E. Coli

• Describe Escherichia coli

  • Type of enterobacteriaceae: gram negative, motile, rod shaped and facultative metabolism
  • Causes diarrhoea, UTI, septicaemia and meningitis
• Describe UPEC pathogenesis

  1. Contamination of periurethral area with a uropathogen from the gut, leading to urethral colonisation and migration to the bladder.
  2. Colonisation of the bladder and fibrinogen accumulation in the catheter due to inflammatory response.
  3. Neutrophils begin to clear infection, but insufficient immune response can lead to ascension of bacteria into the kidneys.
• Describe type I Fimbriae in E. coli pathogenesis

  • Pathogenic UPEC strains mutate FimH, allowing them to alter binding from tri-mannose to mono-mannose, which is expressed in the bladder (uroplakins)
  • Mediates bladder colonisation and invasion
  • Attachment of uroplakins mediates engulfment of bacteria, which allows dormant bacteria to survive within the cell. These bacteria can then be reactivated and lead to recurrent infections.
• Describe the structure of type I Fimbriae

  • Helical array of pillin protein, known as FimA, to form a rod structure
  • Short tip contains the adhesion protein, FimH, and two adaptor proteins, FimF and FimG
  • Mutations in FimH allow UPEC to bind to different tissues and mediates uroepithelial cell invasion
• What is phase and antigenic variation?

  Phase variation- switch genes on or off

  Antigenic variation- alter the gene sequence and therefore protein structure
• Describe different mechanisms of bacterial phase and antigenic variation

  1. DNA inversion- phase variation of UPEC type I pili
  2. DNA methylation- phase variation of UPEC P pili
  3. Complex recombination systems- phase and antigenic variation of N. gonorrhoeae type IV pili
  4. Slipped strand mispairing- e.g., N. gonorrhoeae pili C
• Describe the organisation and regulation of Fim genes

  • All Fim genes encoded within a single operon controlled by a single promoter
  • at 314 bp there is a invertible sequence defined by inverted repeats that flank the promoter sequence
  • upstream of the invertible sequence is regulatory proteins such as recombinases, FimE and FimB
• Describe type I pili phase variation

  • FimE and FimB encode recombinases, which bind to inverted repeats and flip the orientation of DNA
  • The promoter must be facing the right direction in order for fimbriae to be expressed
  • This allows bacteria to turn Fimbriae expression on or off and target specific niches, as well as avoid immune action.
• Describe the structure of P fimbriae

  • Thick helical rod of repeating PapA subunits
  • PapH anchors PapA rod to outer membrane
  • Flexible tip fibrillum contacting PapG and PapE with two adaptor proteins, PapK and PapF
  • PapG variation induced by addition of N-acetyl galactosamine sugars
• Describe phase variation of the Pap genes

  • Promoter for the pap genes contains two GATC sites, which can be methylated by dam methylase
  • distal GATC sites methylated leads to no expression of pap
  • proximal GATc sites methylation leads to inhibition of the binding of inhibitory protein, LRP, which allows pap gene expression.
• Describe type IV pili

  • Initiate infection by attaching to host cells involving CD46 receptors and interaction with nearby pili promotes aggregation.
  • Mediates twitching motility over epithelial surface
  • Composed primarily of pilin E
  • PilC is involved in assembly and adhesion
  • PilE and PilC undergo phase variation and antigenic variation, which prevents the host from generating a protective immune response.
• Describe antigen variation of PilE

  • PilE encodes pilin protein with repeated microcasettes
  • Homologous recombination occurs between PilE and PilS, which leads to DNA exchange
• Describe phase variation of PilC

  • Slipped strand mispairing
  • Guanine nucleotide repeats
  • During DNA replication, slippage of strand leads to addition or deletion of guanine nucleotides resulting in frameshift mutations
  • Premature STOP codon leads to truncated non-functional PilC, which is essential for proper assembly and maturation of pili