Basic Principle of

Microbiology

Jumeri M. Wikarta, Ph.D

 What is Microbiology?
 The science of microorganisms
 Microorganisms : all single-
celled microscopic organisms
and include the viruses
 The discipline is just over a
century old
 Has given rise to molecular
biology and biotechnology
Bacteria : very large group of very small cells

A single colony can contain more than 10 million (107)individual cells

The catalytic and genetic functions of the cell
Microorganisms in modern agriculture
What are Microbes?
 Prokaryotes
• Nucleoid region
(center) contains the
DNA
• Surrounded by cell
membrane & cell wall
(peptidoglycan)
• Contain ribosomes (no
membrane) in their
cytoplasm to make
proteins
Prokaryotes

Morphology of a Gram-Positive Bacterium

Cell walls of Bacteria. (a, b) Schematic diagrams of gram-positive and gram-negative cell walls. The Gram stain photo in the center
shows cells of Staphylococcus aureus (purple, gram-positive) and Escherichia coli (pink, gram-negative). (c, d) Transmission electron
micrographs (TEMs) showing the cell wall of a gram-positive bacterium and a gram-negative bacterium. (e, f) Scanning electron
micrographs of gram-positive and gram-negative bacteria, respectively.
 Eukaryotes
• Cells that HAVE a
nucleus and
membrane-bound
organelles
• Includes protists,
fungi, plants, and
animals
• More complex type
of cells
 Organelles

• Very small (Microscopic)

• Perform various functions for a cell
• Found in the cytoplasm
• May or may not be membrane-bound
Sel prokariot dan eukariot
 Why study Microbiology (1)?
• Microorganisms were the first
life on earth
• Microorganisms created the
biosphere that allowed
multicellular organisms to evolve
• >50% of the biomass on earth is
comprised of microorganisms
• Microorganisms will be on earth
forever
An evolutionary phylogenetic tree
 Why study Microbiology (2)?
• Our understanding of life has arisen largely
from studies of microorganisms (biochemistry
and genetics)
• Studies of microorganisms continue to
contribute to fundamental knowledge of life
processes
 Why study Microbiology (3)?

Health
Agriculture
Food
Environment
Microorganisms in modern agriculture. (a, b) Root nodules on this soybean plant contain
bacteria that fix molecular nitrogen (N2) for use by the plant. (c) The nitrogen and sulfur
cycles, key nutrient cycles in nature. (d) Ruminant animals. Microorganisms in the rumen of
the cow convert cellulose from grass into fatty acids that can be used by the animal.
Biofuels. (a) Natural gas (methane) is collected in a funnel from swamp sediments
where it was produced by methanogens and then ignited as a demonstration
experiment. (b) An ethanol plant in the United States. Sugars obtained from corn or
other crops are fermented to ethanol for use as a motor fuel extender
Endang S. Rahayu Fak.
Teknologi Pertanian UGM
Pathways of Discovery in Microbiology
The Historical Foundations of Microbiology

• The Development of the Microscope

• The Debate over Spontaneous Generation
– Redi
– Needham
– Spallanzani
– Virchow
– Pasteur
 History of Microbiology

 Robert Hooke- First person to peer into the microbial world,

used compound lenses (1664)
 Anton van Leeuwenhoek- First true microbiologist, Dutch -
Simple Microscope
 single lens microscopes capable of 300 to 500X (1632 -
1723)
 drew and published (Proceeding of the Royal Society in
London) accurate pictures microscopic algae, protozoa,
bacteria, demonstrated “wee beasties” (“wee
animalcules”)
Endang S. Rahayu Fak.
Teknologi Pertanian UGM
Endang S. Rahayu Fak.
Teknologi Pertanian UGM
 Spontaneous Generation

• Spontaneous Generation- the idea that life

arose from non-living matter - Abiogenesis
– Aristotle (384-322 BC) believed some simpler
invertebrates could arise by spontaneous
generation
 Spontaneous Generation Debate

Francesco Redi (1626-1697)

 belief at the time- maggots spontaneously arose

from rotting meat

 demonstrated that maggots were dependent on

the presence of fly eggs and of rotting meat
 Controversy Continues

• Leeuwenhoek’s discovery added fuel to the

controversy
• Some proposed microorganisms could
arise from spontaneous generation even if
higher organisms did not
– They pointed out that boiled extracts of hay and
meat could give rise to microorganisms
 Spontaneous Generation
(continued debate)
• In 1748, English Priest John Needham reported
the results of his experiments
– boiled mutton broth and then tightly sealed the
flask
– eventually many of the sealed flask became cloudy
with microorganisms
– he proposed that organic matter possessed a ‘vital
force’ that could give rise to life
 Spontaneous Generation disproved

• Lazzaro Spallanzani - improved on Needham’s

experimental design
• first- sealed seeds and water in glass flask
• second- boiled them
• no growth took place as long as the flasks remained
sealed
• proposed that air carried microbes to the culture
 Controversy finally settled

