Virulence Gene Regulation in Bordetella and Biofilm Development

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Virulence Gene Regulation in Bordetella and
Biofilm Development
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Bordetella

• Bordetella pertussis
• Whooping cough in humans
• Bordetella bronchiseptica
• Has a much broad host range
• Bordetella parapertussis
• Whooping cough-like illness.

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Bordetella pertussis

• Global regulatory system is controlled by BvgAS
• Over 100 different genes
• BvgAS is active at 37oC and inactive below 26oC
• BvgAS is repressed by MgSO4 or nicotinic acid at
  37oC.

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BvgAS Two Component System

• Phosphorelay

BvgS, a membrane bound hybrid sensor
BvgA, a typical response regulator
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Phenotypic Phases
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BvgAS regulated phenotypic phases

• Bvg-
• Expression of Bvg-repressed (class 4) genes (vrg)
• Lack of expression of Bvg-activated genes (vag)
• Bvgi
• Expression of class 2 genes including those
  encoding adhesins and bvgAS itself and class 3
  genes (bipA)
• Lack of expression of class 1 and 4 genes.
• Bvg
• Expression of class 1 genes including those
  encoding toxins (cyaA and ptxA) and class 2 genes
• Lack of expression of class 3 and 4 genes.

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Activation of BvgA-P

• Association of BvgA-P monomers to form dimers
• Recognization of and specific binding to DNA by
  BvgA dimers
• Oligomerization of BvgA dimers along a DNA
  molecule
• Interaction with RNA polymerase

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Model for the DNA-Bound BvgA
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Distinguishable Activities of BvgA-P

• Activation of class 2 genes
• One dimer of BvgA-P binds to a primary,
  high-affinity binding site centered at -88.5
• Two additional Bvg-A dimers bind the secondary
  binding region overlapping -35 region which
  interacts with a-CTD of RNAP

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Three Distinguishable Activities

• Activation of class 3 genes
• One dimer of BvgA-P binds to a high-affinity
  binding site at 65.5 and a second dimer binds at
  the adjacent region overlapping the -35 region.

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Three Distinguishable Activities

• Activation of class 1 genes
• Two Bvga dimers to relatively low affinity
  binding sites located -80-125 or -120-165 bp,
  respectively.

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Selected virulence factors of B. pertussis
activated by BvgAS

• Pertussis Toxin (PTX)
• ADP-ribosylation of G-proteins
• Adenylate cyclase toxin (CYA)
• Invasive adenylate cyclase and hemolysin
• Dermonecrotic toxin (DNT)
• Transglutaminase
• Filamentous hemagglutinin (FHA)
• Adhesion and colonization
• Pertactin (PRN)
• Adhesion and colinization
• Tracheal colonizing factor (TCF)
• Adhesion and colonization
• BrkA
• Serum resistance

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Biofilm Development in Bordetella

• Planktonically growing bacterial cells are
  different from these in biofilms.
• Bordetella often form the biofilms in infected
  areas.

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What is Biofilm?

• Biofilms are composed of microbial communities
  that are attached to an environmental surface. 

Steps Adsorption Irreversible
attachment Extracelular polymer substances
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What is Biofilm?

• Biofilms may form
• 1.  on solid substratums in contact with
  moisture,
• 2.  on soft tissue surfaces in living organisms,
  and 
• 3.  at liquid-air interfaces. 

Rocky Mountains
Dental biofilms
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Biofilm and Industry

• Pipe plugging, corrosion and contamination.
• Water treatment pulp and paper manufacturing
• Use of biofilms to treating sewage and industrial
  waste streams.
• Large scale production of biochemicals

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Biofilm and Human Diseases

• Bacterial endocarditis (infaction of the inner
  surface of the heart), ear infaction and so on.
• Many nosocomial infections
• Highly resistant to antibiotics

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Hypothesis of the Paper

• BvgAS mediates the signal transduction that
  regulates biofilm formation of Bodortella.
• Bordetella forms the biofilms in both Bvg and
  Bvg- states
• The Bvg mediated control of the biofilm
  development is exerted at a step subsequent to
  the initial attachment.

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Phase-Locked Mutants

• Strain Rb50
• A wild type Bordetella strain
• Strain RB53
• Bvg strain, always active
• Strain RB53i
• Bvgi strain, always in an intermediate state
• Strain RB55
• Bvg- strain, a deletion of the entire BvgAS
  locus, always inactive

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Formation of a Bacterial Ring Structure of
Bordetella
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BvgAS is involved in the Biofilm formation
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Biofilm population of the wild type and
phase-locked mutants
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Scanning EM of the B.bronchiseptica
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TABLE 1. Antibiotic sensitivity of biofilm-grown
and planktonically grown wild type B.
bronchisepticaa

• Drug MBC-P (mg/ml) MBC-B (mg/ml)
  MBC-B/MBC-P
• Erythromycin 0.05 gt32b gt640
• Streptomycin 0.75 300 400
• Kanamycin 0.02 4 200
• Gentamicin 0.005 0.8 160
• Ampicillin 0.5 500 1,000
• Ciprofloxacin 0.005 1 200
• a MBC-P, MBC of planktonically grown cells
  MBC-B, MBC of biofilm-grown cells. Values are
  based on results of three different experiments.
• b The antibiotic was partially soluble above this
  concentration.

MBC, minimum bactericidal concentration that
completely inhibits growth.
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What about other Bordetella?
For B. parapertussis
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Biofilm Formation of B. parapertussis
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Biofilm Formation of B. pertussis
For B. pertussis
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Conclusions

• Bodetella of all three species can form biofilms.
• The BvgAS signal transduction system is involved
  in biofilm formation.
• BvgAS and BvgASi are required for the formation
  of biofilms.
• Formation of biofilms increases the resistance to
  antibiotics by 100 to 1000 folds

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Questions

• The mechanism by which the BvgAS controls the
  formation of biofilms of Bodoretella?
• Quorum sensing?
• What genes are essential for the biofilm
  development in Bordetella?
• What are the different roles of BvgAS and BvgASi
  in the biofilm development?