Biofilm is a microbial community characterized by cells attached to a surface and embedded in an extracellular matrix. Biofilms form in root canals and on materials placed in root canals. They are resistant to disinfection and prevent healing. Sodium hypochlorite, chlorhexidine, and MTAD are used to eradicate biofilms, but they often persist. Advanced techniques like lasers, photodynamic therapy, and ultrasound improve disinfection but sometimes biofilms still remain.

1. ENDODONTIC BIOFILM

2.  Biofilm is defined as a sessile multi cellular microbial
community characterized by cells that are firmly
attached to a surface and embedded in a self produced
matrix of extracellular polymeric substances
 Biofilms are formed whenever there is free flow of fluid ,
microorganisms and a solid surface. It is one of the
basic survival strategies employed by bacteria

3. Characteristics of biofilm
 Biofilms should possess
 1. autopoiesis- ability to self organize
 2. homeostasis-resist environmental
pertubations
 3.synergy- effective in association than in
isolation
 4.community- respond to environmental
changes as a unit rather than single individual

4.  Biofilm protects residing bacteria from environmental
threats
 Structure of biofilm traps nutrients
 Displays internal compartmentalization-allows bacterial
species with different growth requirements to survive
 Communicate and exchange genetic materials

5. Ultrastructure of a biofilm
Water channels help in exchange of materials between the
cells

7. Ultrastructure of a biofilm
 Basic structure of a biofilm- heterogenous arrangement
of microbial cells on a solid surface
 Glycocalyx matrix made up of extrapolymeric substance
surrounds the microcolonies and anchors the bacterial
cell to the substrate
 85% of biofilm is made up of matrix and 15% by cells
 A fully hydrated biofilm appears like a mushroom shape/
tower shape
 Water channels are primitive circulatory system in
biofilms

8. How biofilm forms
 First stage of biofilm involves the adsorption of
macromolecules in the planktonic phase to surface- a
conditioning film forms- (transport of microbe to the
substrate surface)
 Second stage – adhesion and co-adhesion of
microbes and attachment strengthened by polymer
production and unfolding of cell surface structures-
(initial non-specific microbial-substrate adherence
phase)
 Third stage involves the multiplication and metabolism
of attached microorganisms -(bacterial growth and
biofilm expansion)
 Fourth stage involves detachment of biofilm micro

9. Stages of biofilm formation
Recognition between a
suspended cell and a cell
already attached to substratum-
co-adhesion
Genetically distinct cells
recognize and clump together-

10. Factors influencing biofilm formation
 PH, temperature, surface energy of substrate, flow rate
of fluid, nutrient availability, bacterial growth stage,
surface hydrophobicity

13. Detachment of biofilm- seeding dispersal
Erosion- continous detachment of single cells
and
small portions of biofilm
Sloughing- rapid massive loss of biofilm

14. Bacterial talk in biofilms
 Communications between bacterial cells residing in a
biofilm is attained through signaling molecules by a
process called as quorum sensing
 Quorum sensing is mediated by low molecular weight
molecules- autoinducers
 Qs leads to
Exchange of genetic materials between species
Antibiotic resistance
Nutrient breakdown
Xenobiotic metabolism
Coordinated behaviour of biofilm

15. Quorum sensing

18. Endodontic biofilms
Endodontic biofilms are less diverse than oral microbiota
Root canal environment is more anaerobic
Microbes persist in isthmuses, deltas and apical parts of
root canal system- so complete disinfection is not
possible
Endodontic films are categorized in to
1.intracanal biofilm
2.extraradicular biofilm
3.periapical biofilm
4.biomaterial centered infection

19. ENDODONTIC BIOFILM FORMATION
 First, there is penetration of the organism in the pulp where it
attaches and spreads further along the root canal.
 Possibly, it is after biofilm formation that the infectious
process gains sufficient power to cause subsequent
destruction of the pulpal tissue. At some point in the
breakdown process, however, a steady state is reached
where the bacterial mass is held up by host defense
mechanisms.
 The demarcation zone may be inside the root canal near the
root canal exit, at the foramen, or, as demonstrated by
scanning electron microscopy (SEM), on the external root
surface near the exit of the foramen to the periapical tissue

20. Intracanal Biofilm
 Forms on root canal
dentin of an infected
tooth
 Identified by Nair 1987
 Cocci, rods , filaments
and spirochetes are
seen
 Morphologically distinct
type of bacteria are seen
 Eg- E faecalis
 Characteristic bacteria-
dentine wall relationship
 Distinct pattern of
organization of microbes

21. Extraradicular biofilms
 Root surface biofilms-
formed adjacent to root apex
of endodontically treated
teeth
 Found in teeth with
asymptomatic periapical
periodontitis and chronic
apical abscess with sinus
tract
 Multispecies in nature- F.
nucleatum, Po. gingivalis,
and Tannerella forsythensis
 Dominated by cocci and
short rods with cocci
attached to tooth substrate
 Calcified extraradicular
 Calcified films lead to
delayed periapical healing

22. Periapical biofilms
 Isolated biofilms in the
periapical area of
endodontically involved
teeth
 Eg- actiomycosis
 P. propionicum
The aggregation
of Actinomyces cells is
influenced by pH, ionic
strength, and cell
concentration which
facilitates biofilm formation

