Chapter 6: Microbial Growth

1
Chapter 6Microbial Growth
2
Microbial Growth

• Microbial growth growth in population
• Increase in number of cells, not cell size
• Two main categories of requirements for microbial
  growth
• Physical requirements (environmental conditions)
• Temperature, pH, osmotic pressure
• Chemical requirements

3
Physical Requirements for Growth Temperature

• Temperature
• Minimum growth temperature
• Optimum growth temperature
• Maximum growth temperature
• Three main classifications
• Psychrophiles (optimum 120C)
• Psychrotrophs (optimum 230C)
• Mesophiles (optimum 370C)
• Thermophiles (optimum above 500C)

4
Physical Requirements for Growth Temperature
Refrigeration
Cause majority of food spoilage
Figure 6.1
5
Hansens Disease(Leprosy)

• Mycobacterium leprae
• Optimal growth temperature 30C
• Grows in peripheral nerves, nasal mucosa and skin
  cells

Figure 22.8
6
Physical Requirements for Growth pH

• pH
• Most bacteria grow between pH 6.5 and 7.5
• Molds and yeasts grow optimally between pH 5 and
  6
• Acidophiles grow in acidic environments (pHlt5.5)
• Alkaliphiles grow in basic environments (pHgt8.5)
• Acidic foods (pickles, sauerkraut) preserved by
  acids from bacterial fermentation
• Growth media used in the laboratory contain
  buffers

7
Physical Requirements for Growth Osmotic Pressure

• Osmotic Pressure
• Hypertonic environments (high osmotic pressure),
  increased salt or sugar, cause plasmolysis
• Obligate halophiles require high osmotic
  pressure
• Facultative halophiles tolerate high osmotic
  pressure (gt2 salt)
• Nutrient agar has a high percentage of water to
  maintain low osmotic pressure (bacterial cells
  are 80-90 water)

Low osmotic pressure
High osmotic pressure
Water flow
High solute concentration/ Low water concentration
Low solute concentration/ High water concentration
8
Physical Requirements for Growth Osmotic Pressure

• Plasmolysis cell growth is inhibited when the
  plasma membrane pulls away from the cell wall
• Added salt or sugar is another method of
  preserving food

Hypertonic solution (high osmotic pressure)
Isotonic solution
Figure 6.4
9
Chemical Requirements for Growth

• Carbon
• Structural organic molecules, energy source
• Heterotrophs use organic carbon sources
• Autotrophs use CO2
• Nitrogen, Sulfur, Phosphorus
• For synthesis of amino acids, nucleotides,
  vitamins, phospholipids
• Most bacteria decompose proteins to obtain N
• Inorganic ions are sources for these elements
  (NH4, NO3-, PO43-, SO42-)

10
Chemical Requirements for Growth

• Trace Elements (Iron, Copper, Zinc)
• Inorganic elements required in small amounts,
  usually as enzyme cofactors
• Often present in tap water
• Organic Growth Factors
• Organic compounds obtained from the environment
  (i.e. the organism cannot synthesize them)
• Vitamins, amino acids

11
Chemical Requirements for Growth Oxygen

• Oxygen (O2)

Obligate aerobes Facultative anaerobes Obligate anaerobes Aerotolerant anaerobes Microaerophiles

12
Chemical Requirements for Growth Oxygen

• Aerotolerance of individual organisms depends on
  their ability to handle oxygen toxicity
• Oxygen radical species O2-, O22-, OH
• Presence/lack of enzymes that neutralize toxic
  oxygen species
• SOD (Superoxide dismutase)
• Catalase/peroxidase

.
13
Chemical Requirements for Growth Oxygen

• Oxygen (O2)

Obligate aerobes Facultative anaerobes Obligate anaerobes Aerotolerant anaerobes Microaerophiles

Require oxygen, but at lower levels than in the
air
Express SOD and catalase
Dont express SOD/catalase
Tolerate oxygen (express SOD/catalase) but
incapable of using it for growth
14
Culture Media

• Culture Medium Nutrients prepared for microbial
  growth
• Source of energy, carbon, nitrogen, sulfur,
  phosphorus, trace elements and organic growth
  factors
• Sterile No living microbes
• Inoculum Introduction of microbes into medium to
  initiate growth
• Culture Microbes growing in/on culture medium

