The document discusses biofilms in ENT diseases. It notes that biofilms are difficult to detect, highly resistant to antibiotics and host defenses, and are suspected in chronic/recurrent infections. Biofilm formation is a multistep process where bacteria attach to surfaces and produce an extracellular matrix. This makes biofilms challenging to treat, requiring both mechanical removal and long-term antibiotic therapy. The document outlines various ENT diseases associated with biofilms like chronic rhinosinusitis, otitis media, and implant infections.

1. BIOFILMS IN ENT
• Presenter – Dr. Mosin B
• Moderator – Prof. Th Sudhiranjan

2. Introduction
Biofilm formation
Pathogenic mechanisms
Biofilm and Antibiotic resistance
Detection of Biofilm
Biofilms Mediated Diseases Of ENT
Treatment
Prevention

3. INTRODUCTION
 The term ‘Biofilm’ was introduced by Bill Costerton in
1978.
 Definition – A structured community of bacterial cells
enclosed in a self – produced polymeric matrix,
adherent to an inert or living surface.
 Biofilms constitutes ~80% of the total number of
microbial infections.
 Multiple species of bacteria often co-exist with fungi.

4. Characteristics -
 * Very difficult to culture using standard
t. techniques
 * Extremely resistant to host defences
 * Highly resistant to conventional antibiotic
therapy

5. BIOFILM FORMATION
 Complex multistep process involving multiple
bacterial species.
 Formed due to a default defence mechanism to
achieve a favourable habitat, retain nutrients and to
ensure survival.
 Biofilms secrete a mixture of polysaccharides,
proteins, fatty acids and nucleic acids called as
Extracellular Polymeric Substance (EPS).

6. Importance of EPS
 80% of biofilm composition is made
of EPS
 It provides a basic platform for
surface attachment
 Act as a medium for the distribution
of nutrients and oxygen
 Protects the bacteria from host
defences and antibiotics
 Facilitate the functioning of
intercellular signalling molecules
such as c-GMP

7. STEPS OF BIOFILM FORMATION
1. Surface attachment
¡. Reversible
¡¡. Irreversible
2. Maturation of biofilm
3. Bacterial detachment/dispersion

8. 1. Surface attachment
• Planktonic cells Come in contact
with a conditioning film(surface)
Adhere via physical forces or by bacterial
appendages such as pili or flagella.
• This stage is reversible due to weak interaction
between the bacteria and surface.
• When the attractive forces are greater than
repulsion, bacterial cells become irreversibly
attached.

9. • Cell to cell connections also occur, known as
cohesion.
• Formation of microcolonies.
• Bacteria starts secreting EPS to fix the
aggregation of a biofilm.
2. Maturation of biofilm
• The adhered cells grow and mature by
interacting among themselves.
• More EPS production, thus stabilising the biofilm
network.
• Microcolonies mature into macrocolonies which
get encased within the EPS where intercellular
signalling or Quorum sensing takeplace.

10. Quorum sensing
• A phenomenon which involves cell density - dependent
control of gene expression.
• It is the ability of bacterial cells to communicate each
other for transfer of genetic material.
• Communication is mediated by small signal molecules
called auto-inducers.
• Importance - Disruption in QS system can inhibit the
growth of bacteria within the EPS.
Disassembly of pre-established biofilms
Increase biofilm susceptibility to antibiotics
to

11. 3. Bacterial detachment/ dispersion
Maturation of biofilm
Resources become limited and
accumulation of toxic products occurs
 In order to expand, get nutrition, and to
eliminate stress-inducing conditions the,
cells disperse to other surfaces.
 Bacterial cells within the biofilm produce
enzymes, which breaks the
polysaccharide in EPS , thereby releasing
the surface bacteria.
 As single cells or in clumps

12.  Important in the progression of biofilm, thereby
spreading infection .
 Causes c/c infection and embolic complications.
 Often referred to as ‘metastatic seeding’.

