Characteristics and study of prokaryotic growth

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Characteristics and study of prokaryotic
growth How do we grow bacteria in the
laboratory? What is required for growth? How do
we measure bacterial growth? What do bacteria do
while they are growing? How do bacteria grow in
the environment?
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What is a pure culture? All of the organisms are
descended from a single cell (and therefore are
identical) Bacteria do not grow that way
naturally Most bacteria cannot be cultured that
way So why do it?
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Historically (19th century) Kochs postulates
applied to prove that a particular
organism caused disease -isolate organism from
diseased animal -establish a pure culture of the
organism (and prove it) - infect another animal
with cultured bacteria does this animal get
sick? are the same microbes isolated from the
second animal? Koch proved that B.anthracis
causes anthrax and that M. tuberculosis causes
tuberculosis this way Koch also developed new
methods for bacterial growth
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What is required for pure culture? Sterile
apparatus Aseptic technique Appropriate
media solid media separable colonies
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Bacteria grow (i.e, divide) by binary
fission With each generation the population
doubles generation time, or doubling time,
varies with bacterium and growing
conditions Can be as little as 20 minutes Thus
bacterial contamination can be rapidly established
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What sorts of factors affect bacterial
growth? temperature
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How can they grow at such a wide range of
temperatures? Implications substances that
function at extreme temperatures for prevention
of contamination for pathogenesis (why do we
develop fevers?)
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Oxygen requirements
Availability varies in the environment and on/in
our bodies
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pH Bacteria maintain an internal neutral pH, but
may survive in a wide range of acidic or
alkaline environments How? H. pylori- produce
urease generates ammonia and raises pH of
immediate environment Other bacteria use proton
pumps acidophiles- out alkalophiles- in
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Salt tolerance Synthesize or pump solutes inside
the cell (and draw water) Osmotolerant
organism- can grow in dry environments (like
skin) Halophiles- require high salt
conditions High solutes concentrations are
historically used to preserve foods
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What do bacteria require to grow? Nutrients Energ
y source Some organisms require growth factors
(fastidious)
Which is easiest to grow in the laboratory?
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Some media are richer than others Some are used
to look at mixed cultures of microorganisms Selec
tive- allow some organisms to grow but not
others Differential- bacteria have different
characteristics Some are both!
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Adjusting atmospheric requirements CO2 levels
(capnophiles) Candle jars (microaerophiles) Anae
robe jars/chambers (guess) reducing agents
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Why would you use selective enrichment medium?
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Measuring bacterial growth Considerations Rapidi
ty Viable vs. total bacteria Large numbers or
small numbers of bacteria Growth in progress?
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Direct counting Rapid Can count organisms that
cant grow in medium Cant distinguish live from
dead cells Motile organisms are hard to
count! Special instruments needed
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Viable counts Of living (and growing)
bacteria requires some skill and time Can be
performed on special media Need fairly
concentrated culture Dilute samples membrane
filtration MPN (most probable number)-
statistical estimate, NOT direct count
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Measuring cell mass Most common method-
turbidity (spectrophotometry) Cells grown in
liquid medium Can measure progress of cell
growth Changes (in turbidity) correlate with
increase in cell numbers Growth can be
measured either by turbidity or by counting to
determine growth curve
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Metabolic activities change in late
log/stationary phase
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Metabolites are often of research/commercial
interest Bacteria can be grown to enhance
production
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How do bacteria actually grow in nature? Are
nutrients always available, and in constant
amounts? They may elaborate protective
structures when necessary glycocalyx endospores
biofilm (protects communities of
microbes) Interactions may be required (one
organism produces substances required by another