là dạng vi sinh vật chiếm ưu thế trong đời sống vi sinh vật trong hệ thống ống tủy. Tương tự, chỉ trong thập kỷ trước, nhiễm trùng tủy răng được thừa nhận là nhiễm trùng sinh học. Sau đó, các nghiên cứu gần đây đã chỉ ra rằng sinh học ống tủy có liên quan đến nhiễm trùng nội nha dai dẳng và như vậy có khả năng không phải là yếu tố góp phần quyết định kết quả của điều trị nội nha. Những nỗ lực quan tâm để nghiên cứu sinh học ống tủy đã được thực hiện trong thập kỷ qua dẫn đến việc công bố các nghiên cứu quan sát và thực nghiệm trình bày chi tiết hình thái và sinh học của các cấu trúc này trong ống tủy bị nhiễm trùng. Ngoài việc xác minh rằng vi khuẩn trong ống tủy không tồn tại ở trạng thái phù du tự do như đã giả định trước đây, thông tin mới này về nhiễm trùng sinh học ống tủy đã tạo cơ hội để đánh giá lại các phác đồ lâm sàng thông thường và cải thiện các biện pháp điều trị nội nha. Mục đích của tập này là cung cấp sự hiểu biết hiện tại về các khía cạnh khoa học cơ bản của sinh học ống tủy răng trong bối cảnh có thể áp dụng được trên lâm sàng. Tập này được chia thành ba phần. Phần I thảo luận về sinh học cơ bản của sinh học ống tủy và giải quyết các câu hỏi chính về các khía cạnh sinh thái và sinh lý có vai trò trong sự hình thành và khả năng đề kháng của vi sinh vật trong ống tủy (chương “Sinh thái và sinh lý học của cộng đồng vi sinh vật trong ống tủy”). Hai chương cuối của phần này xem xét các cơ chế chung của sự kết dính sinh học (chương “Nguyên tắc phân tử của sự kết dính và sự hình thành sinh vật”), và các cơ chế của sự kháng thuốc đối với các mầm bệnh liên quan đến nội nha (chương “Sự kháng thuốc kháng sinh trong các cộng đồng sinh học”). Trong Phần II, sự chú ý tập trung vào bằng chứng quan sát và thực nghiệm về sinh học vi khuẩn ống tủy. Phần II bắt đầu với tổng quan về quan sát sinh học trong ống tủy bằng kính hiển vi điện tử quét (chương “Việc sử dụng kính hiển vi điện tử quét (SEM) trong hình dung sinh học ống tủy”). Bằng chứng về sự hình thành vi khuẩn sinh học trong các chế phẩm mô bệnh học và đánh giá các kỹ thuật phân tử mới để xác định vi khuẩn trong quần thể vi khuẩn sinh học trong các mẫu lâm sàng, được cung cấp trong chương “Vi khuẩn sinh học và bệnh nội nha: Khám phá mô vi khuẩn và phân tử”. Phần II khép lại với mô tả các phương pháp tiếp cận thực nghiệm phổ biến được sử dụng để nghiên cứu sinh học ống tủy bao gồm kỹ thuật mô hình sinh học trong ống nghiệm (chương “Mô hình phòng thí nghiệm của sinh học: Phát triển và đánh giá”) và xem xét những thách thức đằng sau sự phức tạp về giải phẫu trong ống tủy vì chúng có thể đóng vai trò vai trò trong việc khử trùng ống tủy (chương “Giải phẫu ống tủy: Những tác động trong việc khử trùng bằng phương pháp sinh học”). Phần cuối, Phần III, xem xét cách điều trị lâm sàng nhiễm trùng do vi khuẩn sinh học tủy răng và xem xét việc thực hiện các phương pháp tiếp cận kháng sinh mới. Đầu tiên được trình bày tổng quan về kết quả của nhiễm trùng sinh học tủy răng và các lựa chọn điều trị thích hợp (chương “Nhiễm trùng liên quan sinh học trong tủy răng: Điều trị và kết quả”). Tiếp theo là phần giải thích về sự tồn tại của các kỹ thuật tưới tiêu lâm sàng (chương “Tưới kênh gốc”) và tầm quan trọng của việc kê đơn thuốc liên tục đối với phương pháp sinh học ống tủy (chương “Điều trị giữa cuộc hẹn với Canxi Hydroxide trong các trường hợp thường quy của liệu pháp điều trị kênh gốc” ). Cuối cùng, các phương pháp và thiết bị cải tiến hướng tới việc loại bỏ các vi khuẩn sinh học khỏi ống tủy được thảo luận (chương “Các lựa chọn trị liệu nâng cao để khử trùng ống tủy”). Bộ sách này sẽ được nhiều chuyên gia liên quan đến nội nha quan tâm, bao gồm các nhà vi sinh vật học cơ bản, nhà vi sinh vật học và bác sĩ lâm sàng, và sẽ hữu ích cho các nhà khoa học đại học, sau đại học và sau tiến sĩ đang làm việc ở biên giới của sự hiểu biết mới về vai trò của vi sinh vật sinh học trong bệnh nội nha.
Springer Series on Biofilms Luis E. Chávez de Paz Christine M. Sedgley Anil Kishen Editors The Root Canal Biofilm www.pdflobby.com Springer Series on Biofilms Series Editors Mark E Shirtliff, Baltimore, USA Paul Stoodley, Southhampton, United Kingdom Thomas Bjarnsholt, Copenhagen, Denmark www.pdflobby.com More information about this series at http://www.springer.com/series/7142 www.pdflobby.com Luis E Cha´vez de Paz • Christine M Sedgley • Anil Kishen Editors The Root Canal Biofilm Volume www.pdflobby.com Editors Luis E Cha´vez de Paz The Swedish Academy for Advanced Clinical Dentistry Gothenburg Sweden Christine M Sedgley Department of Endodontology Oregon Health & Science University Portland Oregon USA Anil Kishen Department of Clinical Sciences Discipline of Endodontics University of Toronto Faculty of Dentistry Toronto Ontario Canada