CC BY-NC-ND 4.0 · Eur J Dent 2012; 06(01): 043-050
DOI: 10.1055/s-0039-1698929
Original Article
Dental Investigation Society

Development of an intracanal mature Enterococcus faecalis biofilm and its susceptibility to some antimicrobial intracanal medications; an in vitro study

Shehab El-Din Mohamed Saber
1   Department of Endodontic, Faculty of Dentistry, Ain Shams University, Cairo
,
Soha A El-Hady
2   Microbiology and Immunology Department, Ain Shams University, Cairo
› Author Affiliations
Further Information

Publication History

Publication Date:
30 September 2019 (online)

ABSTRACT

Objectives: To develop a mature biofilm of Enterococcus faecalis inside the root canal system and to test its susceptibility to some antimicrobial medications in vitro.

Methods: Single rooted premolars were mechanically enlarged, sterilized, and then infected with a clinical isolate of E. faecalis. Biofilm formation and maturation was monitored using SEM. Biofilm bacteria were exposed to Amoxicillin+clavulanate, Ciprofloxacin, Clindamycin, Doxycycline, and calcium hydroxide as intracanal medications for 1 week. Finally bacterial samples were collected, and colony-forming units were enumerated.

Results: SEM examination confirmed the formation of a mature biofilm at the end of the incubation period. All the chemotherapeutic agents used were significantly better than Calcium hydroxide in elimination of biofilm bacteria. The antimicrobial effect of Amoxicillin + clavulanate, Ciprofloxacin and Clindamycin was significantly better than Doxycycline (P=.05). However the difference in the antimicrobial effectiveness among them was statistically non-significant (P=.05).

Conclusions: The method used for bacterial biofilm development and maturation is reliable and can be used to assess the anti bacterial potential of endodontic materials. Also, the local application of antibacterial agents can be beneficial in resistant cases of apical periodontitis but only after careful culture and sensitivity testing to choose the appropriate agent for the existing flora. (Eur J Dent 2012;6:43-50)

