12/11/2021

40307 Microbiologia | 2021-22

Aula 4
Crescimento, cultivo e controle de microrganismos

Divisão celular e crescimento: quantificação do crescimento; curvas de

crescimento; culturas contínuas.
Fatores que afetam o crescimento microbiano.
Controle do crescimento.

Brock Biology of Microorganisms, 15th Ed. Chapter 5

https://pubhtml5.com/fkzf/nqar

Cell division and growth

• Generation time:
time required for
microbial cells to
double in number

2
40307 Microbiologia

1
 12/11/2021

Growth
• Growth: increase in the number of cells
• Binary fission: cell division following
enlargement of a cell to twice its minimum
size
• During cell division, each daughter cell
receives a chromosome and sufficient copies
of all other cell constituents to exist as an
independent cell.

3
40307 Microbiologia

Generation time
• Generation time: Time
span between two
consecutive cell
divisions

• Septum: inwards
growth of the
cytoplasmic membrane
Time
and cell wall

4
40307 Microbiologia

2
 12/11/2021

Binary fission

5
40307 Microbiologia

Binary fission and

other forms of division
Intercalar growth:
• new cell wall
material is produced
throughout the cell

Polar growth:
• new cell wall
material is
produced at a
single point
• cytoplamic
structures are not
partitioned

6
40307 Microbiologia

3
 12/11/2021

Binary fission and

other forms of division

Salmonella Pirellula Hyphomicrobium

Caulobacter Rhodopseudomonas Ancalomicrobium

7
40307 Microbiologia

The mathematics of
exponential growth
Exponential growth: cell numbers
double within a specific time interval

Nt = N02n
Nt is the cell number in a given moment t.
N0 is the initial cell number.
n is the number of generations during the period of
exponential growth.

8
40307 Microbiologia

4
 12/11/2021

Exponential growth parameters

generation time (g): g=t/n

number of generations (n):

instantaneous growth rate constant

(k) calculated from the generation
time (g):
Nt = N02n
K=ln2/g
Nt is the cell number in a given moment t.
N0 is the initial cell number. K=0.693/g
n is the number of generations during the period of
exponential growth.

9
40307 Microbiologia

The meaning of K - specific growth rate

µ - grams of biomass produced per gram of biomass present per hour of process time

instantaneous growth rate constant

(k) calculated from the generation
time (g):

K=ln2/g

K=0.693/g instantaneous growth rate constant

(k) calculated from the generation
time (g):

K=ln2/g

K=0.693/g

5
 12/11/2021

Growth curve

11
40307 Microbiologia

Growth curve

12
40307 Microbiologia

6
 12/11/2021

Growth curve

µ=0
µmax

µ<µmax
13
40307 Microbiologia

Diauxic growth

40307 Microbiologia
Growth supported by 2 carbon sources 14

7
 12/11/2021

Steady-state growth in
continuous cultures
• Continuous culture
– an open-system microbial culture of fixed volume
• Steady state
– cell density and substrate concentration do not
change over time
• Growth rate controlled by dilution rate
– maintain exponential growth phase for
weeks/months

15
40307 Microbiologia

Chemostat cultures

16
40307 Microbiologia

8
 12/11/2021

Chemostat cultures
• Growth rate and population density
of culture can be controlled
independently and simultaneously,
depending on Dilution Rate (D) and
Concentration of a Limiting Nutrient
(C).
• Dilution rate: F/V
– F is flow rate of adding fresh medium and
removing spent medium
– V is culture volume
• Concentration of a limiting nutrient
– N or C source in the culture
medium

17
40307 Microbiologia

Culturing requires inoculation and

suitable media
• Classification of culture media
– Composition
– Consistency
– Effect

• Cultivation
– In solid media
– In liquid media

18
40307 Microbiologia

9
 12/11/2021

Measuring growth
• Cell counts

19
40307 Microbiologia

Measuring growth
• Colony (viable) counts

20
40307 Microbiologia

10
 12/11/2021

Measuring growth
• Turbidimetry

21
40307 Microbiologia

Environmental factors affecting growth

• Optimum
– The value for an environmental factor that corresponds to the shortest
doubling time and the highest specific growth rate

Sourdough yeast: growth rates at various temperature 22

40307 Microbiologia

11
 12/11/2021

Temperature
Cardinal temperatures

23
40307 Microbiologia

Temperature

Temperature classes of microorganisms

24
40307 Microbiologia

12
 12/11/2021

Temperature
Psycrophiles
• Cytoplasmic membranes
– high unsaturated and shorter-chain fatty
acid content
– some polyunsaturated fatty acids, which
remain flexible at very low temperatures
• Cold shock proteins (chaperones)
• Cryoprotectants
• Exopolysaccharide cell surface slime
• Enzymes that function optimally in
the cold
– more α-helices than β-sheets: greater
flexibility for catalysis at cold temperatures Polaromonas
– more polar and fewer hydrophobic amino and diatoms in
acids
frozen seawater
– fewer weak bonds (e.g., hydrogen and
ionic bonds)

40307 Microbiologia 25

Temperature
Thermophiles
• Membrane stability
– modifications in cytoplasmic membranes
to ensure heat stability
– Bacteria have lipids rich in long-chain and
saturated fatty acids, fewer unsaturated
fatty acids.
– Archaea have C40 hydrocarbons and
membrane forms lipid monolayer rather
than bilayer
• Enzymes and proteins
– Critical amino acid substitutions provide
more heat-tolerant folds.
– Increased number of ionic bonds between
basic and acidic amino acids resists
unfolding in the aqueous cytoplasm.
• Production of solutes
– e.g., di-inositol phosphate, diglycerol
phosphate) helps stabilize proteins.

Archaea in boiling water

40307 Microbiologia 26

13
 12/11/2021

pH

Acidophiles: Alkaliphiles:
• stability of cytoplasmic membrane and • sodium (Na+) motive force rather than
ion pumps proton motive force
40307 Microbiologia
• enzymes 27

Osmolarity and aw
Halophiles Seawater
Osmophiles aw = 0.98
Xerophiles

Hyper-arid
hot desert
soil
aw = 0.1-0.6

28
40307 Microbiologia

14
 12/11/2021

Osmolarity and aw
Accumulation of compatible solutes

29
40307 Microbiologia

Oxygen

30
40307 Microbiologia

15
 12/11/2021

Oxygen
a) Obligate aerobes
b) Anaerobes
c) Facultative aerobes
d) Microaerophiles
e) Aerotolerant

31
40307 Microbiologia

Controlling growth
• Control by heat
– Autoclave
– Pasteurization
– Tindalization
– Appertization
• Radiation
• Filtration
• Chemical control
– Target
• Sterilizers
• Disinfectants
• Sanitizers
– Effect
• Bacteriostatic
• Bactericidal
• Bacteriolytic
32
40307 Microbiologia

16