• 1859, one of the fathers of modern microbiology, Louis Pasteur

offered strong evidence against Spontaneous Generation
• Pasteur was a chemist by training but became interested in
biology during his early studies on fermentations.
• At that time virtually nothing was known about microorganisms
– except that they existed (this was discovered by Anton van
Leeuwenhoek using home-made microscopes in 1684).
• It was known that microorganisms could be found in decaying
matter but it was not clear what role they were playing in the
decomposition process.
 Louis Pasteur (1822-1895)

"Do not put forward

anything that you
cannot prove by
experimentation"

Drew out the necks of

flasks so that they
remained open to the air
added broth and boiled
it to destroy any
• microbes present
flasks were incubated
for months and
remained sterile

Endang S. Rahayu Fak.

Teknologi Pertanian UGM
The defeat of spontaneous generation: Pasteur’s swan-necked flask experiment. In
(c) the liquid putrefies because microorganisms enter with the dust.
 (a) A French 5-franc note
honoring Pasteur

b) The Pasteur Institute, Paris, France.

Today this structure, built for Pasteur
by the French government, houses a
museum that displays some of the
original swan-necked flasks used in
his experiments.
 Pasteur’s Swan Neck Flask Experiment

•
 The Golden Age of Microbiology

• Fermentation and Pasteurization

– Pasteur
• The Germ Theory of Disease
– Agostino Bassi and Pasteur
– Joseph Liter
– Robert Koch
• Vaccination
– Edward Jenner
– Pasteur
 Rapid Advances 1857 -1914
Rudolf Virchow introduced the concept of
biogenesis: living cells can arise only from
preexisting cells (1857).
Louis Pasteur demonstrated that microorganisms
are in the air everywhere and offered proof of
biogenesis (1861).
Agostino Bassi (1835) and Pasteur (1865) showed a
causal relationship between microorganisms and
disease.
 Pasteur’s Contributions to Science

• 1800- French Government (Napoleon III)

enlisted his help in their Wine Industry

– showed the decline in quality was due to a

contaminating microbe
– process of pasteurization
 Pasteur’s Contributions to Science

• Recognized that chickens became immune to

a bacterial pathogen if injected with a weaken
avirulent strain.

• Developed vaccines against rabies and anthrax

Pasteur’s perplexing problem
while working on spontaneous generation he
experienced an anomaly
no matter how long some flasks were boiled, they
always produced growth
German botanist-Ferdinand Cohn discovered the
existence of heat resistant bacterial spores
Pasteur reasoned that the flask that produced
growth contained heat resistant bacterial spores
English physicist John Tyndall dealt the absolute final
blow to Spontaneous Generation and resolved
Pasteur’s problem, he demonstrated the dust did
indeed carry microbes and if the dust were absent,
the broth remained sterile.

He developed the process of tyndallization which is

fractional steam sterilization
similar to the process we use today in the lab.
 The Recognition of the Microbial Role in Disease

Agostino Bassi (1835)

 linked a microorganism to disease

Pasteur (1865)
 disease of silkworms
 The Recognition of the Microbial Role in Disease

Joseph Lister
 English surgeon
 developed an antiseptic system of surgery and
heat sterilized his instruments as well as using
phenol with surgical dressing
 Robert Koch
(1843 –1910)

Direct evidence of the role of bacterial

in disease transmission

Robert Koch
established a relationship between anthrax and the
bacterium- Bacillus anthracis
Koch’s techniques are still used today
Today we know these as Koch’s Postulates
Received the Nobel prize 1905 for his work on
tuberculosis
 Koch’s Postulates
On the basis of these studies Koch proposed a series of criteria
that should be used to establish the link between disease and
microorganisms. These are now known as Koch’s postulates.

1: Microorganisms must always be present in the infected

animal and be absent in healthy individuals.

2: Pure cultures of the microorganism must be obtained from

the diseased animal.

3: Such a culture must be capable, in itself, of causing disease

in healthy animals.

4: The microorganism should be re-isolated from the diseased

animal and should retain the same properties in pure culture
as the original pure culture.
 Koch’s Tuberculosis studies
• In 1881 Koch turned his attention to the study of a human disease
– Tuberculosis (TBC).

• At that time TB killed 1 in 7 people and even today it accounts for

more than 1 in 20 human deaths worldwide. TBC was known to be
contagious but no “contagion” had been identified.

• TBC is a disease of the lungs caused by the bacterium

Mycobacterium tuberculosis. It is acquired by inhalation of
infected droplets or dust particles in the air.

• The bacterium can persist in the lungs for long periods. It survives
and grows within macrophages.

