BIOFILMS AND ENDOSCOPES

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BIOFILMS AND ENDOSCOPES
Dr. Lionel PINEAU BIOTECH-GERMANDE Marseille -
FRANCE
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BIOFILMS AND ENDOSCOPES

• GENERAL INFORMATION ON BIOFILMS
• significance, definition, characteristics, etc
• BIOFILM AND ENDOSCOPES
• WHAT CAN BE DONE ?

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BIOFILM SIGNIFICANCE
Planktonic bacteria 0.02 à 0.04
Sessile bacteria 99.96 à 99.98
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DEFINITIONJ.W. Costerton
Biofilms are defined as matrix-enclosed bacterial
populations adherent to each other and/or to
surfaces or interfaces. Biofilm consists of
single cells and microcolonies of sister cells
all embedded in a highly hydrated, predominantly
anionic matrix of bacterial exopolymers and
trapped extraneous macromolecules.
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BIOFILM FORMATION

• Bacterial colonization of surfaces can be divided
  
• into three principal stages
• the transport of the bacteria from the aqueous
  phase to the boundary layer
• the adhesion, initially in a reversible
  association and eventually in an irreversible
  adhesion.
• Colonization leading to the biofilm formation.

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STAGE 1 THE TRANSPORT OF BACTERIA FROM THE
AQUEOUS PHASE TO THE BOUNDARY LAYER
Boundary layer
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ETAPE 2 INITIAL ADHESION
Specific interactions
Electrostatic interactions
Van der Waals and electrostatic interactions
Van der Waals, electrostatic and specific
interactions
Van der Waals interactions
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ETAPE 3 COLONISATION
Microcolonies
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SUMMARY
Bacterial microcolonyy delineated by their
exopolysaccharide matrix
Network of water channel (convective flow)
Within biofilms bacterial replication and
exopolysaccharide production are regulated
(quorum sensing or complex cell-cell
communication) so that an open system of
microcolonies and water channels is produced and
maintained.
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GLYCOCALYX DEFINITION
Glycocalyx, EPS (extracellular polymeric
substance) water (99) polysaccharide-contain
ing stuctures, of bacterial origin, lying outside
the integral elements of the outer membranes of
Gram-negative cells and the peptidoglycan of
Gram-positive cells. (homopolymères simples ou
hétéropolymères complexes de glucose, mannose,
galactose, xylose, ...
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GLYCOCALYX
The polysaccharides produced vary depending on
the species but are typically made up of
repeating oligosaccharides, such as glucose,
mannose, galactose, xylose, and others.
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RESISTANCE OF BIOFILM BACTERIA
Martin EXNER  Influence of Biofilms by chemical
disinfectants and mechanical cleaning  1987.
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RESISTANCE OF BIOFILM BACTERIA
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BIOFILM AND MICROBIOLOGICAL CORROSION
Microbial corrosion induces by iron oxidizing
organisms
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CONCLUSION
 the highly structured biofilm mode of growth
provides bacteria with a primitive circulatory
system, a framework for the development of
cooperative and specialized cell functions and a
large measure of protection from antibacterial
agents. These advantages led to its functional
predominance in most natural aquatic
ecosystems compare to planktonic cells
Costerton JW
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BIOFILM AND ENDOSCOPES
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BIOFILM AND ENDOSCOPES
Somes references

• Glutaraldehydes-resistant Mycobacterium chelonae
  from endoscope washer-disinfector. Griffiths PA,
  Babb JR, Bradley CR, Fraise AP. J Appl Microbiol
  1997, Apr82(4)519-26
• Audit of bronchoscope disinfection a survey of
  procedures in England and Wales and incidents of
  mycobacterial contamination. J Hosp Infect 1994
  Apr(4)301-8
• Pseudo-outbreak of Mycobacterium chelonae and
  Methylobacterium mesophilicum caused by
  contamination of an automated endoscopy washer.
  Infect Control Hosp Epidemiol 2001
  Jul22(7)414-8

