Chapter 5 Microbial Growth and Nutrition
Microorganisms can be found in the harshest of environments including deep sea thermal
vents, Polar Regions, and hot springs. These organisms survive in what we consider extreme
environments but to the microbe they evolved mechanisms to withstand the environmental
conditions they live in. These organisms have limited environmental ranges in which the species
can grow; for example, every species has a specific temperature range in which the organism can
survive. Organisms found in deep sea thermal vents would be extremely difficult to culture at sea
level due to the pressure and specific nutrients available.
Microbial growth refers to the increase in population of microbes. Therefore the process of
reproduction results in growth. There is no relationship to microbe size and growth as there is
when we consider human growth or plant growth. After a bacterial cell undergoes binary
fission the two daughter cells are fully mature adult cells capable of the same processes as the
parent cell. Microbiologists refer to the amount of time an organism takes to reproduce as the
generation time. The generation time varies based on chemical (ex. food availability) and
physical (ex. temperature) conditions. Escherichia coli for example can reproduce in as little as
20 minutes under optimal conditions, which are the physical and chemical factors that are the
most favorable for that specific organism. Mycobacterium tuberculosis has a generation time of
15-20 hours under ideal conditions, and more than 48 hours in less than optimal conditions. The
human body’s optimal temperature is 37o C, therefore most human pathogens have optimal
temperatures for growth within a few degrees of 37o C. The growth rate is the number of
generations per hour. An important note is that growth rate and generation time are inversely
related, for example, if the generation time of E. coli is 20 minutes, then the growth rate is 3
generations per hour.
Because bacteria grow this way, their population size grows very rapidly. This type of growth,
where the population doubles with each generation is called logarithmic or exponential growth.
Allowed to continue unchecked, exponential growth can quickly lead to a lot of cells. For
example:
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Start with 1 E. coli with a generation time of 20 minutes
In 7 hours there would be roughly 1,000,000 cells
In 10 hours there would be around 1,000,000,000 cells
In 24 hours there would be about 1, 000,000,000,000,000,000,000 cells!
You can see that bacteria growing exponentially expand their populations very rapidly. Luckily,
even with logarithmic growth, we are not completely taken over by bacteria. Their growth is
limited by environmental factors.
Microbial growth is predictable in liquid media given the organism has access to
appropriate nutrients. Once an organism is inoculated into a growth media, the organism will
experience a Lag phase, while the organism is adapting to the environment. Following the Lag
phase, the organism will begin growing at an exponential rate. This stage is referred to as the
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Log phase, during which the organism’s population is doubling during each generation time.
Eventually the nutrients begin to become limited and growth slows. This phase is called the
Stationary phase, during which the number of cells reproducing equals the number of cells that
are dying. Finally the Death phase occurs, when the number of dying cells is greater than the
number of cells reproducing (Figure 5.1).
Figure 5.1. Cell population growth curve generated using spectrophotometry.
Measuring Microbial Growth
Microbiologists are able to measure and monitor cell growth by using spectrophotometry.
Spectrophotometry is considered an indirect method to estimate a population size or density. A
spectrophotometer is used to measure the turbidity of a solution, also called the optical density.
A light beam is directed through a liquid sample, and a sensor records how much light passed
through on the other side. The higher the cell density (correlates to a greater bacterial population)
in a liquid culture, the more turbid the solution will be and less light will travel through the
sample. A computer within the spectrophotometer gives you either an absorbance output reading,
in which a higher absorbance reading indicates a higher cell density (Figure 5.2) or a
transmittance value where a higher transmittance indicates a lower cell density. It is possible to
calculate between %T and absorbance by the following calculation:
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Absorbance = -log(%T/100)
An important note is this technique is only useful when the cell density exceeds 1 million cells
per milliliter, densities below this number do not produce enough turbidity for an accurate
reading. Another important note is that spectrophotometry only correlates to a total number of
cells and does not distinguish between living and dead cells.
5.2.A
5.2. B
Figure 5.2. A) Demonstrates a spectrophotometer measuring a cell sample with 10,000,000 cells.
Figure 5.2. B) Demonstrates a spectrophotometer measuring a cell sample with 1,000,000,000 cells. A
sample with more growth (higher cell density) will give a reading on the spectrophotometer as a
higher absorbance.
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Nutrients for Growth
Microorganisms are capable of using a wide variety of nutrients for their energy needs, in
order to build organic molecules for cellular structures, which are required for growth. The most
common nutrients must contain elements such as carbon, oxygen, nitrogen, and hydrogen. Other
elements needed for growth include phosphorus, magnesium, potassium, iodine, and sulfur.
Elements that are required in very small amounts are referred to as trace elements and include
iron, iodine and zinc. When an organism is growing, the cells are transforming chemical
elements into carbohydrates, proteins, lipids, and nucleic acids: the 4 macromolecules of life also
considered growth factors. Fastidious organisms have very specific and complex nutritional
requirements and when grown in the lab require the use of special media.
Osmotic Pressure
Because most microbes get their nutrients from surrounding water, osmotic pressure is really
important. Cells will experience the following conditions depending on their environment
(Figure 5.3):
Hypertonic: the concentration of solute outside of the cell is in is greater than the concentration
of solute inside the cell. The solute is the dissolved substance such as sugar or salt.
Hypotonic: the concentration of solute outside the cell is in is less than the concentration of
solute inside the cell.
Isotonic: concentration of the solute outside the cell is the same as the concentration of solute
inside the cell.
Figure 5.3. Example of how water
diffuses when a cell is placed into
varying salt concentrations.
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Certain compounds act as growth inhibitors, which affect the osmotic pressure of a cell
and reduce the growth rate or prevent a microorganism from growing all together. A notable
inhibitor is salt. In a hypertonic environment salt inhibits microbial growth by disrupting the
osmotic pressure of the cell. Since water is needed for life, salt reduces the availability of water
to a cell since water will diffuse of a cells cytoplasm when in a high salt environment.
