Microbial Growth

Microbial Growth

1. Growth in Batch Culture

   • "Growth" is generally used to refer to the generally used to refer to the acquisition of biomass leading to cell division or reproduction
   • A "batch culture" is a closed system in growth medium in which no additional nutrient is added after inoculation of the broth
   • Passes through Four Stages:
     • 1) Lag Stage
       • 2) Log Stage- exponential growth
         • 3) Stationary Phase
           • 4) Death Stage

2. Mean Generation Time and Growth Rate

   • Growth during log phase is described by the exponential equation Yt= Yoe^ut
     • Yt- concentration of bacteria at time t
     • Yo- initial concentration of bacteria
     • u- specific growth rate in units of min-1
   • The mean generation time of doubling time, during log phase is given by the equation td= ln(2) / u
     • The doubling time can also be determined from a semilogarithmic plot of growth

3. Measurement of Microbial Growth

   • Microscopic Cell Counts
     • -Calibrated "Petrol-Hausser Chamber", similar to hemacytometer, can be used -Generally very difficult for bacteria since cells tend to move in and out of counting field -Useful for organisms that cants be cultured -Special stains can be used for specific purposes e.g. serological stains or stains for viable cells
   • Serial Dilution & Colony Counting
     • -Also known as "viable cell counts" -Concentrated samples are diluted by serial dilution -The diluted samples can be either plated by spread plating or by pour plating CFU= # of colonies counted/ (dilution factor)( volume plated, in mL) Std.D= SqRt # of colonies counted/ (dilution factor)( volume plated, in mL)
   • Membrane Filtration
     • -Used for samples with low microbial concentration -Sample is filtered through a membrane filter -The filter is placed on nutrient agar medium and incubated -Colonies grow on the filter and can be counted ~0.45um pore size
   • Turbidity
     • -Based on the diffraction of "scattering" of light by bacteria in a growth culture -Light scattering is measured as optical absorbance in a spectrometer -Optical absorbance is directly proportional to the concentration of bacteria in the suspension
   • Mass Determination
     • -Cells can be removed from a broth culture by centrifugation and weighed to determine the "wet mass"
   • Measurement of Enzymatic Activity or Other Cellular Components

4. Continuous Culture

   • -A "continuous culture" is an open system is which fresh media is continuously added to the culture at a constant rate, and old broth is removed at the same rate
   • - This method is accomplished in a device called a chemostat
   • - Typically, the concentration of cells will reach an equilibrium level that remains constant as long as the nutrient feed is maintained

5. Factors Influencing Growth

   • Growth vs. Tolerance
     • Growth- refers to the acquisition of biomass leading to cell division, or reproduction
     • Tolerance- survive under condition in which they cannot grow
     • The suffix "-phile" is often used to describe conditions permitting growth, whereas the term "tolerant" describes conditions in which the organisms survive, but don't necessarily grow
     • For example, a "thermophilic bacterium" grows under conditions of elevated temperature, while a "thermotolerant bacterium" survives elevated temperature, but grows at a lower temperature like Giobacillus Thermophilus
   • Temperature
     • Most bacteria grow throughout a range of approximately 20C, with the maximum growth rate at a certain, "optimum temperature"
     • Psychrophiles: Grows well at 0C; optimally between 0C-15C
     • Psychrotrophs: Can grow at 0-10C; optimum between 20-30C and maximum around 35C
     • Mesophiles: Optimum around 20- 45C
     • Moderate thermophiles: Optimum around 55- 65C
     • Extreme thermophiles (Hyperthermophiles): Optimum around 80- 113C
   • Obligate (Strict) vs. Facultative
     • "Obligate" means that given condition is required for growth
     • "Facultative" means that the organism can grow under the condition , but doesn't require it. The term is often applied to sub-optimal conditions
     • An obligate thermophile requires elevated temperatures for growth, while a facultative thermophile may grow in either elevated temperatures or lower temperatures
   • pH
     • Acidophiles: Grow optimally between ~pH 0 -5.5
     • Neutrophiles: Grow optimally between ~pH 5.5 -8
     • Alkalophiles: Grow optimally between ~pH 8 -11.5
   • Salt Concentration
     • Halophiles require elevated salt concentrations to grow; often require 0.2M ionic strength or greater and some may grow at 1M or greater; example, Halobacterim
     • Osmoyolerant (halotolerant) organisms grow over a wide range of salt concentrations or ionic strengths; for example, Staphylococcus aureus
   • Oxygen Concentration
     • Strict Aerobes: Require oxygen for growth (~20%)
     • Strict Anaerobes: Grow in the absence of oxygen; cannot grow in the presence of oxygen
     • Facultative anaerobes: Grow best in the presence of oxygen, but are able to grow (at reduced rates) in the absence of oxygen
     • Aerotolerant Anaerobes: Can grow equally well in the presence or absence of oxygen
     • Microaerophiles: Require reduced concentrations of oxygen (~2-10%) for growth

6. Growth in Natural Environments

7. Quorum Sensing

   • A mechanism by which members of a bacterial population can behave cooperatively, altering their patterns of gene expression (transcription) in response to the density of the population
   • In this way, the entire population can respond in a manner most strategically practical depending on how sparse of dense the population is
   • Mechanism: As the bacteria in the population grow, they secrete a quorum signaling molecule into the environment (for example, in many gram negative bacteria the signal is an acyl homoserine action, HSL)