ISSN 1863-9607 ISSN 1863-9615 (electronic) Springer Series on Biofilms ISBN 978-3-662-47414-3 ISBN 978-3-662-47415-0 (eBook) DOI 10.1007/978-3-662-47415-0 Library of Congress Control Number: 2015953861 Springer Heidelberg New York Dordrecht London © Springer-Verlag Berlin Heidelberg 2015 This work is subject to copyright All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed The use of general descriptive names, registered names, trademarks, service marks, etc in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use The publisher, the authors and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication Neither the publisher nor the authors or the editors give a warranty, express or implied, with respect to the material contained herein or for any errors or omissions that may have been made Printed on acid-free paper Springer-Verlag GmbH (www.springer.com) Berlin Heidelberg is part of Springer www.pdflobby.com Science+Business Media Luis E Ch avez de Paz would like to dedicate his contribution to Patricia, Luis Diego, Nicol as, and Andre´s, for all their encouragement and support Anil Kishen acknowledges Arunthathi, Abinav, and Aaryan for their encouragement and patience Christine Sedgley dedicates her contribution to Victor, her greatest supporter www.pdflobby.com ThiS is a FM Blank Page www.pdflobby.com Preface Biofilms are recognized as one of the earliest ecosystems on earth They are composed of aggregates of microbial cells enclosed in a self-produced matrix adherent to a surface Root canal biofilms are complex polymicrobial structures adherent to the root canal surface that are formed by microorganisms invading the pulpal space of teeth Important histopathological studies published several decades ago first noted the presence of adherent cells on root canal surfaces However, it was not until the introduction of advanced microscopy and molecular biology techniques that they were recognized to be the dominant form of microbial life in the root canal system Similarly, it was only in the past decade that root canal infections were acknowledged to be biofilm infections Subsequently, recent studies have shown that root canal biofilms are associated with persistent endodontic infections and as such are likely to be significant contributing factors determining the outcome of endodontic treatment Concerted efforts to study root canal biofilms have been made in the past decade resulting in the publication of observational and experimental studies that detail the morphology and biology of these structures in infected root canals In addition to confirming that bacteria in root canals not exist in free-floating planktonic states as previously assumed, this new information on root canal biofilm infections has provided an opportunity to reevaluate conventional clinical protocols and improve endodontic therapeutic measures The aim of this volume is to provide a current understanding of the basic scientific aspects of root canal biofilm biology within a clinically applicable context This volume is divided into three sections Part I discusses the basic biology of root canal biofilms and addresses key questions about the ecological and physiological aspects that play a role in the formation and resistance of biofilms in root canals (chapter “Ecology and Physiology of Root Canal Microbial Biofilm Communities”) The last two chapters of this section review the general mechanisms of biofilm adhesion (chapter “Molecular Principles of Adhesion and Biofilm Formation”), and the mechanisms of antimicrobial resistance in endodontic-related pathogens (chapter “Antimicrobial Resistance in Biofilm Communities”) In Part II, vii www.pdflobby.com viii Preface attention focuses on observational and experimental evidence of root canal microbial biofilms Part II starts with an overview of observations of biofilms in root canals using scanning electron microscopy (chapter “The Use of Scanning Electron Microscopy (SEM) in Visualizing the Root Canal Biofilm”) Evidence for biofilm formation in histopathological preparations, and a review of novel molecular techniques to identify bacteria in biofilm populations in clinical samples, is provided in chapter “Bacterial Biofilms and Endodontic Disease: Histobacteriological and Molecular Exploration” Part II closes with a description of common experimental approaches utilized to study root canal biofilms including in vitro biofilm modeling techniques (chapter “Laboratory Models of Biofilms: Development and Assessment”) and examines the challenges behind anatomic complexities in root canals as these may play a role in root canal disinfection (chapter “Root Canal Anatomy: Implications in Biofilm Disinfection”) The final section, Part III, considers