13. PATHOGENIC MECHANISMS
 Increasing the metabolic efficiency by ‘division of
labour’
 Performing phagocytosis for evading host defences
 Obtaining a high-density population of microorganisms
 Generation of high virulent strains by gene transfer
 Protection against anti-microbial agents
 Transmission of microorganisms to different sites by
detachment of microbial colonies

14. BIOFILMS AND ANTIBIOTIC RESISTANCE
1. Altered micro environment
2. Delayed or failed antibiotic penetrance
3. Persister cells
4. Reduced growth rate
5. Horizontal gene transfer
6. Anaerobic metabolism
7. Extracellular DNA(eDNA)
8. Efflux pump mechanism
9. Interaction b/w bacteria and fungi

15. DETECTION OF BIOFILM
 Very difficult to identify
 Suspect in cases of :
¡. Chronic infections
¡¡. Recurrent/recalcitrant infections
¡¡¡. Implant related infections
¡v. Infections which are resistant to
culture directed antibiotics

16. 1. Routine Microbiological Examinations
* Light microscopy Multiple samples often from
as. different sites may be
requir. required
* C & S High chances of false -ve
Limited magnificatn & resolutn
2. New Techniques of Microbiology
(a) Scanning Electron Microscopy
* Higher resolutn & magnificatn
* Study of biofilm spatial structure
* Lengthy and complex procedure
* Difficult to differentiate b/w
mucus, clot and biofilm.

17. (b) Transmission Electron Microscopy
* Similar to SEM.
* Gives only 2-D images of biofilm
(c) Confocal Laser Scanning Microscopy(CLSM)
* 3-D image
* Qualitative and quantitative
* Mixed-species biofilm
* The effect of antibiotics on
bacterial cells can be studied.

18. (d) Fluorescent in situ hybridization(FISH)
* Specific microorganisms in a
heterogenous biofilm can be identified.
* The main drawback is the need to
presumptively identify the organism
which is then probed.
Fluorescence staining coupled with
CLSM is an ideal method to identify
biofilms.

19. (e) Molecular methods
* PCR Techniques
* DNA– DNA hybridization technique
* Microarray technology
🌀 Detect specific biofilm genes.
🌀 They are more specific, sensitive and rapid
detection methods.
(f) Infrared spectroscopy
(g) Biofilm assay

20. BIOFILM MEDIATED DISEASES OF ENT
 Chronic Rhinosinusitis
 Otitis media with effusion
 Chronic tonsillitis and adenoiditis
 Chronic otitis media
 Cholesteatoma
 Oral Cavity
 Implants and Prostheses

21. 1. Chronic Rhinosinusitis
 Biofilms in 44– 92% of CRS
 S. aureus (M/C), Pseudomonas,
H. influenza, S. pneumoniae
 Fungal (50%) – C. albicans
 S. aureus biofilms – releases superantigenic molecules
 Defective ciliary function and defects in the adaptive
and innate immunity initiates biofilm formation.
 Mucus stasis within the sinus cavity, predisposes to
further biofilm progression.

22.  In this way it becomes a key modulator of the
refractory nature of CRS.

23. • Associated with worst postoperative outcomes after
FESS.
• 75% of patients undergoing revision surgery showed
to have biofilm.
• Presence of biofilms arises the need for more
extensive surgical interventions.
• There will be persistent sinonasal inflammation,
recurrent a/c exacerbations, despite long term
culture-directed antibiotic therapy and well-
performed sinus surgery.
• Also associated with osteitis – biofilms causes bony
inflammation by release of eosinophilic
inflammatory mediators.

24. 2. Otitis media with effusion
 In > 90% cases of recurrent OME
 Common source of biofilms is adenoid tissue
 Endotoxins chronicity of OME
 Pseudomonas, S. pneumoniae

25. 3. Adenoids and Tonsils
• Adenoidectomy is considered beneficial in children
with CRS and OME
• 95% specimens of surgically removed adenoids in
children with CRS showed presence of biofilm.
• S. aureus, Pseudomonas, S. pneumoniae, H.
influenza
• 70% tonsil specimens removed from patients with
c/c or recurrent tonsillitis, contained biofilms.

