Factors Affecting Microbial

Growth
Binary fission in bacteria

Scanning electron
micrograph
 Geometric
progression in the
number of bacteria in
a population resulting
from binary fission

Generation time 
length of time it takes
a single bacterium to
double

E. coli 25 minutes
Mycobacterium spp. 1-3
days.
Binary fission requires the addition
of new material at the growing sites
of bacteria

Gram positive cells such as Gram negative cells such as

this coccus, new material this bacillus, new material is
is added at the division plane added at the division plane and
also throughout the length of
the cell

Bacteria need to synthesize the macromolecules that allow for their

growth and reproduction.
 Physical Requirements for
Growth
 Temperature: most organisms like 37oC
 Oxygen: diversity
 pH: optimum of most organisms is 7.0
 Osmolarity: cytoplasm osmolarity must be
greater than that of its environment
 Pressure
 Water activity: most bacteria require a Aw
between 0.9 and 1.0
 Oxygen Requirements
 Oxygen is essential for obligate aerobes
(final electron acceptor in ETC)
 Oxygen is deadly for obligate anaerobes
 How can this be true?
 Neither gaseous O2 nor oxygen covalently
bound in compounds is poisonous
 The forms of oxygen that are toxic are highly
reactive
 They are strong oxidizing agents. Resulting
chain of oxidations causes irreparable damage
to cells by oxidizing compounds such as
proteins and lipids
Classification of Organisms Based
on Oxygen Requirements
 Aerobes – undergo aerobic respiration
 Anaerobes – do not use aerobic metabolism
 Facultative anaerobes – can maintain life via
fermentation or anaerobic respiration or by
aerobic respiration (E. coli)
 Aerotolerant anaerobes – do not use aerobic
metabolism but have some enzymes that detoxify
oxygen’s poisonous forms (lactobacilli)
 Microaerophiles – aerobes that require oxygen
levels from 2-10% and have a limited ability to
detoxify hydrogen peroxide and superoxide
radicals (H. pylori)
 Obligate anaerobes—oxygen kills the organism
 Capnophiles
There are microorganisms that grow better at a
higher CO2 tension than is normally present in the
atmosphere.

Special CO2 incubators are used for cultivation of capnophile bacteria.

Medically important member of this grope is Streptococcus pneumonia

(agent of pneumonia), Neisseria (agents of gonorrhea and meningitis),
Brucella (agent of undulant fever).
 Four Toxic Forms of Oxygen
 Singlet oxygen – molecular oxygen with electrons
boosted to higher energy state
 Occurs during photosynthesis so phototropic microorganisms
have carotenoids that remove the excess energy of singlet
oxygen

 Superoxide radicals – some form during

incomplete reduction of oxygen in aerobic and
anaerobic respiration
 So reactive that aerobes produce superoxide dismutases to
detoxify them
2O2- + 2H+  H2O2 + O2
 Anaerobes lack superoxide dismutase and die as a result of
oxidizing reactions of superoxide radicals formed in presence of
oxygen
 Four Toxic Forms of Oxygen
 Peroxide anion – formed during reactions
catalyzed by superoxide dismutase and
other reactions
 Aerobes contain either catalase or
peroxidase to detoxify peroxide anion
2H2O2  2H2O + O2
 Obligate anaerobes either lack both enzymes or
have only a small amount of each
 Four Toxic Forms of Oxygen

 Hydroxyl radical – results from ionizing

radiation and from incomplete reduction of
hydrogen peroxide
 The most reactive of the four toxic forms of
oxygen
 Not a threat to aerobes due to action of
catalase and peroxidase
 Aerobes also use antioxidants such as
vitamins C and E to protect against toxic
oxygen products
 Enzyme content of bacteria with
different requirement for oxygen
Name Enzyme content for O2
detoxification
Strict aerobe Catalase – H2O2  H2O + O2
Superoxide dismutase
O2- +2H+ O2 + H2O2  H2O + O2
Facultative Catalase
anaerobe Superoxide dismutase
Microaerophile Small amount of catalase and superoxide
dismutase
Aerotolerant Superoxide dismutase
Strict anaerobe Neither catalase nor superoxide dismutase
 pH
 Acidophiles (grow at low pH (0-5,5)
 Alcaliphiles (8,5-11,5)
 Normal (6,5-7,2)
 pH
Effects of acids on organisms
 energy required to maintain cell's internal
pH
 enzyme activity affected
 proteins, DNA, other molecules denatured
 longer lag, less rapid growth
 Temperature