how infections caused by root canal biofilms are clinically treated and review the implementation of novel anti-biofilm approaches An overview of the outcome of persisting root canal biofilm infections and appropriate treatment options is first presented (chapter “Biofilm-Associated Infections in Root Canals: Treatment and Outcomes”) This is followed by an explanation of the influence of clinical irrigation techniques (chapter “Root Canal Irrigation”) and the importance of interappointment medication on root canal biofilms (chapter “Inter-appointment Medi cation with Calcium Hydroxide in Routine Cases of Root Canal Therapy”) Finally, innovative methods and devices directed towards the removal of biofilms from root canals are discussed (chapter “Advanced Therapeutic Options to Disinfect Root Canals”) This volume will be of interest to a wide range of endodontics-related professionals, including basic microbiologists, clinical microbiologists, and clinicians, and should be useful to undergraduate, postgraduate, and postdoctoral scientists working at the frontier of a new understanding of the role of microbial biofilms in endodontic disease Gothenburg, Sweden Portland, USA Toronto, Canada March 2015 Luis E Cha´vez de Paz DDS, MS, PhD Christine Sedgley MDS, MDSc, FRACDS, MRACDS(ENDO), PhD Anil Kishen www.pdflobby.com Acknowledgements It has been a fascinating journey since we started with the idea to edit a book dedicated to microbial biofilms formed in root canals First of all we wish to thank all the authors for providing their excellent contributions, without their dedication and involvement in this project it wouldn’t have been possible to complete it We wish to give a special acknowledgement to Dr William Costerton, former book series editor, for proposing that it is timely to publish a book on root canal biofilms The editors 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the antibacterial activity of zinc oxide Int J Inorg Mater 3:643–646 Yamazaki R, Goya C, Yu DG, Kimura Y, Matsumoto K (2001) Effects of erbium, chromium: YSGG laser irradiation on root canal walls: a scanning electron microscopic and thermographic study J Endod 27:9–12 Yavari HR, Rahimi S, Shahi S, Lotfi M, Barhaghi MH, Fatemi A, Abdolrahimi M (2010) Effect of Er, Cr: YSGG laser irradiation on Enterococcus faecalis in infected root canals Photomed Laser Surg 28:S91–S96 Yoon KY, Hoon Byeon J, Park JH, Hwang J (2007) Susceptibility constants of Escherichia coli and Bacillus subtilis to silver and copper nanoparticles Sci Total Environ 15:572–575 Zehnder M, S€oderling E, Salonen J, Waltimo T (2004) Preliminary evaluation of bioactive glass S53P4 as an endodontic medication in vitro J Endod 30:220–224 Zehnder M, Luder HU, Schaătzle M, Kerosuo E, Waltimo T (2006) A comparative study on the disinfection potentials of bioactive glass S53P4 and calcium hydroxide in contra-lateral human premolars ex vivo Int Endod J 39:952–958 Zhang L, Mah TF (2008) Involvement of a novel efflux system in biofilm-specific resistance to antibiotics J Bacteriol 190:4447–4452 www.pdflobby.com Index A Actinomyces, A gerencseriae, 30, 246 A israelii, 246 A naeslundii binding to cellular components, 34 biofilm development, 41–42 co-aggregation, 38 FISH technique, 17, 18 A radicidentis, 246 serotype II, and caries development, 31 Adhesins role in biofilm formation, 39–40 role in streptococcal binding, 34 Adhesion, 37–39 regulation of, 39–41 salivary pellicle, 33–34 Adsorbant surface, formation of, 25–26 Affinity matrix, 25 Aggregatibacter actinomycetemcomitans, 39 AI-2 See Autoinducer-2 (AI-2) Alkaline stress, tolerance of microorganisms to, α5β1 integrins, streptococcal, 34 Ampicillin, 67 Amylase, streptococcal binding to, 33 Antibiotics, 25 Antimicrobial photodynamic therapy (APDT) destroy mammalian cells, 334 E faecalis, 334 efflux pump, 337–338 in vitro studies, 336, 339–340 light sources, 335 methylene blue, 336 nanoparticles, 335, 336 phenothiazinium chromophore, 334 photo-oxidative effect, 334 poly-L-lysine, 334–335 potential of, 338 singlet oxygen, 333, 334 spectroscopic singlet, 333 tissue-specific factors, 336–337 tissue toxicity, 334 toluidine blue O, 334 triplet state, 333 wavelength absorption property, 333 Antimicrobial resistance biofilm phenotype, phase variation, and genetic variation, 69–70 efflux pumps, 67–68 EPS matrix, 25–26 eDNA, 60–61 exopolysaccharides, 59 extracellular proteins, 59–60 heterogeneity and oxygen gradients, 66 horizontal gene transfer, 63–65 intracellular communication and quorum sensing, 61–63 in mixed species biofilms, 70–71 penetration ability, 67 persister cells, 68–69 in root canal microflora, 71–76 stress responses, 65–66 tolerance of biofilm communities, 8–11 APDT See Antimicrobial