 
  • REFERENCES

  • 1 Wahlin YB, Holm AK. Changes in the oral microflora in patients with acute leukemia and related disorders during the period of induction therapy. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1988;65:411-417.
  • 2 Rams TE, Feik D, Young V, Hammond BF, Slots J. Enterococci in human periodontitis. Oral Microbiol Immunol 1992;7:249-252.
  • 3 Hancock HH 3rd, Sigurdsson A, Trope M, Moiseiwitsch J. Bacteria isolated after unsuccessful endodontic treatment in a North American population. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2001;91:579-586.
  • 4 Pinheiro ET, Gomes BP, Ferraz CC, Teixeira FB, Zaia AA, Souza Filho FJ. Evaluation of root canal microorganisms isolated form teeth with endodontic failure and their antimicrobial susceptibility. Oral Microbiol Immunol 2003;18:100-103.
  • 5 Siqueira JF, Rôças IN. Polymerase chain reaction-based analysis of microorganisms associated with failed endodontic treatment. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2004;97:85-94.
  • 6 Huycke MM, Sahm DF, Gilmore MS. Multiple-drug resistant enterococci: the nature of the problem and an agenda for the future. Emerg Infect Dis 1998;4:239-249.
  • 7 Hancock HH, Jr, Sigurdsson A, Trope M, Moiseiwitsch J. Bacteria isolated after unsuccessful endodontic treatment in a North American population. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2001;91:579-586.
  • 8 Sundqvist G, Figdor D, Persson S, Sjögren U. Microbiologic analysis of teeth with failed endodontic treatment and the outcome of conservative re-treatment. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1998;85:86-93.
  • 9 Sedgley CM, Lennan SL, Appelbe OK. Survival of Enterococcus faecalis in root canals ex vivo. Int Endod J 2005;38:735-742.
  • 10 Portenier I, Waltimo TMT, Haapasalo M. Enterococcus faecalis – the root canal survivor and ‘star’ in post-treatment disease. Endod Topics 2003;6:135-159.
  • 11 Sedgley CM, Molander A, Flannagan SE. Virulence, phenotype and genotype characteristics of endodontic Enterococcus spp. Oral Microbiol Immunol 2005;20:10-19.
  • 12 Svensater G, Bergenholtz G. Biofilms in endodontic infections. Endod Topics 2004;9:27-36
  • 13 Jett BD, Huycke MM, Gilmore MS. Virulence of enterococci. Clin Microbiol Rev 1994;7:462-478.
  • 14 Elsner HA, Sobottka I, Mack D. Virulence factors of Enterococcus faecalis and Enterococcus faecium blood culture isolates. Eur J Clin Microbiol Infect Dis 2000;19:39-42.
  • 15 Haapasalo M, Orstavik D. In Vitro infection and disinfection of dentinal tubules. J Dent Res 1987;31:1-7
  • 16 Ten Cate JM. Biofilms, a new approach to the microbiology of dental plaque. Odontology 2006;94:1-9.
  • 17 Chávez de Paz L. Redefining the Persistent Infection in Root Canals: Possible Role of Biofilm Communities. J Endod 2007;33:652-662.
  • 18 Mohammadi Z, Abbott PV. On the local applications of antibiotics and antibiotic-based agents in endodontics and dental traumatology. Int Endod J 2009;42:555-567.
  • 19 Stashenko P, Teles R, D’Souza R. Periapical inflammatory responses and their modulation. Crit Rev Oral Biol Med 1998;9:498-521.
  • 20 Abbott PV, Hume WR, Pearman JW. Antibiotics and endodontics. Aust Dent J 1990;35:50-60.
  • 21 Bystrom A, Claesson R, Sundqvist G. The antibacterial effect of campho-rated paramonochlorphenol, camphorated phenol and calcium hydroxide in the treatment of infected root canals. Endod Dent Traumatol 1985;1:170-175.
  • 22 Gilad JZ, Teles R, Goodson M, White RR, Stashenko P. Development of a clindamycin-impregnated fibre as an intracanal medication in endodontic therapy. J Endod 1999;25:722-727.
  • 23 Portenier I, Haapasalo H, Rye A, Waltimo T, Ørstavik D, Haapasalo M Inactivation of root canal medicaments by dentine, hydroxylapatite and bovine serum albumin. Int Endod J 2001;34:184-188.
  • 24 Martohi YA, Agnihorti H, Dubey D. Enteroccocal resistancean overview. Indian J Med Microbiol 2005;23:214-219.
  • 25 Greenwood D, Slack R, Peutherer J. Antimicrobial agents. In: Medical Microbiology a guide to microbial infection; pathogenesis, immunity, Lab. Diagnosis and control. London: Churchill Livingstone; 2007 pp.52-67.
  • 26 Abdullah M, Ng YL, Gulabivala K, Moles DR, Spratt DA. Susceptibilities of two Enterococcus faecalis phenotypes to root canal medications. J Endod 2005;31:30-36.
  • 27 Esterla C, Sydney GB, Figueiredo JP, Esterla ER. A model system to study antimicrobial stratigies in endodontic biofilms. J Appl Oral Sci 2009;17:87-91.
