Dr. Jan Decker

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Dr. Jan Decker

• 235 Veterinary Sci/Micro
• jdecker_at_u.arizona.edu
• 621-8942
• http//microvet.arizona.edu/Courses/MIC419/mic419i
  nstructor.html schedule

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Biofilms

• Microbial Communities and Infectious Disease

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Prokaryotic Cell Structure
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Bacteria Are Prokaryotes

• DNA not surrounded by a membrane
• one circular chromosome
• not associated with histones
• Lack membrane-enclosed organelles

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Prokaryotes

• Unicellular
• Divide by binary fission
• Include bacteria and archaea (no peptidoglycan)
• Cell walls usually peptidoglycan, a complex
  polysaccharide

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NAG and NAM
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Peptidoglycan Structure
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Gram Positive Cell Wall

• Thick peptidoglycan
• Teichoic acid
• ion flow
• protection
• antigen specificity

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Gram Negative Cell Wall

• Thin peptidoglycan
• No teichoic acid
• Outer membrane
• LPS endotoxin
• protection
• porins

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Bacterial Shapes
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Contemporary Endosymbiont

• Photosynthetic cyanobacteria (pink) inside
  flagellated protozoan Cyanophora

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Biofilms

• Biofilm a microbial community attached to a
  surface
• May be one or several organisms
• Sessile organisms attached
• Planktonic organisms free living

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Antoni van Leeuwenhoek
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Benefits to Microbes

• Adherence to hospitable locale
• Syntrophic metabolism
• Horizontal gene transfer
• Antibiotic resistance
• Disease reservoir

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Environmental Biofilms
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Environmental Biofilms
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Environmental Biofilms
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Environmental Biofilms
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Infectious Disease Biofilms
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Infectious Disease Biofilms

• On implants

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Development of Biofilm

• Initial attachment to surface
• Cellcell attachment
• Production of extracellular polysaccharides

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Glycocalyx

• Complex exopolysaccharide
• Adhesion
• Protection from
• biocides
• antibiotics
• bacteriophage
• free-living amoebae
• WBC

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Adhesion

• Depends on surface
• shape
• charge
• hydrophobicity
• hydrodynamics

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Surfaces for Biofilms

• Abiotic (pipes, rocks, implants)
• Host tissues
• Other microbes

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Biological Surface Conditioning

• Plasma proteins
• albumin
• complement
• fibronectin
• Inorganic salts
• Glycocalyx

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Adhesins

• Proteins on bacterial pili

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Biofilm Architecture

• Hydrated 73-98 noncellular
• Water channels
• Architecture varies with species

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Climax Biofilm

• Reaches optimal size
• External organisms become planktonic, leave to
  colonize elsewhere
• Cells nearest the surface become quiescent or die
  due to limited O2 and nutrients, increased waste

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Cell-Cell Communication

• Quorum sensing
• cells secrete molecules
• nearby cells sense population density
• Regulates biofilm architecture
• Regulates gene expression
• Horizontal gene transfer increases 10-600 X

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Quorum Sensing
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Quorum Sensing
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Gene Regulation in Biofilms

• Change in gene expression in attached cells
• Increased production of glycocalyx
• Change in energy metabolism
• P. aeruginosa changes expression of 40 genes
• S. aureus increases expression of genes for
  glycolysis, fermentation (low O2)
• Increased antibiotic resistance

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Horizontal Gene Transfer
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F Pilus and Conjugation

• Bacteria with F plasmid can form pilus and
  transfer plasmid and sometimes part of chromosome
  to another bacterium

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Antimicrobial Resistance

• 103-104 higher doses needed to kill biofilm
  compared with planktonic organisms
• Biofilm is a molecular filter?
• Low metabolism of cells?
• Reduces drug activity?

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Resistance to Immune System Clearance

• Phagocytosis (macrophages, PMNs)
• surface binding
• engulfed
• killed with digestive enzymes and reactive
  oxygen molecules (O2., H2O2, NO)

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Resistance to Immune System Clearance

• Vaccinated rabbit to bacteria
• Increased antibody levels
• No increase in phagocytosis of biofilm

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Resistance to Immune System Clearance

• Biofilm E. coli are less likely to be killed by
  human PMN (neutrophils) in vitro
• Resistant to active oxygen species produced by PMN

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Some Medically Important Biofilms

• CDC estimate 65 of human bacterial infections
• Dental plaque
• Infectious kidney stones
• Endocarditis
• Catheters
• Cystic Fibrosis

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Infectious Kidney Stones

• 15-20 involve urinary tract infection
• Bacterium ? biofilm ? mineralization
• StruviteMg(NH4)PO4.6H20
• Carbonate-Apatite Ca10(PO4)6CO3

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Infectious Kidney Stones
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Infectious Kidney Stones

• Causative organisms have urease
• Urea ? NH4 H2CO3
• Biofilm concentrates urease ? crystal formation

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Endocarditis

• Biofilm of bacteria host components on valve
  vegetation
• Requires prior valve injury
• 200X increase in antibiotic resistance
• Rabbit model block biofilms ? acute virulent
  infection

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Cardiac Vegetation
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Cystic Fibrosis

• Mutation in chloride channel in epithelial cells
• 1sr stage intermittent infections
• 2nd stage permanent infection Pseudomonas
  aeruginosa
• Mucoid type overproduces alginate
• Antibiotic resistance

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Pseudomonas aeruginosa
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Key Concepts

• Describe a biofilm and give some examples.
• Discuss the advantages to bacteria of living in
  biofilms.
• Explain the endosymbiotic theory of eukaryotic
  cell evolution.

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Key Concepts

• Describe the development and climax status of a
  biofilm.
• Discuss interbacterial communication in
  biofilms. Correctly use the term quorum sensing
  in your answer.
• Describe some of the changes in gene function
  associated with biofilm formation.

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Key Concepts

• Describe three methods of horizontal gene
  transfer (HGT) in bacteria and explain how HGT is
  enhanced in biofilms.
• Describe how biofilms play a role in infectious
  disease. Discuss a specific example.