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Chapter 6 – Microbial
Growth and Death
• Batch growth patterns
• Batch growth kinetics
• Environmental conditions of kinetics
• Quantifying cell concentration
• Death kinetics
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• Growth → replication + Δ cell size
• Nutrients → energy production +
biosynthesis + product formation
• Substrates + cells → Productsout + “cells”
ΣS + X → ΣP + nX
• Autocatalytic reaction
6.2
Microbial growth
Section 6.1 of textbook
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6.3
Microbial growth patterns
A. Growth-associated product formation
Section 6.2.2 of textbook
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2.4
B. Non-growth-associated
product formation
Penicillin production on glucose
by Penicillium chrysogenum
S
X
P
Section 6.2.2 of textbook
O2 CO2
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6.5.5
Diauxic growth of Pseudomonas oxalaticus (●) on oxalate (▲) while adapting to acetate (■)
C. Diauxic growth
Section 6.2.2 of textbook
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A: Regular, linear graph
B: Semi-logarithmic graph
Exponential phase
Microbial growth curve
Section 6.2.2 of textbook
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Growth rate =dt
X d Slope
phaselExponentiaconstant
dt
X d
X
dt
X Lnd
dt
X Log d Slope
1
)(
3.2
1
Specific
growth rate =
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2.8
Microbial growth kinetics
• First-order kinetics for exponential growthrate and death rate, µ = (1/X) dX/dt
• Yields
growth YX/S = - ΔX/ ΔS
product YP/S = - ΔP/ ΔS
Section 6.2.2 of textbook
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2.9
Microbial growth kinetics vs
environmental conditionsSection 6.2.3 of textbook
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2.10
Microbial count – direct
methods number of cells
Petroff-Hausser (hemocytometer) slide
Plate count giving CFU
“Particle” counter – flow cytometer
Section 6.2.1.2 of textbook
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2.11
Microbial count – direct
methods cell massconcentration
Dry weight
Packed cell volume
Absorbance
Section 6.2.1.2 of textbook
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2.12
Microbial count – indirect
methods cell massconcentration
Protein measurement after extraction ATP measurement with luciferin-luciferase
Section 6.2.1.2 of textbook
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2.13
Microbial death kinetics
Food sterilization, canning, retort
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Death rates – D-value
Decimal Reduction Time
N
time
T
o
o Dk N
N Log 303.21.0
T
T
o t D N
N Log
1
T o t
k
N
N Log
303.2
For a population of micro-
organisms, N, being killed:
When 90% destruction, tT = DT
No
0.1No
11.0
o
o
N N Log
T givenaat N k dt
N d
DT
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Death rates – z-value
Thermal Destruction Temperature
time
temperature
For a given No/N, less time
is required to kill at higher T
21
21
T T
t Log t Log
Slope
T T
tT2
T2
tT1
T1
For 90% reduction in heating
time, T2 – T1 = z:
121 T T t Log t Log
z T T
t Log t Log T T
12
21 1
z Slope
1
z
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Microorganism Tref. (
C) DT, ref.
(sec) zT, ref.
(
C) Typical Food
Pasteurization
Mycobacterium tuberculosis 82.2 0.018 5.6 Milk
Coxiella burnetii 65.0 36 5.0 Milk
Salmonella spp. 82.2 0.192 6.7 Milk
Staphylococcus spp. 82.2 0.378 6.7 Milk
Coxiella burnetti 82.2 0.0131 5.0 Milk
Lactobacillus spp. 82.2 0.57 6.7 Milk
Sterilization (“Appertization”)
Clostridium botulinum 121.1 12.6 10.0 Milk
Clostridium sporogenes 121.1 6 – 9.0 10.0 Milk
Mesophiles 121.1 11 10.5 Whole milk
Mesophiles 121.1 31 10.5 Cream (30% fat)
Thermophiles 121.1 25 10.5 Whole milk
Thermophiles 121.1 46 10.0 Cream (30% fat)
Heat Resistance of
Microorganisms
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Heat Resistance of
Microorganisms
Microorganism Tref.
(
C)
DT, ref.
(sec)
zT, ref.
(
C)
Typical Food
Sterilization – low acid foods
Thermophi l ic (35 – 55°C)
Bacillus stearothermophilus 121.1 4.0 10.0 Vegetables, milk
Clostridium thermosaccharolyticum 121.1 3.0-4.0 7.2-10.0 Vegetables
Mesophi l ic (10 – 40°C)
Clostridium sporogenes 121.1 0.8-1.5 8.8-11.1 Meats
Bacillus subtilis 121.1 0.5-0.76 4.1-7.2 Milk products
C. botulinum toxins A and B 121.1 0.1-1.3 5.5 Low-acid foods
Listeria monocytogenes 18-42 (70°C) 5.6-5.7 Salmon, cod
Escherichia coli H157:O7 70-152
(68°C)
Pepperoni
Psychrophi l ic (-5 – 1.5°C)
C. botulinum toxin E 121.1 3.0 (60°C) 10.0 Low-acid foods
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Death rates – P-value
Pasteurization value
• Amount of time at a given temperature required
to denature enzymes or destroys non-pathogenic,
vegetative microorganisms
T
T
o t D N
N Log 1Replace tT by P, and DT by Dreference in:
and re-arrange
N
N
Log D P
o
reference
Reference temperature is normally 65ºC - 82ºC, for pasteurization
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• An alternate expression is more common:
z T T
t Log t Log T T 1
21
21
z
T T
T
z
T T
T
referenceT
ref
ref
P t
P
t
T T z
P Log t Log
1
1
10
10
(1
1
1
11
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Death rates – F-value
Sterilizing value
• Amount of time at a given temperature of
121.1ºC to destroys pathogenic microorganisms
• Expressions similar to those of pasteurization
can be derived:
z
T T
T
z
T T
T
referenceT
ref
ref
F t
F
t
T T z
F Log t Log
1
1
10
10
(1
1
1
11
where Treference is now 121.1ºC
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Death rates – L-value
Lethal value
• Amount of time calculated from autoclaveexperiments, equivalent to an F-value(theoretical, or reference value) for killing a
given microorganism• Sum of all individual times spent at a given
temperature during heating and some partof cooling
• For adequate sterilization: L > F
z
T T
T i
ref
t L
1
1011
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Heat Penetration in Cans during
Sterilization
2.22
T1 = final autoclave T
Autoclave T
Inside Can T
To = initial autoclave and/or can T
Tc = final autoclave and/or can T
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Practice Problems
• Calculate the D-value for Mycobacterium
tuberculosis at 60ºC using values in Table 1.1
• Calculate the pasteurization value at 82.2ºCbased on reducing the population of M.
tuberculosis by a factor of 9
• Calculate how much time one should pasteurize
M. tuberculosis at 71ºC