MICROBIOLOGY

DHV 20043
8 Unit Theories (55% T1, T2 & TF)
6 Practical (35% Skills & Report)
1 Task (10% Affective Domain from Report)

Tuesday (8.30am-9.30am)
Wednesday (2.00-5.00pm)

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Contents
• Unit 1: Introduction to microbial cell biology
• Unit 2: Classification of microorganisms
• Unit 3: Basic microbiological techniques (√)
• Unit 4: Interactions & impact of microorganisms & humans
• Unit 5: Correct use of standard microbiology lab equipment (√)
• Unit 6: Proper use of aseptic techniques & demo of appropriate
microbiological media & test system (√)
• Unit 7: Estimate the number of microbes (bacterial counts) (√)
• Unit 8: Antimicrobial sensitivity testing (√)
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Contents
Lecture Practical Assessment
Unit Title TT FT P Affective
(hours) (35%) (20%) (35%) (10%)
1 Introduction to microbial cell biology 8 - / /
2 Classification of microorganisms 8 - / /
3 Basic microbiological techniques 12 6 / / / /
4 Interactions and impact of microorganisms / /
4 -
and humans
5 Correct use of standard microbiology /
4 3
laboratory equipment
6 Proper use of aseptic techniques & demo of /
appropriate microbiological media & test 5 3
system /
7 Estimate the number of microbes (bacterial
5 3 /
counts)
8 Antimicrobial sensitivity testing 5 4 /
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Overview on Microbiology

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 Do you know that bacteria can heal crack concrete?

Bacteria that produce spore that are tough to harsh condition (Bacillus pseudofirmus)
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Unit 1
Introduction to microbial cell biology

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Introduction to microbial cell biology
1.1 Information flow within a cell
1.2 Regulation of cellular activities
1.3 Basic cell structure and function
1.4 Growth & cell division
1.5 Microbial Metabolism
1.6 Bacterial genetics

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1.1 Information flow within a cell
Topic learning outcome:

- Describe the process involved in Central

Dogma of molecular biology.

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Central Dogma of Biology

Ribozyme

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1.2 Regulation of Cellular Activities
Topic learning outcome:

- Describe the genetic control mechanisms.

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Overview: Cellular Regulation
• Bacterial cell carries out large numbers of metabolic reactions
(anabolic & catabolic: release & require energy).
• All metabolic reactions are catalyzed by enzymes.

• A cell activity can be stop or continue through the feedback inhibition,

repression or induction of the enzymes that catalyzed the process.
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Overview: Cellular Regulation
• Genes, through transcription & translation, direct the synthesis of
proteins, (gene expression) many of which serve as enzymes used for
cellular metabolism.
• Tremendous energy is required for protein synthesis. Therefore, to
conserve (save) energy, cell only produce needed protein only. Thus,
controlling the synthesis of enzymes.
• Gene regulation describe any mechanism used by a cell to increase or
decrease the production of specific gene products (protein or RNA) by
modifying the gene expression.
• Mostly occur at the levels of transcription.

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Repression & Induction
Control the formation & amounts of enzymes in the cells, not the activities of the
enzymes.
Repression Induction
• What: Inhibit gene expression • What: Stimulate gene
& decrease the synthesis of expression & increase the
enzymes. synthesis of enzymes.
• Why: Due to overabundance • Why: For utilization of lactose
of end-product of metabolic
reactions. (food for E. coli)

• How: Repressor block RNA • How: Inducer turns on

polymerase that initiate transcription of a gene (lac
transcription. operon genes)
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1.3 Basic Cell Structure & Function
Topic learning outcome:

- Compare and contrast the overall cell structure

of prokaryotes and eukaryotes

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BACTERIA VIRUS

MICROORGANISMS

FUNGI
ALGAE

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Comparing prokaryote & eukaryote: An Overview
• Chemically similar
(contain nucleic acids,
proteins, lipids &
carbohydrate)
• Same chemical reactions
(to metabolize food, build
proteins, & store energy)
• Differences: size,
structure of cell wall,
membranes, the presence
of organelles, cell division.