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BIOFILM AND ENDOSCOPES
Internal structure of an endoscope
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BIOFILM AND ENDOSCOPES
Internal structure of an endoscope
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BIOFILM AND ENDOSCOPES
Evaluation of the contamination level and surface
conditions of two different type of endoscope
biopsy channels (Teflon vs Goa) according to
the number of exams performed with the endoscope
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BIOFILM AND ENDOSCOPES
Internal surface of an endoscope biopsy channel
after 300 exams (Teflon)
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BIOFILM AND ENDOSCOPES
Internal surface of an endoscope biopsy channel
after 300 exams (Teflon)
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BIOFILM AND ENDOSCOPES
Internal surface of an endoscope biopsy channel
after 600 exams (Teflon)
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BIOFILM AND ENDOSCOPES
Internal surface of an endoscope biopsy channel
after 600 exams (Goa)
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BIOFILM AND ENDOSCOPES
Internal surface of an endoscope biopsy channel
after 600 exams (Goa)
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BIOFILM AND ENDOSCOPES
Internal surface of a brand new endoscope biopsy
channel (Goa)
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BIOFILM AND ENDOSCOPES
Internal surface of a brand new endoscope biopsy
channel (Teflon )
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BIOFILM
Influence of surface condition
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BIOFILM
Influence of surface condition
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CONCLUSION

•  Endoscopes are not designed to be easily cleaned
  and disinfected
• Some medical biomaterials facilitate attachment
  and colonization of microorganisms
• The use of fixative agent during the disinfection
  stage may contribute to biofilm formation

There is a real risk of biofilm formation in
endoscope channels which may constitute
uncontrolled foci but biofilm bacteria may be
difficult to recover with standard sampling
procedures
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WHAT CAN BE DONE?
 Select chemicals and/or cleaning and
disinfecting procedures which are able to remove
and/or kill biofilm bacteria.
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BIOFILM FORMATION
P1 Pump for the adhesion broth supply (2,5 to 3
ml/min).
P2 Agitation pump (100 ml / min).
B Zone d'inoculation de la boucle.
A-B Zone de prélèvement des portions de tube.
(90 cm).
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BIOFILM FORMATION
Biofilm formed experimentally on Tygon tube
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CHEMICALS EVALUATION
P
After each contact time
Tested solution
Tygon Tube biofilm
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THEORITICAL RESULTS
Log(CFU/cm2)
(proteins µg/cm2)
Log(Nb. of released bacteria/cm2)
Log(Nb. of fixed bacteria/cm2)
Proteins
Contact time (min.)
Theoretical evolutions of the number of viable
bacteria (log.) still adherent or removed from
the internal surface of the Tygon tube and of
the amount of residual proteins on the support
according to the contact time with the tested
solution.
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EXPERIMENTAL DATA
Comparaison of the activity of several detergent
solutions against Pseudomonas aeruginosa biofilm
Log (Nb. bacteria/cm2)
9,0
8,0
7,0
6,0
5,0
4,0
3,0
2,0
1,0
0,0
0,0
2,0
4,0
6,0
8,0
10,0
12,0
14,0
16,0
Contact time (min.)
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ENDOSCOPE WD
Log. CFU/ cm2
9,0
Evaluation of the efficacy of an endoscope
cleaning/disinfection cycle against biofilm
8,0
7,0
6,0
5,0
4,0
3,0
2,0
1,0
0,0
Control
Rinsing
Washing
Rinsing
Rinsing
Rinsing
Drying
Disinfection
Log CFU/ cm2 of tygon tube (E. coli biofilm).
Log CFU/ cm2 of tygon tube (S. aureusbiofilm).
Log CFU/ cm2 of tygon tube (P. aeruginosa mucoïd
biofilm).
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ENDOSCOPE WD
HTM 2030 Washer-disinfectors Validation and
verification
 The piping used to convey rinse water to the
endoscope, if contaminated, may easily develop a
layer of biofilm containing many micro-organisms
in a state which is highly resistant to chemical
disinfection   for chemical self disinfection
cycle the test required is designed to ensure
that the self disinfection cycle will disinfect
contaminated tubing by evaluating the effect of
the cycle against a biofilm containing
Pseudomonas aeruginosa 
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CONCLUSION

• Biofilms are universal, occurring in aquatic and
  industrial water systems as well as a large
  number of environments and medical devices
  relevant for public health (biomedical implant
  infection, hemodialysis, catheter associated
  infection, ).
• Effective strategies to prevent or control
  biofilms on medical devices need to be developed
  
• Medical biomaterials which minimize microbial
  cell attachment
• Chemicals and/or process which penetrate the
  biofilm and kill and/or remove the associated
  cells.
• New treatment based on inhibition of genes
  involved in cell attachment and biofilm formation
  (Donlan RM, 2002)

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CONCLUSIONThe last one!!!
Performing antimicrobial susceptibility tests
using pure cultures and in planktonic growth mode
should be questioned (Dunne WM, 2002)