photodynamic therapy (APDT) Apical foramen, 95 Apical pathosis, 97 Apical periodontitis, 96, 103–104, 106 classification, 115–117 © Springer-Verlag Berlin Heidelberg 2015 L.E Cha´vez de Paz et al (eds.), The Root Canal Biofilm, Springer Series on Biofilms 9, DOI 10.1007/978-3-662-47415-0 www.pdflobby.com 357 358 Index Apical periodontitis (cont.) histobacteriological analysis, 107–115 and root canal system, 87 Apical plaque, 95 Apical vapor lock, 272 Atomic force microscopy (AFM), 148–149 Autoinducer-2 (AI-2), 41–42, 63 B Bacillus B subtilis, extracellular proteins in, 660 B thuringiensis, 68 Bacterial biofilm antimicrobial resistance, 327, 328 chemical components, 328 endodontic biofilm management antibiofilm strategies, 329, 330 antimicrobial strategies, 330 APDT (see Antimicrobial photodynamic therapy (APDT)) disease, 329 enzymes, 347–348 laser-assisted (see Laser-assisted root canal disinfection) nanoparticles, 330–333 ozone, 344–346 from plants, 346–348 meticulous chemomechanical preparation, 329 nutrient and gas gradients, 328 obturation, 329 persister cells, 328 schematic diagram, 328 Bacterial endotoxin, 309 Bacteriocins, 42–43 Barrier effect, 130 Beta-lactams, 66, 71, 76 bfrC gene, 40 bfrG gene, 40 Bifidobacterium spp., and caries development, 30 Biofilm-associated infectionssss See Intraradicular microbiota Biofilms, 23–24 C Calcium hydroxide advantage, 317 apical periodontitis, 316, 317 Brown and Brenn stain, 320 clinical observations apical healing, 313 CBCT, 313 clinical outcome, 313 convincing and unequivocal support, 313–314 culture-negative samples, 310, 311 periapical healing, 313 radiographical signs, 310 randomized controlled study, 312 controlled clinical trials, 316 dentin, 314–316 historical perspective, 305–308 microbial host interaction, 319 microscopic examination, 319–320 necrotic infected pulps, 318 necrotize pulp tissue remnants, 304 non-suppurating apical lesions, 318 non-vital pulps, 317 paste-like form, 319 periapical disease, 319 pulpectomy, 317 root filling, 318 scientific evidence, 304 Actinomyces israelii, 309 anaerobic Gram-negative bacteria, 309 Enterococcus faecalis, 308–310 Gram-positive microorganisms, 310 LPS, 309–310 LTA, 309 treatment, 318 visual endorsement, 320 wet root canal, 318 Candida albicans, 24, 26, 27, 71 Carbohydrates and facultative anaerobic bacteria, and microbial homeostasis, 29 Caries development, 30–31, 36, 93 Cavitation bubbles, 279 Cellular components, binding of biofilms to, 34–35 Challisin, 41 Checkerboard DNA–DNA hybridization, 118 Chemical effect of irrigants chlorhexidine biofilm, 286–287 definition, 282–283 dentin, 288 pulp tissue, 289 EDTA biofilm, 286 definition, 282 www.pdflobby.com Index 359 dentin, 287–288 pulp tissue, 289 NaOCl biofilm, 284–285 dentin, 287, 288 hypochlorite ion, 282 hypochlorous acid, 282 pulp tissue, 288–289 uses, 281 physical properties, 283–284 second Damk€ohler number, 280 Zehnder, 280 Chlorhexidine (CHX), 25 biofilm, 286–287 definition, 282–283 dentin, 288 pulp tissue, 289 Ciprofloxacin, 68–69 CLSM See Confocal laser scanning microscopy (CLSM) Coadhesion, 129 Co-aggregation, 37–39, 130 Colonization resistance, 24 Colony-forming units (CFU), 139–140 comAB gene, 44 comCDE gene, 44 Communication See Intercellular communication Community-as-pathogen concept, 104–106 Comparative genomics, Competence-stimulating peptide (CSP), 43–44 Competence system, 43–45 Computed tomographic (CT) imaging, 157–158 comR gene, 44 Cone beam computed tomography (CBCT) C-shaped canals, 169 periapical bone loss, 313 Confocal laser scanning microscopy (CLSM), 15–16, 69, 142, 143, 145–146 Conjugated gene transfer, 63–64 Conventional biofilm models, 131 Cryogenic pulverization, 117 CSP See Competence-stimulating peptide (CSP) Culture-dependent/independent methods of mircobial composition, 30 Cytoplasmic housekeeping enzymes, response to environmental stress, D Dead-water/stagnation zone, 272 Demineralizing method, 156 Dens evaginatus, 164–165 Dens invaginatus, 164 Dental anomalies accessory canal, 172 apical canal, 173–175 C-shaped configuration Hertwig’s epithelial sheath, 166 mandibular first premolars, 166, 168 mandibular molar canals, 166–167 mandibular second molars, 166–168 Melton’s method, 167 micro-CT models, 166 preoperative diagnosis, 169 pulp floor and orifice, 168 Vertucci’s type V configuration, 168 dens evaginatus, 164–165 dens invaginatus, 164 developmental stages, 162 isthmuses configurations, 170 dentin debris, 172 lateral interconnection/transverse anastomosis, 169 mandibular and maxillary molars, 170, 171 morphology and prevalence, 170 posterior teeth, 170 lateral canals, 172–173 radix entomolaris, 165 taurodontism, 163 Dentinal tubules, 175–177 in endodontic infections, 107, 