• In individuals with a low resistance acute pulmonary infection can

occur with extensive damage to lung tissue. Spread of the
bacterium to other parts of the body can then lead to death.
 Robert Koch (1843 – 1910)
• Developing pure culture technique
• Use potato slices – first found individual
bacterial colonies growing with different
appearance
• Microscopic examination revealed cells within
a single colony were similar
 Pure Culture Key to Studying Microbes

Definition : Pure culture is a population of

organisms, all of which are the progeny of a
single organism
In nature, microbes almost never occur as
pure cultures
 Development of solidifying agents and complex
media

• Koch wanted to culture pathogens so he used

something similar to the body tissue
• Meat extracts
• Initially, gelatin (jell-o) was used to solidify
beef extract broth but : (1) many organisms
can digest gelatin and (2) it melts at 37 oC, the
favored incubation temperature for most
pathogens
 Development of solidifying agents

Agar was suggested by Fannie Hesse wife

of Walther Hesse working in Koch Lab
Story : Walther was working with gelatin
plates in summer and was having troubles
Asked wife : Why do your jellies and
pudding stay solid in warm weather?
Fannie learned to use Agar-agar from a
dutch neighbour in New York who spent
time in Asia
Agar-agar had been used as a gelling
agent in Asia for centuries
 Agar
Agar is complex polysaccharides
derived from seaweed
Its only melts at 100 oC, and
solidifies at 45 oC
Non-toxic to most
microorganism
Stable at sterilization
temperature
Only a very few bacteria have
enzymes that digest it
Petri dish was invented in 1877
By an assistant of Koch –
R.J. Petri

Shallow glass plates

 Alexander Fleming

• Observed that the mold Penicillium inhibited

the growth of a bacterial culture.

• Penicillin has been used clinically as an

antibiotic since the 1940s
Anthrax & Bioterrorism
 Bioterrorism 1
Use of biological material to inflict suffering
and/or death on enemies
(board definition)

• Biological warfare is not a new idea; In 184BC Hannibal

threw baskets of snakes onto enemy ships!

• In 1346 Tartar soldiers catapulted plague victims into

besieged Kaffka and successfully started an epidemic.

• In 1763 the British army gave smallpox-contaminated

blankets to Indians who were loyal to the French.
 Bioterrorism 2
• In 1925 the Geneva convention outlawed the use of
biological weapons.

• In 1969 the US “officially” ended its program of developing

biological weapons.

• In 1972 the Biological and Toxin weapons convention

approved a treaty to outlaw biological weapons
(140 countries signed this treaty)

• It is likely that many countries still have

the capacity for biological.
 What is Anthrax?
• Anthrax is a disease that is caused by the bacterium
Bacillus anthracis (the link was first proved by Koch).
• B. anthracis is a Gram-positive, spore-forming common
soil microorganism.
• Primarily a disease of livestock; catlle, sheep etc.
• Pasteur devised the first vaccine against anthrax.
• Today vacccination is still the primary means of control.
 Anthrax in Humans
• Contrary to reports in the popular press – B. anthracis is not a
good human pathogen
• Humans are fairly resistant to infection. Even textile workers
exposed to infected animal products have a low infection
rate.
• Large numbers of spores (8,000-10,000) are required to infect
humans. This exposure is very rare in the “normal”
environment.
• Breathing air around infected animal carcasses would lead to
the inhalation of around 2-10 spores every 10 minutes!
 Infection by B. anthracis
There are 3 routes of infection by this
pathogen:

1) Entry by ingestion
• Gastrointestinal anthrax can arise by
the ingestion of contaminated meat, usually from an
animal that died of anthrax.

• Infection reaches intestinal lymph nodes, then causes

septacemia. Exact pathogenesis is not understood.

• Very rare – but in cases that present clinically the

mortality rate is about 50%.
2) Entry through skin lesions.

• Spores enter damaged areas of skin.

• Erythematous papule after 2-5 days (red bump).

• Centre of lesion darkens after 7-10 days.

• Usually heals without

intervention
(spread to blood occurs
in less than 5% of cases).
 3) Entry by inhalation
• Inhalation of spores can lead to pulmonary anthrax – also
called “woolsorters disease”.

• Malaise within 1-5 days, including cough, fever and chest

pain.

• Difficulty with breathing, blueish skin colour, swelling of

the chest.

• Once swelling is evident the

disease beyond treatment
and death usually occurs.
 Anthrax and bioterrorism

Advantages:

• Very easy to obtain and to grow.

• Spores very stable.
• Spores easily inhaled.
• High fatality rate in cases of pulmonary anthrax.

Disadvantages:

• Dispersing spores over a wide area is difficult.

• Not contagious – no person-to-person spread.
• If diagnosed early it is easily treated with antibiotics.
 Concept Check
Microorganisms can be both
beneficial and harmful to humans.
Although we tend to emphasize
the harmful microorganisms
(infectious and disease agents),
many more microorganisms are
beneficial than harmful.
In what ways are microorganisms
important in the food industries ?