26. 4. Chronic otitis media
 In 75% of squamosal and 35% of mucosal types –
explains its recurrent and recalcitrant course.
 S. aureus, Pseudomonas, S. pneumoniae
 May found in damaged tissue, such as ulcerated
middle ear mucosa or exposed osteitic bone .
 Presence of biofilms is high in middle ear mucosa
compared with the mastoid and ossicles.
 Resistant bacterial colonies in mastoid cavity can
influence graft success after surgery.
 Residual biofilms is one of the reason for failure and
persistent ear discharge after mastoidectomy.

27. 5. Cholesteatoma
 In 80 – 90% cases
 Keratin matrix gives an ideal environment for the
formation of biofilms
 M/C Organism – Pseudomonas
Have the ability to adhere to keratinocytes
 Chronic inflammation, which is caused by recurrent
infections, is one of the factors contributing to the
pathogenesis of cholesteatoma.
 Induce mucin hypersecretion, hyperproliferation of
epithelial cells and keratinocytes, bone resorption by
activation of collagenases and osteoclasts.

28.  If autologous ossicle chain reconstruction is attempted,
inactivation of biofilm should be done.
6. Oral cavity
 Dental plaque
 Gingivitis
 Periodontitis

29. 7. Implant and Prostheses
1. Cochlear implants
2. Tracheostomy tubes
3. Tympanostomy tubes
4. Speech Prostheses
1. Cochlear implants
 Very rarely, the internal component can become
infected.
 Mostly, implants to be removed and replaced.
 Very few cases – successfully managed with local
therapies.

30. • Irregular surfaces of the cochlear
implants serve as a platform for
biofilm formation.
• Preventive measures :-
* Use of coated implants
* Proper positioning of CI on smooth skull
surface with no pressure on the scalp.
* Tight periosteal flap closure to
compress cochlear implant.
* Use of skin glue and sterile strip to avoid
stitch abscess.
* H2O2 dressing to destroy biofilms.

31. 2. Tracheostomy tubes
 90% of tracheostomy tubes removed after 7 days
show biofilms
 Pseudomonas, S. epidermidis
3. Tympanostomy tubes
 Can act as a surface for biofilm formation.
 Methods to prevent :
* Minimizing bleeding during surgery. ( Clots
provide an apt environment for Pseudomonas)
* Use of Ionized, coated fluoroplastic grommets .
* Ion-bombarded silicon tubes and coating with
albumin.

32. 4. Speech Prostheses
 S. aureus, S. epidermidis, C. albicans
 Measures to prevent –
* Topical use of N-acetyl cystine / 7% Silver oxide
or use of a prosthesis made of Silicon
modified with Per-fluro-alkyl-siloxane.

33. TREATMENT
 Difficult to completely eradicate biofilms.
 Common strategies :-
1. Mechanical Removal
2. Early and aggressive antibiotic treatments
3. Dispersion of the existing biofilm
4. Disruption of Quorum sensing

34. 1. Mechanical Removal
 Surgery :-
* Mechanically disrupts biofilms
* Assists the host’s natural defences to clear the
infections
* Increases O2 tension
 Debridement
 Removal of infected implants

35. 2. Early and aggressive antibiotic treatments
 Biofilm bacteria are 10 - 1000 times more resistant
 Requires sensitive and well penetrating antibiotics
 Systemic + Topical More
 Combination of antibiotics with diff MOA beneficial
oSystemic Antibiotics :-
 Macrolides, Fluoroquinolones
( Active against non-growing bacteria’s)
 Long term therapy will be required
 Macrolides - * Antibacterial & immunomodulatory effect
* Enhance phagocytic properties of
neutrophils against biofilms
* Inhibits inter-cellular signalling