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1.4 Growth & Cell Division
Topic learning outcomes:
- Explain the requirements for microbial
growth.
- Describe bacterial growth

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Bacterial Growth: Overview
• Increase in
bacterial number,
not in size.
• As number of
bacteria increase,
it forms into
colonies.
• Why do we study
bacterial growth?

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Water in the deep sea are not boiled even at
temperature above 100○C?

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Bacterial Growth: Requirement
Physical Chemical

• Temperature • Carbon
• pH • Nitrogen, Sulphur,
Phosphorus
• Osmotic pressure
• Trace elements
• Oxygen
• Organic growth factors

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 The red, orange, yellow, and green are all due to
microbial growth – the temperature gradient by the edge
of the hot spring created a unique niche for different
microbes that can survive in that specific environment

National Yellowstone Park, US Grand Prismatic Spring

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Physical Requirement: Temperature
A. Classification:
i. Psychrophiles :
 Cold-loving microbes
 Seldom cause disease or food microbes
Location: North pole/ ocean depth

ii. Mesophiles :
Moderate-temperature-loving microbes
Include most pathogens and common spoilage
organisms.
Location: Many have adapted in the body.

iii. Thermophiles :
Heat-loving microbes
Location: Adapted to live in sunlit soil, Range: 30°C apart
compost piles, and hot springs
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Physical Requirement: Temperature
B. Min. optimum & max. growth
temperature range:
i. Min. growth temperature
 Lowest temperature at which the
species will grow

ii. Optimum growth temperature

 Temperature at which the species
grows best.

iii. Max. growth temperature

 Highest temperature at which growth
is possible.

Range: 30°C apart

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 Why do you think so?
• The bacteria grow in the soil and sporulates upon
unfavorable conditions.
• The spores are transferred in the soil deposited on
the plants. (1)
• If the canning temperatures are sufficient then the
bacteria will die. If not, the bacteria will begin to
grow under the anaerobic conditions inside the
can/jar.
• The nutrients are eventually exhausted and the
bacteria die, leaving the toxin. Home-canned foods
are most often a source because the canning
temperatures are less rigorously monitored.
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Size of the container use to keep
food might influence the chance
of food spoilage?

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Food Preservation Temperature

 Refrigeration is the most

common method of
preserving food
 Growth is not fully inhibited
by the temps of the
refrigerator.
 Growth is slow
 Results in mold and slime on
food surfaces, off-tastes, off-
colors, and odors (gases being
produced by the microbe)

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Physical Requirement: pH
i. Most bacteria prefer neutral pH
(6.5-7.5).
ii. Molds and yeast grow in wider
pH range, but prefer pH between
5 and 6.
iii. Alkalinity & acidity inhibits
microbial growth.
 Which pH is preferable for food
preservation?

When bacteria are cultured in the laboratory, they often produce acids that
eventually interfere with their own growth. To neutralize the acids and
maintain the proper pH, chemical buffers are included in the growth
medium.

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Food Preservation: Application

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Physical Requirement: pH
Classification
• A. Acidophiles: “Acid loving”.
– Grow at very low pH (0.1 to 5.4)
– Lactobacillus produces lactic acid, tolerates mild acidity.

• B. Neutrophiles:
– Grow at pH 5.4 to 8.5.
– Includes most human pathogens.

• C. Alkalophiles: “Alkali loving”.

– Grow at alkaline or high pH (7 to 12 or higher)
– Vibrio cholerae and Alkaligenes faecalis optimal pH 9.
– Soil bacterium Agrobacterium grows at pH 12.
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Physical Requirement: Osmotic pressure
• Microbes require water for
growth, and their composition
is 80-90% water.
• In a hypertonic solution, water
passes out the cell through the
plasma membrane, causing
plasmolysis (osmotic loss of
water in which the cell’s
cytoplasm shrink)

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Food Preservation: Application

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 Physical Requirement: Osmotic pressure
Halophiles: Require moderate to large salt
concentrations.
High salinity combined with the presence of a salt- • Ocean water contains 3.5% salt.
loving algae species known as Dunaliella salina and
pink bacteria known as halobacteria. • Most bacteria in oceans.