112 SEM of microorganisms/biofilm in, 92, 94 Development of biofilms attachment to tooth surfaces, 33–34 binding to cellular components, 34–35 ecological aspects communication and consequences, 41–42 competence and genetic exchange, 43–45 interspecies antagonism, 42–43 EPS production, 35–36 regulation of, 39–41 DGCs See Diguanylate cyclases (DGCs) Diaphanization, 156 Diffusion barrier, formation of, 25–26 Diguanylate cyclases (DGCs), 40 Dns evaginatus, 164–165 Dormancy, 12–14, 66 Dual-species biofilms, antimicrobial resistance in, 71 www.pdflobby.com 360 Index E Ecological disturbance of bacteria, 7–12 Ecological plaque theory, 29 Ecosystems analysis methods, 15–18 confocal scanning laser microscopy, 15–16 fluorescence in situ hybridization, 17–18 fluorescent probes, 16–17 scanning electron microscopy, 15 eDNA See Extracellular DNA (eDNA) EDTA See Ethylenediaminetetraacetic acid (EDTA) Efflux pumps, 67–68 Electron microscopy See Scanning electron microscopy (SEM) EndoActivator® system, 206 EndoActivator/Vibringe, 269 Endodontic biofilm models antibiofilm strategies, 329, 330 antimicrobial resistance, 128 antimicrobial strategies, 330 APDT (see Antimicrobial photodynamic therapy (APDT)) bacterial adherence, 128–129, 131 bacteria-substrate interaction, 129 CFU, 139–140 chemostats, 133 colorimetric techniques, 140, 141 conditioning layer, 128 constant-depth reactors, 133 disease, 329 ELISA, 147 enzymes, 347–348 extracellular matrix, 146–147 flow cell system, 132–133, 138–139 in vitro biofilm models aerobic and anaerobic environments, 131 antimicrobials and root canal irrigation strategies, 131, 134–138 biofilm bacteria and host immune cells interaction, 131 collagen-coated hydroxyapatite disk, 131, 132 conventional biofilm models, 131 glass cover, 131, 133 microbial biofilm formation, 131 planktonic killing tests, 132 static and dynamic biofilm models, 131 structure and development, 131, 139 laser-assisted (see Laser-assisted root canal disinfection) microarray analysis, 147 microbial biomass, 146–147 microscopic techniques AFM, 148–149 atomic force microscopy images, 142, 144 CLSM, 142, 143, 145–146 epifluorescence microscope, 144 ESEM, 143 fluorescence, 142 FTIR spectroscopy, 149 light microscopic image, 141 NMR spectroscopy, 149 scanning electron microscopy, 132, 133, 142, 144 transmission electron microscopy, 142 nanoparticles, 330–333 ozone, 344–346 PCR, 148 physical parameters, 146 from plants, 346–348 ultrastructure, 128–131 Endodontic infections, 104 bacterial species/phylotypes commonly detected in, 119–120 biofilm communities histobacteriological analysis of, 107 interindividual/intraindividual variability, 120 molecular analysis of, 117–120 morphology, 107, 109–110 profiling, 118–119 soft tissue lesions, 113–114 Endodontic treatment See Calcium hydroxide EndoVac®, 207 Enterobacter agglomerans, 59 Enterococcus faecalis, 17, 18 adaptation to starvation, 13 antibiotic resistance gene transfer in, 65 APDT, 334 calcium hydroxide, 308–310 calorimetric assay, 141 confocal laser scanning microscopy, 143 eDNA of, 60 failed root canal treatment, 244 horizontal gene transfer in, 64 inherent resistance to alkaline stress, 11–12, 131, 134–136, 138 www.pdflobby.com Index 361 light microscopic image, 141 Environmental scanning electron microscopy (ESEM), 89, 143 Enzyme-linked immunosorbent assay (ELISA), 147 Enzymes, role during environmental disturbance, EPS See Extracellular polymeric substances (EPS) Escherichia coli antimicrobial resistance, 56, 65 efflux pumps in, 68 persister cells, 68–69 subcellular localization of ribosomes in, 16–17 ESEM See Environmental scanning electron microscopy (ESEM) Ethylenediaminetetraacetic acid (EDTA), 286 and chlorhexidine, 289 definition, 282 microbial biofilm degradation, 247 pulp tissue, 289 vs NaOCl, 287–288 Exopolysaccharides, 59 Extracellular DNA (eDNA), 36 and antimicrobial resistance, 60–61 ultrastructural analysis, 61 Extracellular matrix (ECM), 194, 196 Extracellular polymeric substances (EPS), 15, 24, 57–58, 104–105 antimicrobial resistance, 25–26 eDNA, 60–61 exopolysaccharides, 59 extracellular proteins, 59–60 irrigants chlorhexidine, 286–287 critical loads, 278 diffusion coefficient (Drs), 284 EDTA, 286 FDG technique, 278 footprints, 277 NaOCl, 284–285 ultraasonic/laser activation, 278–279 viscoelastic properties, 277 production, 35–36 Extracellular polysaccharides, 25 Extracellular proteins, and antimicrobial resistance, 59–60 Extra-radicular microbiota, 244–246 Extraradicular microorganisms/biofilm, SEM visualization of, 95–98 F Facultative anaerobic bacteria and nutrients, recovery after treatment, FH See Fungal hyphae (FH) in root canal biofilm Fibronectin, streptococcal binding to, 34 FIB-SEM See Focused ion beam