Fundamentals and Applications of Biofilms Bacterial Biofilm...
Transcript of Fundamentals and Applications of Biofilms Bacterial Biofilm...
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Fundamentals and Applications of BiofilmsBacterial Biofilm Control: Cell-to-Cell CommunicationChing-Tsan Huang (黃慶璨)Office: Agronomy Building, Room 111Tel: (02) 33664454E-mail: [email protected]
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The Discovery of Cell-Cell CommunicationStage 1 (1970s): Discovery and ecological explanation
Quorum sensingLuminous bacteria (e.g. Vibrio fischeri ) contain high levels of luciferase only when cultures reach the late-log phase of growth.
AutoinductionMicroorganisms producing pheromones and communicating with one another, was proposed.
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The Discovery of Cell-Cell CommunicationStage 2 (1980s): Proof of the Autoinduction
Physiological, chemical and genetic evidences were foundThe structure of the autoinducer singal, N-3-(oxohexanoyl) homoserine lactone was solved. Genes required to enable autoinduction of luminescence were identified.
luxR and luxI model was provedluxR: encodes an autoinducer-responsive transcriptional activator. luxI: encodes a protein required for autoinducer synthesis.
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Stage 3 (1990s): Discovery of luxR-luxI-type systems in other bacteria.
The Discovery of Cell-Cell Communication
Bacterium Phenotype Molecule
Photobacterium fischeri Bioluminescence OHHL
Vibrio harveyi Bioluminescence HBHL
Erwinia carotovoraMultiple exoenzymesvirulence, antibiotic
OHHL
Pseudomonas aeruginosaMultiple exoenzymes, virulence OdDHLVirulence, secondary metabolites BHL & HHL
Agrobacterium tumefaciens Ti plasmid conjugal transfer OOHL
Streptomyces spp. Antibiotic, differentiation -butyrolactones
& more
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Quorum Sensing Signal MoleculesAcylated homoserine lactones
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Mechanism of Autoinduction
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More detailed
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I Gene and R Gene LuxI-type proteins recruit substrates from general
metabolismAt least three LuxI-type proteins (LuxI, RhlI, TraI), the substrates are S-adenosylmethionine (AdoMet) and the appropriate fatty acid substrate conjugated to acyl carrier protein (ACP). (more)
Substrate interactions of acyl-HSL synthetasesThe active site of an acyl-HSL synthetase must accommodate AdoMet and acyl chain carried by ACP.
Regulation of acyl-HSL synthetaseRegulation of enzyme activity Control of acyl-HSL synthetase
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Pseudomonas aeruginosa lasI/rhlI Regulation
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Microbiology. 2008 Aug;154(Pt 8):2184-94.
PMN: polymorphonuclearleukocyte
Quorum Sensing in Acute or Chronic Infection
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Cell-to-Cell Communication & Biofilm Formation
Biofilms are perfect situation for cell-to-cell communicationHigh cell density may be in the region of 1010 cell per cm3
Increased physical proximity
Evidence of cell-to-cell communication within biofilms
Davies et al. (1998) Science 280: 295-298.
& more
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Bio
film
den
sity
log(
cfu/
cm2 )
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Time (h)0 12 24 36 48 60 72
Bio
film
den
sity
log
(cel
ls/c
m2 )
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A
B
WT
QS mutants
Time(h)24 36 48 60 72
g p
olys
acch
arid
e /
g pr
otei
n
0.0
0.1
0.2
0.3
0.4
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0.6 PAO1 PDO100 JP1 JP2
* * *WT
QS mutants
Shih and Huang (2002) J. Antimicrobiol. Chemother. 49: 309-315.
Biofilm Formation: Wild Type vs Mutants
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WT
rhlI
lasI
Cell/EPS Distribution: Wild Type vs QS MutantsEtBr Calcofluor white
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Cell-Cell Communication & Biofilm Resistance
Little was known
Cell-to-cell communication
Biofilm formation
Polysaccharide production
Biofilm resistance
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0.0
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1.0
Time (min)0 30 60 90 120
Sur
vivi
ng fr
actio
n
0.0
0.2
0.4
0.6
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0 30 60 90 120
PAO1 JP1
PDO100 JP2
Shih and Huang (2002) J. Antimicrobiol. Chemother. 49: 309-315.
Biofilm Resistance : Wild type vs Mutants
& others
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Appl Microbiol Biotechnol. 86(5):1267-79, 2010.
Bacterial QS systems
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Quorum Sensing in Gram Positives
Two-component signal transduction
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AIP Structure-Function
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Three canonical quorum-sensing circuits in bacteria
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Interspecies Quorum Sensing by AI-2
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Vibrio harveyi AI-1 & AI-2 Quorum Sensing
Auto-phosphorylate De-
phosphoralate
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Cell Microbiol. 11(3):363-9, 2009.
Three autoinducing strategies in Gram-negatives.
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Trends Microbiol. 18(2):73-80, 2010.
Signaling interactions between different groups of bacteria
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Biofilm ControlCell-to-Cell Communication approach
Center for Biofilm Engineering, MSU-Bozeman
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Potential targets and intervention strategies disrupting quorum-sensing (QS) in bacteria
FRAY R G Ann Bot 2002;89:245-253
AHL
AHL analog
Cyclic dipeptides
activate or repress AHL receptor
Scanning electron micrographs of biofilms
Beighton D ADR 2009;21:69-73
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Quorum Sensing
Engineered QS Systems
Biosensors
Biofouling
Production of biochemicals
Tissue Engineering
Mixed species fermentation
Pathogen diagnostics
Cancer detection
Pathogen therapeutics
Biocontrol
Surgical implants
Membrane bioreactors
Heat exchangers, tanks, pipes
Quorum Sensing on Biotechnology
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Appl Microbiol Biotechnol. 86(5):1267-79, 2010.
Engineered QS systems