23. Biomaterial centered biofilm
 Bacteria adheres to
artificial biomaterial
surface and forms biofilms
 Usually reveals
opportunistic invasion by
nosocomial organisms
 Eg- coagulase negative
staphylococcus, s. aureus,
enterococci, streptococci,
p.aeruginosa
 A-Microbial film on
guttapercha
 B-E. feacalis on film
 serum plays a significant role
in biofilm formation

24. Biomaterial centered infection
 Bacterial adherence to a biomaterial surface is also
described in three phases:
 Phase 1: Transport of bacteria to biomaterial surface,
 Phase 2: Initial non-specific adhesion phase, and
 Phase 3: Specific adhesion phase.
E. faecalis, Str.
sanguinis, Streptococcusintermedius, Streptococcus
pyogenes, Staphylococcus aureus form biofilm on GP
points.
F. nucleatum, Propionibacterium acnes, Po. gingivalis,
and Pr. intermedia do not form biofilm on Gutta-
Percha(GP) points.


27. Eradication Of Biofilms
 SODIUM HYPOCHLORITE
 Effective against biofilms containing p.intermedia,
peptostreptococcus micros, s.intermedius and
fusobacterium
 Disrupts oxidative phosphorylation and inhibits DNA
synthesis of bacteria
 The antibacterial effectiveness and tissue dissolution
capacity of aqueous hypochlorite is a function of its
concentration, and so is its toxicity
 Fresh hypochlorite consistently reaches the canal system,
and concentration of the solution may thus not play a
decisive role

28.  One of the methods to improve the efficacy of sodium
hypochlorite was to use heated solution. This improves
their immediate tissue-dissolution capacity.
 Ultrasonic activation of sodium hypochlorite has also
been advocated, as this would “accelerate chemical
reactions, create cavitational effects, and achieve a
superior cleansing action”

29.  CHLORHEXIDINE DIGLUCONATE
 Effective against gram positive and gram negative
bacteria
 Denatures bacterial cell wall causes leakage of
intracellular organisms
 Substantivity effect
 Eg- e. fecalis
 IODINE
 Bactericidal, fungicidal, virucidal and sporicidal
 Attacks proteins , nucleotides and fatty acids resulting in
cell death

30. Despite its usefulness as a final irrigant, chlorhexidine cannot be
advocated as the main irrigant in standard endodontic cases,
because (a) chlorhexidine is unable to dissolve necrotic tissue
remnants, and (b) chlorhexidine is less effective on Gram-negative
than on Gram-positive bacteria

32. Eradication Of Biofilms
 EDTA
 Extracts bacterial proteins by combining with cell envelope
proteins and results in bacterial cell death
 Inhibits growth of bacteria and ultimately destroys them by
starvation
 EDTA chelates with metallic ions. Chelators may detach
biofilms adhering to root canal walls. An alternating irrigating
regimen of NaOCl and EDTA may be more efficient in
reducing bacterial loads in root canal systems than NaOCl
alone
 TETRACLEAN
 More effective than MTAD against E. faecalis
 Contains cetrimide for antimicrobial properties

34.  MTAD
 Tetracycline- Bacteriostatic broad spectrum antibiotic
 Low ph, calcium chelator
 Substantivity
 Promotes healing
 Citric acid- removes smear layer
 Detergent- decreases surface tension
 Kills most E. faecalis strains
 High binding of doxycycline- prolongs antibacterial effect

35.  CALCIUM HYDROXIDE
 Ineffective in killing E. feacalis on its own
 Effective when combined with 2% chx
 Combination completely eliminates E. faecalis
 ULTRASONIC ACTIVATED IRRIGATION
 Improves root canal cleaning and shaping- isthmus and
deltas cleaning
 OZONE
 Ozone in 0.1-0.3 ppm is able to kill bacteria after 15- 30 mins
of contact time

36. OZONE
ULTRASONIC
IRRIGATION

37.  LASERS
 Induce thermal effect producing an alteration in the bacterial
cell wall- change in the osmotic gradients and cell death
 ER- YAG irradiation reduces the number of viable cells
 Eg- A. naeslundi, E feacalis, P. acnes, F. nucleatum
 PHOTOACTIVATED DISINFECTION
 Combination of photosensitizer solution and low power laser
light
 Photosensitizer selectively accumulated in the target cell is
activated by a visible light of appropriate wave length
 ENDOACTIVATOR SYSTEM- debrides deep lateral anatomy ,
removes smear layer and dislodges simulated biofilm

38. Biofilm Detection
 The forces of interaction among bacterial cells and
between bacterial cells and substrates has been studied
by atomic force microscopy –AFM
 Micromanipulators have been used to sample individual
cells or biofilm compartments.
 Laser-based optical tweezers are noninvasive and non-
contact tools that can probe the interaction between
microscopic objects such as bacteria and collagen.
 Fourier transform infrared (FTIR) spectroscopy is used
to characterize the chemical composition of mature
biofilm structures qualitatively and quantitatively.

39.  Solid-state nuclear magnetic resonance (NMR) is a
powerful analytical tool to study the constituents of
bacterial biofilm, as well as to obtain metabolic
information in planktonic cells, adherent bacterial cells,
and in situ biofilm bacteria
 Recent advances in micromanipulator-assisted analysis,
green fluorescent protein (GFP) tagging, confocal laser
scanning microscopy (CLSM), flow cytometry, and
fluorescence in situ hybridization (FISH) have made
biofilm characterization very comprehensive.