scanning electron microscopy (FIB-SEM) fimA, 35 FISH See Fluorescence in situ hybridization (FISH) Fluid dynamic gauging (FDG) technique, 278 Fluid wall interaction See Wall shear stress Fluorescein, 16 Fluorescence in situ hybridization (FISH), 17–18, 99, 117, 145 Fluorescent probes, 16–17 Fluorophores, 16 Focused ion beam scanning electron microscopy (FIB-SEM), 89 Formation of biofilms, 24–28 biofilm-specific development of genetic resistance, 26 diffusion barrier and adsorbant surface, 25–26 persister cells, 27–28 Fourier transform infrared (FTIR) spectroscopy, 149 Fungal hyphae (FH) in root canal biofilm, 94 Furcation canals, 159 Fusobacterium nucleatum, 130 G Gemella, 30 Gene expression, and survival, 10 General stress response (GSR), 13–14 Genetic exchange, 43–45 Genetic resistance, biofilm-specific development of, 26 Genetic variation, and antimicrobial resistance, 69–70 Gene transfer See Horizontal gene transfer (HGT) Gingiva, 28 Gingival crevice fluid, 28 Glucans, 35–36 Glucose, and tolerance to antimicrobials, 10–11 Glucosyltransferases (Gtfs), 35–36 www.pdflobby.com 362 Index Glycocalyx See Extracellular polymeric substances (EPS) Gram-positive bacteria, tolerance to environmental disturbances, Granulicatella, 30 Green fluorescent protein (GFP), 145 GSR See General stress response (GSR) Gtfs See Glucosyltransferases (Gtfs) Guanosine pentaphosphate (pppGpp), 13 Guanosine tetraphosphate (ppGpp), 13 Gutta-percha (GP) cone, 96, 268 H Habitat filtering, 5–7 Heterogeneity, and antimicrobial resistance, 66 HGT See Horizontal gene transfer (HGT) HMP See Human Microbiome Project (HMP) Consortium H2O2, and bacteriocin production, 43 Homeostasis, microbial, 29 Horizontal gene transfer (HGT), 63–65 Host behavior, environmental changes due to, 28 Host–biofilm interface, 24–25 Housekeeping enzymes, Human Microbiome Project (HMP) Consortium, 31–32 Hydroxyapatite, 33 Hypochlorite ion (OCl-), 282 Hypochlorous acid (HOCl), 282 I Inherent resistance, to environmental disturbances, 11–12 Insurance effects, 66 Integrins, streptococcal interaction with, 34–35 Inter-appointment medication See Calcium hydroxide Intercellular communication, 27–28, 41–42 Interspecies antagonism, 42–43 Intracellular communication, and antimicrobial resistance, 61–63 Intraradicular endodontic infections, 107, 116–117 Intraradicular microbiota bacterial occupants, 196 bacterial population, 197 bioburden, 192 biofilm eradication and wound dressing, 198 chemomechanical intervention, 198 chronic periapical lesions, 196 climax community, 193 coronal breaches, 191 coronal leakage, 193 dentine surface, 197 distribution and diversity, 191 diversity determination, 191 ECM, 194, 196 fluid-based nutritional resources, 193 fresh unreacted fluid, 198 infection continuum, 192 microbial interactions, 192 microscopy, 191 morphological complexity, 198 nutrition depletion, 197 patchy and variable distribution, 193, 194 periapical tissue status, 192 PMN phagocytosis, 194, 197 polymicrobial community, 193 root canal treatment (see Root canal treatment) slow-burning nature, 193 speculative hypotheses, 192 thick and rich biofilm layer, 193, 195 thickness of, 193, 195 vs extra-radicular infections, 196 Irrigation systems chemical effect (see Chemical effect of irrigants) conventional irrigation, 261 definition, 259–260 endodontic outcome, 289 extrusion, 273–275 flow characteristics dentinal tubules, 273 isthmuses, 273 laser-activated irrigation, 265, 271–272 lateral canals, 273 manual dynamic activation, 264, 268 negative-pressure irrigation, 264, 268 oval extensions, 273 SAI, 264, 269–271 syringe irrigation, 262, 264, 266–267 UAI, 264, 269–271 flow phase, 260, 261 fluid wall interaction (see Wall shear stress) intraradicular microbiota acoustic micro-streaming, 207 EndoVac®, 207 laser-induced agitation, 206 magnetostrictive transducers, 207 www.pdflobby.com Index 363 manual agitation, 206 mechanical shaping, 205, 206 post-mechanical flushing, 205 pressure/vacuum agitation, 206 pure flushing action, 204–205 RinsEndo, 207 sonic agitation, 206 sonic devices, 206–207 ultrasonic agitation, 206 non-instrumentation technique, 261–262 objectives of, 260 predefined effect, 261 randomized controlled trial (RCT), 260 rest phase, 260 techniques, 262, 263 Isthmuses configurations, 170 dentin debris, 172 lateral interconnection/transverse anastomosis, 169 mandibular and maxillary molars, 170, 171 morphology and prevalence, 170 posterior teeth, 170 L Lactic acid, 37 Lactobacillus, 9, 30 caries-associated species, 31 L salivarius, 14, 17, 18 preobturation cultures, 239 Laser-activated irrigation (LAI), 342, 344 Laser-assisted root canal disinfection biofilm bacteria killing, 340 chemomechanical disinfection, 344 depth of penetration, 340 