Extreme or Obligate Halophiles: Require very

high salt concentrations (20 to 30%).
• Bacteria in Dead Sea.

Facultative Halophiles: Do not require high salt

concentrations for growth, but tolerate 2% salt or
more. 33
Chemical Requirement: Overview
• Microbes require energy and nutrients to build protein &
structural membranes and to drive biochemical processes.
• Water, carbon, nitrogen, sulphur and phosphorus are
needed in large quantities for microbial growth.

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 Chemical Requirement

Carbon Nitrogen, Trace Element Oxygen Organic Growth

Sulphur, • small amount of iron, • Obligate aerobes: require
Factor
• 50% of bacterium dry
weight is carbon Phosphorus copper, molybdenum, high concentration of • Many bacteria can
and zinc essential for oxygen. Ex: respiration. synthesize all their own
• Chemoheterotrophs & • To synthesize cellular the functions of certain • Obligate anaerobes: vitamins & are not
photoheterotrophs: materials (amino acids, enzymes, usually as cannot grow if oxygen dependent on outside
Gained from organic protein, DNA, RNA, cofactors. available. sources.
compounds such as nucleic acid,
carbohydrate, protein, • Facultative anaerobes: can • Ex: vitamins, amino
phospholipid) acid, purines,
lipid grow in both conditions
(presence & absence of pyrimidines.
• Chemoautotrophs &
photoautotrohs: oxygen)
Gained from CO2 • Microaerophiles: require
oxygen at low
concentration.

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Bacterial Growth: Bacterial Division

 Bacteria normally reproduce by

binary fission

 Few bacterial species reproduce by

budding

 Yeast: Mitosis

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 Bacterial Growth: Generation time

What is generation time?

The time required for a cell to divide and its

population to double.
 Varies among organisms and with environmental conditions, such as temperature.
 Generation time for most bacteria: 1-3 hours/generation
 Logarithmic scales are generally used to graph bacterial growth.

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How do bacteria grow?

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Bacterial Growth Curve

Binary fission occur here 40

You are already familiar with:
• Structure of prokaryotic cells.
• Requirements of the cells to survive.

Do you know the activities that allow them to thrive?

- Even the most structurally simple organism involve complex
biochemical reactions.

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1.5 Microbial metabolism
Topic learning outcomes:
- Define microbial metabolism
- Discuss several biochemical reactions
involved in the metabolisms

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Metabolism: Overview
Sum of all chemical reactions within a living organism.

• This process generate energy

from nutrients and the energy
will be used for the activity of the
cell.
• Two types of metabolic reactions:
a. Catabolic reaction
b. Anabolic reaction

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 Metabolism: Types of reaction
Anabolic Catabolic
• build up of molecules by using • breakdown of molecules to
the energy (ATP) produce energy (ATP)
• endergonic • exergonic
• Dehydration reactions (release • Hydrolytic reactions (use water)
water)
• Ex: Cell break down sugars
• Ex: Formation of proteins from (carbohydrate) into CO2 & H2O.
amino acids.

Chemical composition of a living cell is constantly changing: some molecules

are broken down while others are being synthesized. 44
 How do you make sure sugars will be dissolved in the
water?

Do you think that microbes can survive in such condition?

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Enzymes: Overview
The metabolism process is controlled by enzymes (enzyme-regulated activities).

 Enzymes are biological

catalysts that can speed up
biochemical reactions at a
temperature that is
compatible with the normal
functioning of the cell.
 Components:
 Apoenzyme (protein portion,
inactive)
 Cofactor (nonprotein portion,
active)

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Enzymes: Reactions
The metabolism process is controlled by enzymes (enzyme-regulated activities).

 Enzyme-substrate complex
(temporary binding).
 Ex: Enzyme sucrose can catalyze
sucrose (substrate) to form
glucose and fructose (product).

Enzyme

Sucrose Glucose + Fructose

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The growth or activity of the microbes can be regulated by
controlling the activity and synthesis of the enzymes.

