MGY277H1 Lecture Notes - Lecture 5: Crystal Violet, Archaea, Spectrophotometry

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Lecture 5 Microbial Growths
Principles of Microbial Growth
Prokaryotic cell divide by the process Binary Fission
Exponential growth population doubles each division
oCome with important consequences
o10 cell of food born pathogen in potato salad at picnic
can become 40,000 cells in 4 hours
Microbial/Prokaryotic growth = increase in cell numbers,
NOT size
Generation time time it takes to double population
oVaries among species and influenced by environmental conditions
(temperature and nutrient availability)
oDepends on species and growth condition
A single E.coli cell, through binary fission, can grow into a population of cells with
the mass of the entire Earth within 43 hours!!!
Principles and Formula of Prokaryotic
Growth
Growth can be calculated Nt=N0 X 2n
Nt = # of cells in population at time t
N0 = original # of cells in population
N=number of division
Most bacteria will NOT grow at low temperature (4C)
Fast growth is NOT required for a microbe to become pathogen; they can grow for
hours and days
Prokaryotic growth in Laboratory Condition
Prokaryotes grown in culture medium
Laboratory Growth Condition System
Closed
Batch cultures
Open
Continuous Culture
Nutrition NOT renewed Nutrition renewed
Waste NOT removed Waste Removed
Characteristic growth
curve
Much more complicated
process (Use a pump)
Shaking for oxygenation Use a chemostat
machine as monitoring
(Usually grow in liquid with ASEPTIC technique)
oTubes or Flask or both Liquid
oAgar plates (Petri dishes) Solid
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Characteristic growth curve of a CLOSED SYSTEM (5 stages)
1. Lag phase
No growth
Begin synthesizing enzymes required for growth
Delay depends on conditions
2. Log /Exponential phase
Phase used to measure generation growth
Cell divide at constant rate
Generation time can be measures
Medically & Commercially Important
oMedical Most sensitive/vulnerable to antibiotics
oCommercial Production of primary metabolites
Primary metabolites made during growth phase (lipids, nucleotide)
Later stage Nutrient gradually deplete, waste accumulate
oCell activate endospore formation when possible and prepare for
starvation
oMaking secondary metabolites
Secondary metabolites NOT used for growth (toxin, antibiotics)
3. Stationary phase
Not died but No division (Total # stay the samewhen other dies it
release nutrient for other to grow)
Nutrient levels too low to sustain growth (Some may experience late log)
Duration of this phase depends on environmental condition and species
4. Death phase
Total # of viable cells decrease (cell die at constant rateExponentials
but much slower than growth rate)
When cell die they become food for other bacteria at stationary phase
which is WHY bacteria take a long time to die
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5. Phase of prolonged decline
Some survive death phase as stated below
Us nutrient from dead cells
Survivor can persist but most die off
Since closed medium change the environment, if you want
to have a consistent environment you need to use…
CONTINUOUS GROWTH / OPEN SYSTEM
Prolonged phase of exponential growth
Chemostats can maintain continuous growth
oContinuously drip fresh medium into culture in chamber
oRemove equivalent waste (cells, spent medium)
oSpeed of addition & removal is controlled (constant rate and cell
density)
oProduce relatively uniform population for study
How do you culture individual microbes from complex microbial samples?
Obtaining a pure culture
Pure culture population of cells derived from a single cell
oAllow study of single species
Limitation
oOften behave differently than in natural environments
oApprox. 1% of prokaryotes can be grow in lab, BUT…
oRecent advances including genomics have enable us to cultivate far
more microbes than before (about 90% associated with human can
be cultures)
Most medically relevant bacteria and human associated microbes can be
grown in pure culture
Pure culture require ASEPTIC techniques to minimize contamination
Up till now we talk about growing bacteria in liquid
NOW we are going to talk about solid media
Growth on Solid Media
Require culture medium, container, aseptic condition, method d to separate
individual cells
With correct conditionsingle cell will multiply
Usually form visible colony (~1 million cells is easily visible)
Agar used to solidify culture media (Hold in Petri Dish)
oFew microbes can degrade
oNot destroyed by high temp
oLiquefies above 95 C
oSolidifies below 45 C
Solid media CAN separate microbes in complex samples
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