Er,Cr:YSGG laser, 342 Er:YAG laser, 341 infrared lasers, 340 in vivo studies, 342, 343 LAI and PIPS, 342, 344 limitations, 341 longer pulses of, 342 Nd:YAG laser irradiation, 341, 342 optical amplificatio, 338 thermal properties, 338 tissue response, 338 treatment, 340 ultrastructure, 342 water/irrigation solution, 342 Lectins, linked to fluorophores, 16 Lipopolysaccharide (LPS), 59, 309 Lipoteichoic acid (LTA), 309 LIVE/DEAD BacLight viability probe, 16 LPS See Lipopolysaccharide (LPS) Lumen, root canal, 90 lytT gene, 36 M Macroscopic anatomy configurations, 161–162 dental anomalies (see Dental anomalies) pulp canal space, 160 pulp chamber, 158–160 Manual dynamic activation (MDA), 264, 268 MBEC See Minimal biofilm eradication concentration (MBEC) assays Mechanical instrumentation, as ecological distribution, Melton’s method, 167 Metabolic reactivation of biofilm cells, 14 Methicillin, 25 Microbiome, 29 Microflora, antimicrobial resistance in, 71–76 Microscopic anatomy, 175–177 Mineral trioxide aggregate (MTA), 315 Minimal biofilm eradication concentration (MBEC) assays, 56 Minimum inhibitory concentration (MIC), 130 Mixed species biofilms, antimicrobial resistance in, 70–71 Molecular sieving, 25 Monkey-model study, 240 Monoclonal antibodies, linked to fluorophores, 16 Morinda citrifolia (MCJ), 347 Multidrug efflux pumps (MEP), 338 Multidrug resistance efflux pumps, 67 Mutacin IV, 43 N ndvB gene, 26 Neutrophils, 5, 25 Nitric oxide (NO), role in selection of root canal bacteria, Nuclear magnetic resonance (NMR), 149 Nutrients, role in selection of root canal bacteria, 6–7 Nutrition depletion, 197 O Oral environment, challenges for biofilms in, 28–29 www.pdflobby.com 364 Index Oral mucosal epithelium, 28 Oscillatory component, 269 Oxygen gradients, and antimicrobial resistance, 66 role in selection of root canal bacteria, P Paper-point-sampling technique, 118 Passive ultrasonic irrigation (PUI), 269 PCR See Polymerase chain reaction (PCR) PDEs See Phosphodiesterases (PDEs) Penetration ability, of antimicrobials, 67 Peptides, antimicrobial, 25 Periapical lesion, 97 Persistence, definition, 14 Persister cells, 14, 27–28, 68–69, 328 Phase variation, phenotype, 69–70 Phenotype, biofilm adaptation, to environmental disturbance, and antimicrobial resistance, 69–70 Phenotypic switching, Pheromones, 64 Phosphodiesterases (PDEs), 40 Photon-initiated photoacoustic streaming (PIP), 342, 344 Photo-oxidizable amino acid residues, 334 Physiological adaptive mechanisms, 9, 11 Planktonic cells, 24 PNAG See Polysaccharide polyNacetylglucosamine (PNAG) Polymerase chain reaction (PCR), 117–118, 148 Polymicrobial synergy and dysbiosis (PSD) model, 29 Polysaccharide poly-Nacetylglucosamine (PNAG), 59 Porphyromonas gingivalis, 25, 29, 30, 41–42 binding to cellular components, 34, 35 co-aggregation, 38 genetic exchange in, 44–45 regulation of gene expression, 39 ppGpp See Guanosine tetraphosphate (ppGpp) pppGpp See Guanosine pentaphosphate (pppGpp) Primary endodontic infection, 104 Protein expression, and survival, 10 PSD See Polymicrobial synergy and dysbiosis (PSD) model Pseudomonas aeruginosa, 24, 25, 40 antibiotic-resistant phenotypic variants of, 69 antimicrobial resistance of, 56, 65 dormancy in, 66 eDNA of, 60 efflux pumps in, 67 GFP, 145 hypermutability of, 70 oxygen depletion in, 66 quorum sensing, 63 Pulp necrosis, 96–97 Pulp, status during microbial invasion, Pyrosequencing, 71, 118 Q Quantitative reverse transcriptase real-time PCR (qRT-PCR), 148 Quorum sensing, and antimicrobial resistance, 61–63 R Radix entomolaris/radix paramolaris, 165 Red complex bacterial species, 30 Resazurin, 16 Resilience, definition, Resilience of root canal microbial communities, 12–14 dormancy and adaptation to starvation, 12–14 metabolic reactivation, 14 scanning electron microscopy, 13 Resistance definition, to environmental disturbances adaptive mechanisms, 9–10 inherent resistance, 11–12 Rifampin, 26, 67 Root canal treatment antibacterial action against, 247 biofilm existence, 246 biological rationale, 199–201 chemomechanical procedures antibacterial action, 209 culture-dependent and independent approaches, 209–225 root-treated teeth, periapical disease, 225–233 direct and indirect killing effects, 246 ECM-enclosed bacteria, 247 efficacy of, 199 factors, 240–242 failed root canal treatment www.pdflobby.com Index 365 extra-radicular microbiota, 244–246 intra-radicular microbiota, 243–244 guidelines, 248–249 heal after termination, 249, 251–252 irrigation system acoustic micro-streaming, 207 EndoVac®, 207 laser-induced agitation, 206 magnetostrictive transducers, 207 manual agitation, 206 mechanical shaping, 205, 206 post-mechanical flushing, 205 pressure/vacuum agitation, 206 pure flushing