 Effect of temperature on proteins

 Effect of temperature on lipid-containing
membranes of cells and organelles
 If too low, membranes become rigid and
fragile
 If too high, membranes become too fluid and
cannot contain the cell or organelle
 Temperature (characteristic
ranges)
 Psychrophiles:
 Psychrophilic
Opt temp below 15oC
Capable of growth at 0oC
Cannot grow above 20oC
Found: SNOW FIELDS, POLAR ICE, DEEP OCEAN
EXAMPLES: Pseudomonas, Flavobacterium, &
Alcaligenes
Achromobacter sp.
 FACULTATIVE PSYCHROPHILE:
Grow slowly in cold conditions
BUT have opt temp above 20oC
CONCERN: Staphylococcus
EXAMPLES: Contaminants of aureus, L. products
food/dairy
 Temperature

Mesophiles: between 15 and 45 with

optimum around 37 C

Thermophiles: between 30 and 75 with

optimum around 55 C

Hyperthermophiles: T greater than 100 C

Effects of Temperature on
Growth

Figure 6.4a
Effects of Temperature on
Growth

Figure 6.4b
Food Preservation Temperatures
TEMPERATURE RANGES (OPTIMA GROWTH) OF SOME
BACTERIA

Bacterium Growth Temperature oC

Min Max Optimum
1. Pseudomonas fluorescens 2-4 36-38 25-30
2. Pseu. aeruginosa 10-15 41-44 c. 37
3. Escherichia coli 15-20 45 37
4. Bacillus polymyxa 5-10 35-45 30-32
5. B. stearothermophilus 30-45 65-75 c. 55
6. Thermus sp. 40 79
 WATER ACTIVITY (Aw)

ALL BACTERIA require water (growth & reproduction)

Essential solvent, biochemical reactions

Water activity = index amount of water free to react

Equivalent to atmospheric measure (Relative Humidity)
Absorption & Solution factors reduce availability ( Aw)
Pure distilled water (Aw =1)

E.g., Saturated soln NaCl (Aw = 0.8)

Seawater [NaCl] 3% (Aw = 0.98)

RH = 100 Aw Therefore, 90% RH = 0.90 Aw

Most bacteria Aw >0.9 (active metabolism)

Most microbes - grow opt Aw = 1.0

 Aw = slow growth rate
Below Aw 0.9 Bacteria unable to grow

EXCEPTIONS:
XEROTOLERANT: lower Aw
Fungi able to grow Aw 0,7 osmotolerant 0.60
Yeasts (conc sugar soln’s Aw = 0.60)
Salt-tolerant Bacteria - Halophiles (High [Solute], low Aw)
Effect of Aw on growth of Staphylococcus aureus in
medium containing hydrolysate
The Interrelationships of Aw of various foods &
susceptibility to microbial spoilage
 OSMOTIC PRESSURE

Results from: water diffusing across cell membrane in

response to [solute]
Association with [salt] = SALINITY

OSMOTOLERANT: withstand high osmotic pressure

high [solute]

OSMOPHILES: require high [solute] for growth

E.g., Xeromyces (opt Aw =0.9)
SALINITY:
Most microbes - HYPOTONIC or ISOTONIC conditions

HALOPHILES (exceptions) - Require High [NaCl]

Moderate Halophiles: Marine bacteria 3% [salt]
Require 1.5% NaCl (maintain membrane integrity)

Extreme Halophiles: saturated brine soln’s

OBLIGATE HALOPHILES Halobacterium, sp
Halococcus
Both 25% NaCl
Drying, Salting, Jamming (achieve low Aw)
EFFECTIVE methods of preservation
HYDROSTATIC PRESSURE

Normal pressure: 1 atmosphere (atm)

Ocean Depth (1000m +) HP = 600-1100 atm (2-3oC)
Bacteria survive & adapt BAROPHILES

BAROTOLERANT:  pressure not adversely affected

Industrial case: P> 1,5 atm affects metabolism

Nutritional requirements
 Bacteria need to synthesize
macromolecules that allow for their
growth and reproduction— what are bacterial
cells made up of ?

