Microbial Growth

• Binary fission:

  • Involved in cell division
  • Mutations in FTS genes result in filamentous forms
  • Temperature sensitive: "Filamentous temperature sensitive"
  • Found in archaea and bacteria
  • FTS proteins interact to form division apparatus called "divisome"
• Proteins involved in cell division:

  • FtsZ forms the divisome (attached to cytoplasmic membrane)
  • ZipA & FtsA anchor FtsZ to the cytoplasmic membrane, FtsA has ATPase activity
  • FtsI (penicillin-binding protein) is a peptidoglycan biosynthesis protein
  • FtsK separates chromosomes for daughter cells to have chromosomal material
• MreB protein:

  • Major shape-determining protein in rods, spirillum, spirochetes
  • Absent in cocci
  • Forms actin-like cytoskeleton
• Peptidoglycan synthesis and cell division:

  • Coccus has FtsZ ring
  • Peptidoglycan synthesis in cocci localized in one point
  • Peptidoglycan synthesis in bacilli occurs in different places of the cell
  • Autolysis prevention requires coordination of different molecules:
  • Autolysin introduces breaks in peptidoglycan layer
  • Transglycosylase forms glycosidic bonds for new peptidoglycan materials
  • Transpeptidase forms crosslinking of tetrapeptide chains
  • Bactoprenol is a carrier molecule for peptidoglycan precursor molecules
• Transpeptidation:

  • Terminal D-alanine is removed for transpeptidation
  • Transpeptidase enzyme is the penicillin binding protein
• Effect of binary fission on population:

  • Exponential growth: number of cells doubles in constant time interval
  • Generation time is the time a bacterial cell takes to divide into two cells
  • Bacterial growth curve:
  • Lag phase: lag in population increase, cells prepare for division
  • Exponential phase: active cell division
  • Stationary phase: highest number of cells, cells divide and die
  • Death phase: more cells die than divide, spore formers may be found
• Measurements of microbial growth:

  • Direct total count:
  • Microscopic counts using DAPI or acridine orange stains
  • Flow cytometer counts every particle
  • Viable cell count:
  • Viability staining with LIVE/DEAD Baclight Viability Stain
  • Plate count method for growing colonies
• Continuous culture:

  • Maintains culture in exponential phase for long periods
  • Useful for physiologic studies
• Biofilms:

  • Aggregates of microorganisms at solid-liquid interface
  • Cells embedded in extracellular polymeric substance (EPS)
  • EPS produced by extracellular DNA, polysaccharides, lipids, proteins
  • Quorum-sensing mediated activity of cells in biofilm formation
  • Presence of EPS deters antibiotics from penetrating biofilms
  • Biofilm formation involves initial adhesion, formation of EPS, cell division, and mature biofilm
• FTS proteins are involved in cell division and mutations in FTS genes can result in filamentous forms
• FTS proteins are temperature sensitive, known as "Filamentous temperature sensitive", and are found in archaea and bacteria
• FtsZ is structurally similar to eukaryotic tubulin and defines the division plane in the cell
• FtsZ ring forms after DNA replication
• Proteins involved in cell division:

  • FtsZ forms the divisome and is attached to the cytoplasmic membrane
  • ZipA and FtsA anchor FtsZ to the cytoplasmic membrane, with FtsA also having ATPase activity
  • FtsI is a peptidoglycan biosynthesis protein
  • FtsK separates chromosomes so each daughter cell will have chromosomal material
• MreB protein is a major shape-determining protein in rods, spirillum, and spirochetes, but absent in cocci
• Peptidoglycan synthesis during cell division:

  • In cocci, peptidoglycan is localized in one point during cell division
  • Peptidoglycan synthesis in bacilli occurs in different places of the cell
  • Autolysin introduces breaks in the peptidoglycan layer
  • Transglycosylase forms glycosidic bonds for new peptidoglycan materials
  • Transpeptidase forms crosslinking of tetrapeptide chains
• Carrier molecule bactoprenol allows transfer of peptidoglycan precursor molecules along the cytoplasmic membrane
• Exponential growth results in the doubling of cells in a constant time interval, with generation time being the time a bacterial cell takes to divide into two cells
• Batch culture involves setting up medium in glassware, inoculating, and incubating without adding or removing products
• Bacterial growth curve stages:

  • Lag phase: cells prepare for division
  • Exponential phase: active cell division
  • Stationary phase: highest number of cells, some divide and die
  • Death phase: more cells die than divide, spore formers may be present
• Measurements of microbial growth:

  • Total count methods include microscopic counts, flow cytometer, and plate count
  • Viable cell count methods include viability staining and plate count
• Turbidity measurement indicates growth in the medium but is not a viable count
• Continuous culture in a chemostat maintains culture in exponential phase for extended periods, useful for physiologic studies
• Biofilms are aggregates of microorganisms at a solid-liquid interface, with high cell density and composed of different species that interact with each other
• Biofilm formation involves initial adhesion of planktonic cells to a surface, followed by the formation of EPS and heterogeneity, leading to mature biofilms with social interactions
• • FtsZ:
  • Sutrcturally similar to eukaryotic tubulin
  • Defines division plane in the cell
  • FtsZ ring forms after DNA replication