Some genera of microorganisms can live and even thrive in the presence of salt while
others quickly die due to crenation, or shriveling of the cell’s cytoplasm. This results when
water diffuses out of the cell due to a high solute concentration outside of the cell membrane.
Salt tolerant organisms are considered halophiles. Obligate halophiles are adapted to grow
under high osmotic pressure and have an absolute requirement for a high salt concentration.
Obligate halophiles would be found in environments such as the Great Salt Lake in Utah, the
salinity varies greatly from 5% to near 30%, microbes living in the Great Salt Lake would burst
if placed into freshwater. Facultative halophiles do not require high salt concentrations however
they can tolerate high salt conditions. A high salt condition would be considered 5% and above.
Staphylococcus aureus can tolerate salt up to 20%, which allows this organism to colonize the
surface of the skin. The role that salt plays on the osmotic pressure of cells accounts for the
preserving action in jerky and salted fish.
Temperature
When cultivating microorganisms especially novel organism’s microbiologists perform a battery
of tests, which are used to characterize and classify the microorganism. Organisms can be
classified into one of four categories based on their temperature requirements for growth.
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Psychrophiles are cold loving organisms that prefer temperatures between -5 and 20 oC.
Mesophiles are moderate temperature loving organisms and are capable of surviving in
temperatures between 15 and 45 oC.
Thermophiles are heat loving microbes that are capable of growing in temperatures
between 45 and 80 oC.
Hyperthermophiles are extreme heat tolerant and can survive boiling temperatures
between 70 and 105 oC.
Psychroduric these are bacteria that prefer warmer temperatures, but can survive in
colder temps.
Thermoduric these are mesophiles that can survive short periods at higher temperatures.
These bacteria may contaminate food and survive canning and Pasteurization processes.
An important note when classifying organisms by temperature requirements is that the optimal
temperature for growth is usually in the middle of the range discussed above, for example a
psychrophile would have an optimal growth rate at around 10 oC (Figure 5.3). Bacteria can
generally survive if they moved to a lower temperature range than optimum, as this will most
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likely slow their metabolic processes. However, moving bacteria to a higher temperature range
usually results in death as proteins and enzymes are denatured and nucleic acids are damaged.
Even within the given temperature ranges there is room for variation. For example, while
technically a mesophile, the microbe that causes leprosy, Mycobacterium leprae has an optimum
temperature of 33°C, lower than the 37°C that is human core temperature. This lower
temperature accounts for the fact that M. leprae typically affects the extremities, cheeks and
noses of its victims.
Figure 5.3. Four categories of classification based on temperature ranges for growth,
with example microorganisms.
Oxygen
As mentioned in the earlier, oxygen is often an important element for growth. You will
learn later in the semester that oxygen is used as an electron acceptor during aerobic respiration.
For now we can learn how to classify microorganisms based on their oxygen requirements.
Microbes can be classified as obligate aerobes, facultative anaerobes, obligate anaerobes,
microaerophiles, or aerotolerant. Obligate aerobes have an absolute requirement for oxygen,
meaning they cannot grow without oxygen. Facultative anaerobes tend to grow better with
oxygen however they can grow without oxygen. Facultative anaerobes can grow without oxygen
since they either have the ability to switch their metabolism. Obligate anaerobes cannot grow in
the presence of oxygen. Oxygen is actually toxic to these types of organisms. Microaerophiles
have a very specific requirement for oxygen. High concentrations of oxygen can inhibit the
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growth of microaerophiles. Aerotolerant also considered fermenters are organisms are that are
indifferent to oxygen, they can grow in the presence of oxygen yet they do not use oxygen to
transform energy. In lab we will use a special media called fluid thioglycolate (FT) media to
determine oxygen requirements of microbes (Figure 5.4).
Figure 5.4. A) Growth of an obligate aerobe in FT media. B) Growth of a facultative
anaerobe in FT media. C) Growth of an obligate anaerobe in FT media. D) Growth of a
microaerophile in FT media. E) Growth of an aerotolerant organism/fermenter in FT
media. Note: you cannot tell the difference between a facultative anaerobe and an
aerotolerant organism, other biochemical tests would need to be performed to determine
metabolic characteristics.
Organisms capable of surviving in the presence of oxygen will experience damaging effects of
oxygen. Toxic superoxides (O2-) are formed and can oxidize important cell chemicals with in a
cell. Superoxides are neutralized by an enzyme called superoxide dismutase, which forms
hydrogen peroxide in the cell (H2O2), which is also toxic to the cell. Catalase, an enzyme, breaks
down hydrogen peroxide into water and oxygen, neutralizing the peroxide. Obligate anaerobes
lack these enzymes, which is why they cannot survive in the presence of oxygen.
Aerotolerant/fermenters such as organisms from the genus Streptococcus are catalase negative
yet survive just fine in the presence of oxygen. Streptococcus sp. produce a different enzyme
called peroxidase which neutralizes hydrogen peroxide.
The last condition for growth we will discuss is how pH affects growth. Organisms can
be classified based on their ability to tolerate different salt concentrations and acid/base levels.
These concepts are important for understanding how humans preserve food. Many bacteria
including human pathogens are considered neutrophiles and grow in a neutral pH around 6.67.9. Organisms growing in a pH above 8.0 would be considered Alkalophiles. Acidophiles can
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tolerate acidic conditions below pH of 6.5 and below (Table 5.1). Few organisms can grow
below a pH of 4.0, which is important in preserving foods. Sauerkraut, pickles, cheeses, canned
tomatoes are all preserved from spoilage by acids produced during a fermentation process or
natural low acidity.
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