Cells consist of WATER and MACROMOLECULES

Macromolecules are made up of smaller monomeric molecules

Small monomeric molecules are made up of atoms

 Macromolecules of the bacterial cell
Proteins—The most abundant class of macromolecules and comprise
most of the structures of the cell as well as enzymes
amino acids—monomeric subunits of proteins
consists of carbon, hydrogen, oxygen, nitrogen, sulfur and
sometimes selenium
amino acids are covalently linked to form a peptide bond
H H H
H
NH3 C COOH C N C COOH
NH3 C
R R1 O R2

Peptide bond
 Macromolecules of the bacterial cell

Polysaccharides—2nd most abundant of the bacteral macromolecules

sugars (monosaccharides)—monomeric units
consists of Carbon, Hydrogen and Oxygen atoms at a ratio
of 1:2:1
individual sugars are linked by a glycosidic bond

Polysaccharides form covalent linkages with other macromolecules

with proteins—glycoproteins
with lipids—glycolipids, lipopolysaccharides
 Macromolecules of the bacterial cell

Nucleic Acids
DNA—deoxyribonucleic acid
RNA—ribonucleic acid
Backbone of nucleic acids= polymer of phospho-ribose (RNA)
or phospho-deoxyribose (DNA)
The sugars are covalently attached to each other by phospho-
diester bonds
Bases are attached to a carbon atom of the sugar moiety
cytosine, adenine, guanine (DNA/RNA); thymine (DNA)
uracil (RNA)
Comprised of the atoms Phosphorous, Carbon, Hydrogen, Oxygen
and Nitrogen
 Macromolecules of the bacterial cell

Lipids—made up of a long carbon chain—fatty acids—(14-20

carbons and one carboxylic acid group)

Saturated—Hydrogen atoms attached to most or all carbon

moieties.
Unsaturated—fewer Hydrogen atoms associated with carbons

Complex lipids are attached to simple sugars like phoshoglycerol

(ie phospholipids) or complex polysaccharides (LPS)

C, H, O, P
 Macromolecules (Stouthamer, 1973)

Organic compounds % dry weight

Proteins 52,4
Carbohydrates 16,6
Lipids 3,4
RNA 15,4
DNA 3,2
Others 2,7
 Bacterial Nutritional
Requirements
NUTRITION::act of supplying microorganisms with the molecules
and atoms they require for the biosynthesis of small molecules
and macromolecules
Macronutrients: nutrients required in high amounts
Carbon, Nitrogen, Phosphorous, Sulfur
ALSO
Potassium, Magnesium, Calcium, Sodium
Micronutrients: nutrients required in small or even trace amounts
Chromium, Cobalt, Copper, Manganese, Molybdenum
Nickel, Selenium, Tungsten, Vanadium, Zinc, Iron
Growth Factors::organic compounds required in very small amounts

Vitamins, amino acids, purines and pyrimidines

 Bacterial Nutritional
Requirements
Autotroph: an organism capable of biosynthesizing all cell
material from CO2 as a sole carbon source
Heterotroph: an organism that requires carbon from preexisting
organic material
Photo--: solar energy converted to chemical energy
Chemo--: energy derived from chemical compounds
Catabolism: Act of breaking down complex molecular material
for energy or biosynthetic material
Anabolism: The act of biosynthesizing complex material from simpler
organic compounds
Culture media for artificial cultivation of bacteria in the lab
1. Complex (undefined)—enzymatic digests of milk protein
(casein), beef, yeast
2. Chemically defined—precise amounts of purified organic
and inorganic compounds are added to distilled water
 Macronutrients
 Carbon (CO2 or organic compounds)
 Hydrogen (H2O or organic compounds)
 Oxygen (H2O or organic compounds)
Nitrogen (NH3, NO3-, organic N-compounds)


