chapter 6s

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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