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Biofilms

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Mariette Barbier

on 16 September 2014

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Transcript of Biofilms

Dental
plaque
Coronary
implants
Venous
catheter
Hip
implants
Urinary
catheter
http://biofilm.ku.dk/biofilm/
What are Biofilms?
Biofilms
are not
simply passive assemblages of cells that are stuck to surfaces

They
are
structurally and dynamically complex interactive communities
1. Attachment (reversible)
loose/transient association
weak and reversible forces (Van der Waals,...)
How?
2. Attachment (irreversible)
pilus
fimbria
flagela
...
How?

3. Production of matrix and maturation
Biofilm structure
Environment
Nutrient
availability
Genetics
Wild type
Water flow
Streamer biofilm
Banin et al. Iron and Pseudomonas aeruginosa biofilm formation.
PNAS 2005 and Hall-Stoodley et al. Bacterial Biofilms:
from the natural environment to infectious diseases.
Nature Reviews Microbiology
2004.
Mushroom-like biofilm
formed in fast moving water
filamentous
quiescent water
Flat biofilm
Mushroom-like biofilm
minimal medium with citrate
minimal medium with glucose
Klausen et al. Biofilm formation by Pseudomonas
aeruginosa wild type, flagella and type IV
pili mutants. Molecular Micro
2003
Formation of micro-colonies by clonal growth
Fimbria mutant
Pilus mutant
1 day
4 days
Klausen et al. Biofilm formation by Pseudomonas
aeruginosa wild type, flagella and type IV
pili mutants. Molecular Micro
2003
7 days
4. Detachment
Swarming/Seeding
Clumping dispersal
Surface dispersal (gliding/twitching)
Why?
How?
Exogenous factors:
(increased fluid shear,...)
Endogenous factors:
cell lysis (prophages)
enzymatic degradation
release of extracellular polymeric substances
release of surface-binding proteins
- P. aeruginosa
- S. aureus
UV- exposure
Metal
Toxicity
Acid
Exposure
Dehydration
Salinity
Phagocytosis
Predators
(amoeba and bacteria)
Biofilm
Antimicrobial
agents
1. Surfaces provide stability
2. Protection from a wide range of environmental challenges
How?
Barrier properties: neutralizes, binds or dilutes (creating a concentration gradient)
reactive antimicrobials (bleach, superoxides,...)
charged antimicrobials (metals)
large antimicrobials (immunoglobulins, surfactant proteins,...)
Physiological state
dormant/stationary phase bacteria are resistant to antibiotics affecting cell wall synthesis
Theoretically, all antibiotics require a degree of cellular activity to be effective since their function is to disrupt a bacterial process.
Subpopulations
subpopulations of resistant phenotypes known as "persisters"
mixed population of sensitive and resistant strains
Matrix
Composition = hydrated EPS (extracellular polymeric substances)
mediates adhesion to surfaces
provides mechanical stability
connects cells
eDNA (extracellular DNA)
released by prophage activity and cell lysis
structural component
source of nutrients
Intercellular Nanotubes
Intercellular Nanotubes Mediate Bacterial Communication. Dubey et al. Cell 2011
Exopolysaccharides
Alginate Psl Pel
Biosynthesis of the Pseudomonas aeruginosa extracellular polysaccharides, alginate, Pel, and Psl. Franklin et al. Frontiers in Microbiology 2011.

Main regulators of the Planktonic-to-Biofilm lifestyle switch
Secondary messenger cyclic diguanosine monophosphate
(c-di-GMP):
Secreted in early stationary growth phases, regulates the transcription factor RpoS which regulates biofilm formation
Quorum sensing:
Dependent on cell density, regulates bacterial adhesion and matrix secretion

Biofilms
M. Barbier
Formation
Role in bacterial survival
Impact
Structure
Stages of formation:
1. Attachment (reversible)
2. Attachment (irreversible)
3. Production of Matrix and Maturation
4. Detachment
This process is conserved among prokaryotes
Secretion of a viscoelastic gel. The biofilm becomes resistant to shears.
Formation of channels, death of the bacteria at the center of the biofilms
exopolysaccharide
DNA
debris, blood,...
What?
Bacterial attachment and transition from "swimmers" to "stickers"
Cellulose (EPEC),..
amyloid fibers (EPEC)
amyloid-like fibers ( )
Proteins
S. aureus
Lipids
Function:
Allow for Intercellular and Inter-species communication
( )