  • 28 Sen BH, Piskin B, Demirci T. Observation of bacteria and fungi in infected root canals and dentinal tubules by SEM. Endod Dent Traumatol 1995;11:6-9.
  • 29 Andrews JM. Determination of minimum inhibitory concentrations. J Antimicrob Chemother 2001;48:5-16.
  • 30 Edgar S, Marshall JG, Baumgatner JC. The Antimicrobial Effect of Chloroform on Enterococcus faecalis After Gutta- Percha Removal. J Endod 2006;32:1185-1187.
  • 31 Chavez de Paz LE, Dahlen G, Molander A, Moller ,Bergenholtz G. Bacteria recovered from teeth with apical periodontitis after antimicrobial endodontic treatment. Int Endod J 2003;36:500-508.
  • 32 Johnson SA, Goddard PA, Iliffe C, Timmins B, Rickard AH, Robson G, Handley PS. Comparative susceptibility of resident and transient hand bacteria to para-chloro-meta-xylenol and triclosan. J Appl Microbiol 2002;93:336-344.
  • 33 Dunavant TR, Regan JD, Glickman GN, Solomon ES, Honeyman AL. Comparative evaluation of endodontic irrigants against Enterococcus faecalis biofilms. J Endod 2006;32:527-531.
  • 34 Spratt DA, Pratten J, Wilson M, Gulabivala K. An in vitro evaluation of the antimicrobial efficacy of irrigants on biofilms of root canal isolates. Int Endod J 2001;34:300-307.
  • 35 George S, Kishen A, Song KP. The role of environmental changes on monospecies biofilm formation on root canal wall by Enterococcus faecalis. J Endod 2005;31:867-872.
  • 36 Kishen A, George S, Kumar R. Enterococcus faecalis-mediated biomineralized biofilm formation on root canal dentine in vitro. J Biomed Mater Res A 2006;77:406-415.
  • 37 Duggan JM, Sedgley CM. Biofilm Formation of Oral and Endodontic Enterococcus faecalis. J Endod 2007;33:815-818.
  • 38 Murray BE. Beta-Lactamase-producing enterococci. Antimicrob Agents Chemother 1992;36:2355-2359.
  • 39 Windley W 3rd, Teixeira F, Levin L, Sigurdsson A, Trope M. Disinfection of Immature Teeth with a Triple Antibiotic Paste. J Endod 2005;31:439-443.
  • 40 Torabinejad M, Shabahang S, Aprecio R, Kettering JD. The antimicrobial effect of MTAD: an in vitro investigation. J Endod 2003;29:400-403.
  • 41 Chan Y, Chan CH. Antibiotic resistance of pathogenic bacteria from odontogenic infections in Taiwan. J Microbiol Immunol Infect 2003;36:105-110.
  • 42 Sjögren U, Figdor D, Spångberg L, Sundqvist G. The antimicrobial effect of calcium hydroxide as a short-term intracanal dressing. Int Endod J 1991;24:119-125.
  • 43 Esberard RM, Carnes DL Jr, del Rio CE. Changes in pH at the dentin surface in roots obturated with calcium hydroxide pastes. J Endod 1996;22:402-405.
  • 44 Nerwich A, Figdor D, Messer HH. pH changes in root dentin over a 4-week period following root canal dressing with calcium hydroxide. J Endod 1993;19:302-306.
  • 45 Voel D, Voel JG, Pratt CW. Fundamentals of biochemistry. 1st ed. Hoboken, NJ: John Wiley & Sons, 1999:93-123.
  • 46 Khan AA, Sun X, Hargreaves KM. Effect of Calcium Hydroxide on Proinflammatory Cytokines and Neuropeptides. J Endod 2008;34:1360-1363.
  • 47 Evans M, Davies JK, Sundqvist G, Figdor D. Mechanisms involved in the resistance of Enterococcus faecalis to calcium hydroxide. Int Endod J 2002;35:221-228.
  • 48 Cogulu D, Uzel A, Oncag O, Aksoy S, Eronat C. Detection of Enterococcus faecalis in necrotic teeth root canals by culture and polymerase chain reaction Methods. Eur J Dent 2007;1:216-221.
  • 49 Staehle HJ, Pioch T, Hoppe W. The alkalizing properties of calcium hydroxide compounds. Endod Dent Traumatol 1989;5:147-145.
  • 50 Siren EK, Haapasalo MP, Ranta K, Salmi P, Kerosuo EN Microbiological findings and clinical treatment procedures in endodontic cases selected for microbiological investigation. Int Endod J 1997;30:91-95.
  • 51 George S, Kishen A, Song KP. The role of environmental changes on monospecies biofilm formation on root canal wall by Enterococcus faecalis. J Endod 2005;31:867-872.
  • 52 Orstavik D, Haapasalo M. Disinfection by endodontic irrigants and dressings of experimentally infected dentinal tubules. Endod Dent Traumatol 1990;6:142-149.
  • 53 Heling I, Steinberg D, Kenig S, et al. Efficacy of a sustainedrelease device containing chlorhexidine and Ca(OH)2 in preventing secondary infection of dentinal tubules. Int Endod J 1992;25:20-24.
  • 54 Baik JE, Kum KY, Yun CH, Lee JK, Lee K, Kim KK, Han SH. Calcium hydroxide inactivates lipoteichoic acid from Enterococcus faecalis. J Endod 2008;34:1355-1359.
  • 55 Brandle N, Zehnder M, Weiger R, Waltimo T. Impact of growth conditions on susceptibility of five microbial species to alkaline stress. J Endod 2008;34:579-582.