36. o Topical medications
 Mupirocin irrigation – S. aureus
 Tobramycin irrigation – Pseudomonas Shown to
 Colloidal Silver nasal sprays have
 Sodium hypochlorite irrigation beneficial
 Xylitol (5-Carbon Alcohol) effects in
 Hyaluronic acid nebulization CRS
 Non-antibiotic microbial agents –
* N,N-dichloro-2-2-dimethyl taurine
* Manuka honey
* Gentian violet

37. Manuka honey
 Active agent – Methylglyoxal
 Most lethal for biofilms
 Active against a broad spectrum of gram-
positive and gram-negative bacteria
 1.8mg/ml
• Biofilms under the effect of electric currents,
ultrasonic radiation, pressure waves have found
to be more susceptible to antibiotics.
(Increases the penetration of antibiotics)

38. • Correction of acid-base balance :-
Inflammation metabolism acidosis
pH Effect of antibiotics
• Role of Probiotics :-
* Eradication of pathogenic organisms without
disturbing useful micro environment.
* Also releases antimicrobial agents.
* Consumption of probiotic drinks, containing
Lactobacillus, 3 times daily for 6 months found to
reduce the need for voice prosthesis replacement.

39. • Antimicrobial photodynamic therapy :-
* Under trial
* Involves the act of destroying cells in the
presence of a photo reactive dye and a laser.
• Role of Steroids :-
* Not well established
* Higher concentrations of fluticasone,
budesonide and mometasone shown to have
some beneficial effects in CRS.
• Nanoparticles with intrinsic antimicrobial activity :-
* eg: nanoparticles coated with Silver
* Increased penetration into biofilms.
• Bacteriophage therapy.

40. 3. Dispersion of the existing biofilm
 Surfactants were used as dispersion agents
 They dissolves EPS.
 1% baby shampoo – o causes nasal irritation
o ed mucociliary clearance time
 Citric acid zwitterionic surfactant – ciliary toxic
 SinuSurf – causes anosmia
Matrix degrading enzymes
* DNase I - degrade eDNA
* Dispersin B - degrade matrix
* a-amylase – degrade polysaccharides

41. 4. Disruption of Quorum sensing
 Macrolides – Azithromycin
 4-Phenyl butanoic acid, extracted from Bacillus
pumilis
 Cinnamaldehyde and Baicalin hydrate - decreases
resistance of Pseudomonas and B. cepacia towards
tobramycin.
 Halogenated furanone.

42. PREVENTION
 Extreme aseptic precaution during implant
procedures
 Ultraclean operating theatres
 Sterilisation of surgical garments, instruments and
implants
 Use of prophylactic antibiotics and antibiotic coated
implants.

43. SUMMARY
• Biofilms are highly resistant to antibiotics and host
defences
• Difficult to detect in a normal clinical setting
• Highly suspect in cases of chronic/recurrent/
recalcitrant/implant related infections.
• Mechanical removal of biofilm followed by medical
therapy remains the cornerstone of management.
• Difficult to completely eradicate.

44. REFERENCES
• Scott Brown’s Otorhinolaryngology Head & Neck Surgery – 8th edition
• Cummings Otolaryngology Head & Neck Surgery – 6th edition
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differentially structured metallic surfaces using confocal laser scanning
microscopy. Eng Life SCI. 2019;19:513-521.
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Arch Otolaryngol Head Neck Surg. 2002;128:1129-1133.
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after endoscopic sinus surgery. Am J Rhinol Allergy. 2010;24:169-174.
• Abu Bakar MB, McKimm J, Haque M. Otitis media and biofilm: An overview.
Int J Nutr Pharmacol Neurol Dis. 2018;8:70-8.

45. • Hong W et al. Strategies for combating bacterial biofilm infections. Int J
Oral SCI. 2015 Mar;7(1):1-7.
• Zohra K et al. Bacterial biofilm formation on implantable devices and
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46. THANK YOU