15
Culture MediaAgar

• Complex polysaccharide
• Used as solidifying agent for culture media in
  Petri plates, slants, and deeps
• Generally not metabolized by microbes
• Agar is not a nutrient
• Liquefies above 100C
• Can incubate at a wide range of temperatures

16
Culture Media
17
Anaerobic Culture MediaBroth cultures

• Reducing broth media
• Contain chemicals (thioglycollate) that combine
  with dissolved O2 to deplete it from the media

18
Anaerobic Culture MethodsAgar Cultures

• Anaerobic jar
• Oxygen and H2 combine to form water

Figure 6.5
19
Culture MediaSelective and Differential Media

• Selective media suppress growth of unwanted
  microbes and encourage growth of desired microbes
• Differential media make it easy to distinguish
  colonies of different microbes

Enterobacter aerogenes on EMB
E. coli on EMB
Figure 6.9b, c
20
Obtaining Pure Cultures

• A pure culture contains only one species or
  strain
• A colony is a population of cells arising from a
  single cell or spore or from a group of attached
  (identical) cells
• One colony arises from one colony-forming unit
  (CFU)
• Specimens (pus, sputum, food) typically contain
  many different microorganisms
• Common way to isolate a single species from a
  mixture of microorganisms Streak plate method

21
Streak Plate Method for Isolation of a Pure
Species

• Use loop to pick colony
• Inoculate broth
• Pure culture

Figure 6.10a, b
22
Microbial Growth in HostsBiofilms

• Microbial communities
• 3-dimensional slime
• i.e. dental plaque, soap scum
• Share nutrients
• Sheltered from harmful factors
• Cell-to-cell communication quorum sensing

Figure 6.5
Bacterial biofilm growing on a micro-fibrous
material
23
Microbial Growth in HostsBiofilms Quorum
Sensing

• Quorum sensing allows a form of bacterial
  communication
• Individual cells can sense the accumulation of
  signaling molecules (autoinducers)
• Informs individual cells about surrounding cell
  density
• May change the behavior (gene expression) of
  individual cells
• Results in a coordinated response by the whole
  population

http//biofilmbook.hypertextbookshop.com/public_ve
rsion/
24
Prokaryotic ReproductionBinary Fission
Figure 6.11
25
Reproduction in ProkaryotesGeneration Time

• Generation time the time required for one
  population doubling
• Varies with species and environmental conditions

26
Reproduction in ProkaryotesGeneration Number

• Generation number the number of times a cell
  population has doubled in a given time under
  given conditions

Figure 6.12b
27
Reproduction in ProkaryotesGrowth Plot
Logarithmic
Arithmetic
Figure 6.13
28
Bacterial Growth Curve

• Lag little/no cell division
• Adapting to new medium
• Metabolically active
• Log exponential growth
• Most metabolically active
• Gen. time at constant minimum
• Stationary equilibrium phase
• Growth rate death rate
• Nutrients exhausted, waste accumulation, pH
  changes
• Death logarithmic decline

Figure 6.14
29
Measuring Microbial Growth

• To determine the size of a bacterial population
  in a specimen, cell counting techniques are used
• Often there are too many cells per ml or gram of
  specimen
• A small proportion of the specimen (a dilution)
  is counted
• The number of cells in the original specimen can
  be calculated based on the count in the small
  dilution

30
Direct Measurements of Microbial GrowthViable
Cell Count

• Plate Counts Perform serial dilutions of a
  sample
• How many cells are in 1 mL of original culture?

DF1
DF
10-3
10-5
10-2
10-4
10-1
Figure 6.15, top portion
31
Direct Measurements of Microbial GrowthViable
Count

• Inoculate one agar plate with each serial dilution

Figure 6.16
32
Direct Measurements of Microbial GrowthViable
Count

• After incubation, count colonies on plates that
  have 30-300 colonies (CFUs)

Figure 6.15
33
Direct Measurements of Microbial Growth

• Filtration
• Ideal when microbial density is low in a sample

Figure 6.17a, b
34
Direct Measurements of Microbial Growth
Disadvantages -Likely to count dead
cells -Motile cells can be difficult to count
Figure 6.19
35
(No Transcript)