 Phosphorus (PO43-)
 Sulfur (H2S, SO42-, organic compounds)
 Potassium (K+)
 Magnesium (Mg2+, salts)
 Sodium (Na+)
 Calcium (Ca2+, salts)
 Iron (Fe3+, Fe2+, or salts)
 Iron as a nutrient
 Needed for aerobic metabolism
(cytochromes, iron-sulfur proteins)
 Insoluble under aerobic conditions
 Fe(OH)3, FeOOH
 Solubilized by siderophores
Siderophore
 Micronutrients and growth
factors
 Micronutrients: Metals and metalloids
 Generally not necessary to add to medium
 Deficiencies can arise when medium
constituents are very pure
 Growth factors: organic requirements
 Vitamins, amino acids, purines, pyrimidines,
acetate
 Growth Requirements

 Organisms use a variety of nutrients for

their energy needs and to build organic
molecules and cellular structures
 Most common nutrients – those containing
necessary elements such as carbon,
oxygen, nitrogen, and hydrogen
 Microbes obtain nutrients from variety of
sources
 Nutrients: Chemical and Energy
Requirements

 Sources of carbon, energy, and electrons

 Oxygen requirements
 Nitrogen requirements
 Other chemical requirements
 Sources of Carbon, Energy, and
Electrons

 Organisms are categorized into two groups

based on source of carbon
 Those using an inorganic source of carbon
(carbon dioxide) are autotrophs
 Those catabolizing reduced organic molecules
(proteins, carbohydrates, amino acids, and fatty
acids) are heterotrophs
 Sources of Carbon, Energy, and
Electrons (continued)

 Organisms categorized into two groups

based on whether they use chemicals or
light as source of energy
 Those that acquire energy from redox reactions
involving inorganic and organic chemicals are
chemotrophs
 Those that use light as their energy source are
phototrophs
Four Basic Groups of Organisms

Figure 6.1
 Nitrogen Requirements
 Growth limiting nutrient for many
organisms: Anabolism often ceases due to
insufficient nitrogen needed for proteins
and nucleotides
 Nitrogen acquired from organic and
inorganic nutrients; also, all cells recycle
nitrogen from amino acids and nucleotides
 The reduction of nitrogen gas to ammonia
(nitrogen fixation) by certain bacteria is
essential to life on Earth because nitrogen is
made available in a usable form
 Other Chemical Requirements
 Phosphorus required for phospholipid
membranes, DNA, RNA, ATP, and some proteins
 Sulfur is a component of sulfur-containing amino
acids, disulfide bonds critical to tertiary structure
of proteins, and in vitamins (thiamin and biotin)
 Trace elements – usually found in sufficient
quantities in tap water (Se, Co, Cu, Mn, Si, Zn)
 Growth factors – organic chemicals that cannot
be synthesized by certain organisms (vitamins,
certain amino acids, purines, pyrimidines,
cholesterol, NADH, and heme)
 Principal elements of the cell and
their physiological functions
Element Cell dry Physiological functions
weight
Carbon (C) 50 Constituent of all organic cell components
Oxygen (O) 20 Constituent of cellular water and most organic cell
components; molecular oxygen serves as an
electron acceptor in aerobic respiration
Nitrogen (N) 14 Constituent of proteins, nucleic acids, coenzymes
Hydrogen (H) 8 Constituent of cellular water and organic cell
components
Phosphorus 3 Constituent of nucleic acids, phospholipids,
(P) coenzymes
Sulfur (S) 1 Constituent of some amino acids in proteins and
some coenzymes
Potassium (K) 1 Important inorganic cation and cofactor for some
enzymatic reactions. It stabilizes ribosomes
 Principal elements of the cell and their
physiological functions
Element Cell dry Physiological functions
weight
Sodium (Na) 1 One of the principal inorganic cations in
eukaryotic cells and important in membrane
transport
Calcium (Ca) 0,5 Important inorganic cation and cofactor for
some enzymatic reactions
Magnesium (Mg) 0,5 Important inorganic cation and cofactor for
many enzymatic reactions
Chlorine (Cl) 0,5 Important inorganic cation
Iron (Fe) 0,2 Constituent of cytochromes and some
proteins
Trace elements: 0,3 The elements are required in very small
Cobalt, Zinc, amounts.
Molybdenum, Part of enzymes, required for enzyme
Manganese (Mn) activity