E. coli, B. subtilis and S. aureus
- gain new traits
- exchange nutrients (synergistic degradation of complex coumpounds,...)
- increase population fitness
- coordination (gliding motility,...)
Gradients
oxygen
nutrients
Device-related infections
Intravenous catheters
Prosthetic valves
Joint prostheses
Peritoneal dialysis catheters
Cardiac pacemakers
Cerebrospinal fluid shunts
Endotracheal tubes
Staphylococcus epidermidis
Staphylococcus aureus
Pseudomonas aerguinosa
Chronic polymer associated infections
Infective endocarditis
Inner tissue of the heart
(more frequently the valves)
Streptococcus
Infective endocarditis
Staphyloccus aureus
(acute, fulminant - days or weeks)
(subacute, more frequent, slow progression)
presence of damaged tissue
pre-existing heart condition
prostetic heart valves
vascular grafts
catheter
What are biofilms?
Gunk
Slime
Schmutz
Glue-like substance
"River slime"
"Schmutz"
"Gunk"
Dental plaque
Single or
multi-species
Can include many species of bacteria as well as fungi, algae, yeasts, protozoa, debris and corrosion products
Can be as thin as a few cells layers or many inches thick
3D resilient attached communities
Held together by "extracellular polymeric substances" or "EPS."
Health problem
Biofilms cost the U.S. billions of dollars every year in:

energy losses
equipment damage
product contamination
medical infections
Impact of biofilms
Biofilms also offer huge potential for:

bioremediating hazardous waste
biofiltering wastewater
forming biobarriers to protect soil and groundwater from contamination.
Potential
Problems
1943
1969
1978
1973
1940
1684
Zobell
Observes that the number of bacteria on surfaces is dramatically higher than in the surrounding medium (in this case, seawater).
First use of scanning and transmission electron microscopy to examine biofilms on trickling filters in a wastewater treatment plant.
They showed them to be composed of a variety of organisms and that the matrix material surrounding and enclosing cells in these biofilms was polysaccharide.
Characklis et al.
They studied microbial slimes in industrial water systems and showed that they were not only very tenacious but also highly resistant to disinfectants such as chlorine.
Costerton et al.
Explains the mechanisms by which microorganisms adhere to living and nonliving materials and the benefits accrued by this ecologic niche.
Van Leeuwenhoek
"The number of these animicules in the scurf of a man's teeth are so many that I believe they exceed the number of men in a kingdom."
Heukelekian and Heller

(bacterial growth and activity are substantially enhanced by the incorporation of a surface to which these organisms could attach)
A little bit of history
Observation of the “bottle effect” for marine microorganisms
Jones et al.
3. Metabolic advantages
Negatively charged matrix allows bacteria to concentrate cations and dissolved organic compounds to supports growth in low nutrients conditions
Metabolic collaboration (syntrophism): bacteria can rarely metabolize complex substrates (complex polysaccharides, proteins, glycoproteins,...). Various bacterial species with complementary metabolic process can collectively metabolize complex compounds
From Stewart and Franklin, 2008. Physiological heterogeneity in biofilms; Nature Review Microbiology; Vol. 6: 199-210
metabolic products
metabolic intermediates
pH
...
Sugars
Lactate
Streptococci
Actinomyces
Veillonella
Propionate
Acetate
Why do prokaryotes form biofilms ?
4. Adaptation advantages
From Stewart and Franklin, 2008. Physiological heterogeneity in biofilms; Nature Review Microbiology; Vol. 6: 199-210
a. Physiological adaptation
b. Genotypic variation and selection
c. Lateral gene transfer (more frequent in biofilms than planktonic cells)
Regulation of Biofilm formation
www.cs.montana.edu
Cell to cell signaling - Quorum sensing
Cell dependent production of auto-inducer
Regulation of gene expression
Virulence
exopolysaccharide (i.e. alginate -> prevents phagocytosis)
extracellular enzymes (hemolysins, elastases, nucleases,...)
toxins
pili
flagellae
adhesins
...
Adherence
Structure
Mobility
Dental plaque
irreversible -> cannot be removed by gentle rinsing
Where?
irregular material (diminished shear forces)
hydrophobic/non polar surfaces
Water channels
Interstitial voids

Allow the flow of water and nutrients and oxygen within the biofilm
www.colinmayfield.com
www.microbe.net
Full transcript