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 Normal Enzyme Reaction

Enzymes: Factors
How do these factors affecting the growth of microbes?
Substrate bind to active site, have reaction
Substrate
Temperature pH Inhibitors
Concentration
Types of inhibitors:
Non-competitive inhibitors

Inhibitor bind to allosteric site, cause shape changes in active

site, so substrate cannot bind, thus block enzyme reaction

Optimal pH of enzyme is pH7

Competitive inhibitors

Enzyme denatured if pH is
too acidic/ too basic

Optimal temperature of
enzyme is 35°C Inhibitor compete with substrtae for active site, thus
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block enzyme reaction
The enzymes-regulated activities will involved energy-release
& energy-requiring reactions.

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Energy Production: Oxidation-Reduction Reactions
To extract & store energy (metabolism), these involve many reactions in which electrons are
passed from one molecule to another.

• A redox reaction is when

one molecule loses
electrons and is oxidized,
while another molecule
gains electrons (the ones
lost by the first molecule)
and is reduced.
• Reactions involving
electron transfers are Handy mnemonic: “LEO goes GER”: Lose Electrons, Oxidized; Gain Electrons, Reduced.
known as oxidation-
reduction reactions (or
redox reactions). 51
Metabolism: Energy Production
• The reactions that extract energy from molecules like
glucose are called catabolic reactions.

• As a glucose molecule is gradually broken down, some of the

breakdowns steps release energy that is captured directly as
ATP.

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Metabolism: Carbohydrate Catabolism
Breakdown of carbohydrate (ex: glucose) molecules to produce energy

• Most microorganisms
oxidize carbohydrates
as their primary Substrate-level phosphorylation

source of cellular
energy. Ex: Glucose.
• 2 general processes:
Cellular respiration &
Fermentation

Oxidative phosphorylation

Cellular respiration involves a

long series of oxidation-reduction
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process
54
Carbohydrate Catabolism: Cellular Respiration
 After glucose broken down into
pyruvic acid, the pyruvic acid can
be channeled into either cellular
respiration or fermentation.
 2 types of cellular respiration:
 aerobic respiration: uses
oxygen. (glycolysis, Krebs
Cycle, electron transport
chain)- generate 38 ATP
 anaerobic respiration:
does not use oxygen-
generate less ATP (varied
among microbes)

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Carbohydrate Catabolism: Cellular Respiration
 After glucose broken down into
pyruvic acid, the pyruvic acid can be
channeled into either cellular
respiration or fermentation.
 Fermentation:
 Release energy from sugar or
other organic molecules.
 Does not require O2.
 Does not require Krebs cycle
and electron transport chain.
 Produce only small amounts
of ATP.
 Ex: alcohol fermentation,
lactic acid fermentation.

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What does you inherit from your parents?

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1.6 Bacterial genetics
Topic learning outcomes:
- Explain the mechanism involve in gene
transfer of bacteria

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Genetic recombination refers to the
exchange of genes between two
DNA molecules to form new
combinations of genes on a
chromosome.

How does the genes brought close together to recombine?

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• In plant & animal, genes can be
passed to the offspring by
vertical gene transfer (from The movement of
parent to offspring). genes between
unicellular or
• In bacteria, genes can be passed multicellular is called
not only to the offspring, but gene transfer.
also to other microbes of the
same generation by horizontal
gene transfer (from parent to
offspring & other organism).
• This involves donor & recipient
cells.
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Bacterial Gene Transfer: Mechanism
• Transformation – genetic transfer
process by which free/naked DNA is
incorporated into a recipient cell
and brings about genetic change.

• Transduction – a bacterial virus

(bacteriophage) transfers DNA from
one cell to another.

• Conjugation- mechanism of
genetic transfer that involves cell-to-
cell contact, conjugated cells must
be opposite mating type.(donor cells
have plasmid, recipient do not) 61
One Genetic
Engineering Story

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 UNIT 1 - SUMMARY
1. Information flow within a cell
2. Central dogma of molecular biology
3. Purpose of cellular regulation
4. Differences between prokaryotic and eukaryotic
cells
5. Requirements for microbial growth
6. Bacterial growth curve
7. Microbial metabolism
8. Bacterial genetics – mechanism of gene transfer

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