action, 204–205 RinsEndo, 207 sonic agitation, 206 sonic devices, 206–207 ultrasonic agitation, 206 mechanical intraradicular preparation complex canal systems, 203, 205 coronal access cavity, 201 tapered canals, 201–202 thermoplasticised gutta-percha techniques, 202 Type A canal systems, 202 Type B canal systems, 203 Type C canal systems, 203, 204 microbial biofilm, 247 obturation, 208 obturation procedures, 225 periapical healing process, 208 periapical radiolucencies, 242, 243 preobturation culture test bacteria in, 239 chemomechanical phase, 238 effect of, 225, 234–237 focal infection, 225 monkey-model study, 240 negativevs positive culture, 238 root canal debridement, 249, 250 root canal dressing, 208 rRNA targeted oligonucleotide probing, 16 S Saliva, 28, 33–34 Salmonella enterica serovar Typhimurium, 26 Salmonella typhimurium, 309 Sanger sequencing, 118 Scanning electron microscopy (SEM), 15, 56, 88–89 of resilient root canal microbial community, 13 visualization of microorganisms/biofilm in dentinal tubules, 92, 94 extraradicular microorganisms/biofilm, 95–98 in root canal, 89–94 Secondary endodontic infection, 104 SEM See Scanning electron microscopy (SEM) Sensitivity, definition, Signaling molecules, 27–28 Slaked lim See Calcium hydroxide Soft tissue lesions, 113–114 Sonic Air Endo Handpiece, 206 Sortase A, 40 Species composition, of human oral biofilm, 29–32 microbiome era, 31–32 pre-microbiome era, 30–31 succession in biofilms, 37–39 Species-specific polymerase chain reaction, 118 16S rRNA gene amplification, 30 oligonucleotide sequences, linked to fluorophores, 16 Stable cavitation, 271 Staphylococcus S.aureus, 25, 26 antimicrobial resistance of, 56 eDNA of, 60 exopolysaccharides, 59 phase variation in, 70 S epidermidis, 24–25 antimicrobial resistance of, 71 exopolysaccharides, 59 phase variation in, 70 Starvation, adaptation to, 12–14, 27 Streptococcus, 9, 30, 32 See also Development of biofilms co-aggregation, 37–38 composition, 30 S anginosus, dormancy of, 14 S gordonii, 17, 18, 30 adhesins, 34, 40 antibiotic resistance gene transfer in, 65 and caries development, 31 co-aggregation, 38 competence development in, 44, 45 eDNA, 36–37 glucosyltransferase, 36 intercellular communication, 41 interspecies antagonism, 42–43 regulation of gene expression, 40 www.pdflobby.com 366 Index Streptococcus (cont.) S mitis, 30 S mutans antimicrobial resistance of, 71 binding to cellular components, 34 biofilm formation in, 40 in caries development, 30, 31, 36 competence development in, 45 competence system, 43 horizontal gene transfer in, 64 intercellular communication, 41 interspecies antagonism, 42–43 quorum sensing, 63 regulation of gene expression, 39–40 S oralis, 7, 30, 41–42 S salivarius, 30 S sanguinis, 30 biofilm development in, 40 and caries development, 31 competence development in, 44, 45 eDNA, 36–37 interspecies antagonism, 42–43 Stress responses and antimicrobial resistance, 65–66 Stringent response, 13 Super-resolution microscopy, 16 Swimmer cells, 68 Syringe irrigation, 262, 264, 266–267 T TA See Toxin–antitoxin (TA) systems Tannerella forsythia, 29, 30 Taurodontism, 163 TCSs See Two-component systems (TCSs) TEM See Transmission electron microscopy (TEM) Tetracycline, 71, 76 Tetrazolium salts, 16 Three-dimensional wax models, 156 TLR2, 25 TLR9, 25 Tobramycin, 59, 69 Toluidine blue O (TBO), 334 Tooth eruption, 28 Tooth surfaces, attachment of biofilms to, 33–34 Toxin–antitoxin (TA) systems, 65 Transduction, gene transfer, 63, 64 Transformation, gene transfer, 63, 64 Transmission electron microscopy (TEM), 89 Transport-based mechanisms, and antimicrobial tolerance, 11 Treponema denticola, 30 Turmeric (Curcuma longa ), 347 Two-component systems (TCSs), 39–40 V Vancomycin, 26, 71, 76 Veillonella, 30, 37 preobturation cultures, 239 V parvula, antimicrobial resistance of, 71 Viable but nonculturable (VBNC), 145 Virulence, bacterial, 106 W Wall shear stress biofilm disruption, 277–279 LAI, 265, 277 MDA, 264, 276 negative-pressure system, 264, 276 sonic and ultrasonic devices, 265, 276–277 syringe irrigation, 264, 275–276 Water, in EPS matrix, 57–58, 104 www.pdflobby.com ... microbial biofilm communities in the pathobiology of root canal infections The first of these processes occurs after the invasion of bacteria from the oral cavity into the root canal, in which the root. .. in the biofilm below The use of SEM to analyze oral biofilms and infected root canals is further reviewed in chapter ? ?The Use of Scanning Electron Microscopy (SEM) in Visualizing the Root Canal. .. and the 3D architecture of the oral biofilm Subsequently, other species are able to bind to the glucans and increase the species richness of the biofilm The role of S mutans in the oral biofilm