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Microbiology
The study of microorganisms
The discovery of microbes
• Roman philosopher Lucretius (about 98–
55 B.C.) and the physician Girolamo
Fracastoro (1478–1553) suggested that
disease was caused by invisible living
creatures.
• The earliest microscopic observations
made by the Italian Francesco Stelluti
The first person to observe and describe microorganisms accurately
was the amateur microscopist
Antony van Leeuwenhoek (1632–1723) of Holland
His hobby was making microscopes…!!!
• Constructed simple microscopes composed of
double convex glass lenses held between two
silver plates
• Could magnify around 50 to 270 times,
• Illuminated his liquid specimens by placing them
between two pieces of glass and shining light on
them at a 45° angle to the specimen plane.
• A form of dark-field illumination
• Bacteria clearly visible.
• Sent detailed letters describing his discoveries to
the Royal Society of London.
• Classification: The orderly arrangement
of units under study into groups of larger
units.
• Nomenclature
• Carolus Linnaeus, a Swedish botanist.
• Until 18th century, the classification of
living organisms placed all organisms into
one of two kingdoms, plant and animal.
Kingdom Protista
• E. H. Haeckel, a German zoologist in 1866
• A third kingdom - Protista.
• To
include
those
unicellular
microorganisms that are typically neither
plants nor animals.
– Bacteria, Algae, Fungi, and Protozoa
– Viruses are not cellular – So, not classified as
Protists
Kings Play Chess On Fine Glass Stools
– Domain
– Kingdom
– Phylum / Division
– Class
– Order
– Family
– Genus
– Species
???
• What criteria could be used to distinguish a
bacterium from a yeast or certain microscopic
algae?
• Aid of Microscopy - observation of internal cell
structure
• In bacteria, the nuclear substance was not
enclosed by a nuclear membrane.
• In algae and fungi, the nucleus was enclosed in
a membrane.
• The presence / absence of membrane-bound
internal structures is the criterion.
– Lower protists: Bacteria
– Higher protists: Algae, Fungi, and Protozoa.
PROKARYOTES
&
EUKARYOTES
FEATURE
PROKARYOTES
EUKARYOTES
Groups
Bacteria
Algae, Fungi, Protozoa, Plants
and Animals
Size range
1-2 by 1-4 μm
Greater than 5 μm in width or
diameter
Cell Wall
Peptidoglycan (murein or
mucopeptide) as component
Absence of peptidoglycan
Cell Membrane
Generally do not contain sterols;
contain part of respiratory and, in
some, photosynthetic machinery
Sterols present; Respiration and
photosynthesis do not occur
Location of Genetic content
Nucleoid, chromatin body or nuclear
material
Nucleus, mitochondria,
chloroplasts
Nuclear Structure
Not bounded by nuclear membrane;
one circular chromosome
Bounded by nuclear membrane;
more than one chromosome
Chromosome does not contain
histones
Chromosomes have histones
Nucleolus absent
Nucleolus present
Refer Pelczar,
for a detailed list of differences.
Classification Systems
• Two Kingdom System.......
– Plantae / Animalia
• Five Kingdom System, 1969
• Three Domain System, 1978
Whittaker’s Five-kingdom Concept
• A recent system of classification
• Proposed by R. H. Whittaker (1969).
• Based on 3 levels of cellular organization
that evolved to explain the 3 principal
modes of nutrition:
– photosynthesis,
– absorption. and
– ingestion.
• Kingdom Monera:
– The Prokaryotes
– They lack the ingestive mode of nutrition.
• Kingdom Protista:
–
–
–
–
Unicellular eukaryotic microorganisms
All 3 nutritional types are represented here
The nutritional modes are continuous
The mode of nutrition of the microalgae is
photosynthetic
• Kingdom Plantae
– The mode of nutrition of the protozoa is ingestive
• Kingdom Animalia
– The mode of nutrition in some other protists is
absorptive
• Kingdom Fungi
• The Three Domains
– Eukarya - all eukaryotic orgnaisms
» Kingdoms:
» Animalia
» Plantae
» Fungi
» Protista
– Bacteria
» Bacteria, spirochetes, cyanobacteria etc
– Archaea - "ancient" bacteria living extreme environments
» methanogens
» extreme halophiles
» extreme thermophiles
Bergey’s Manual of Systematic Bacteriology
The international standard for bacterial taxonomy
• Kingdom Prokaryotae: All bacteria included
• Divided into 4 divisions as follows:
• Division 1: Gracilicutes
– Prokaryotes with a complex cell-wall structure characteristic of
Gram-negative bacteria
• Division 2: Firmicutes
– Prokaryotes with a cell-wall structure characteristic of Grampositive bacteria
• Division 3: Tenericutes
– Prokaryotes that lack a cell wall
• Division 4: Mendosicutes
– Prokaryotes of an earlier phylogenetic origin.
Viruses
• Very small noncellular obligate parasites or
pathogens of plants, animals, and bacteria as
well as other protists.
• So small that they can be visualized only by the
electron microscope.
• Can be cultivated only in living cells.
• Range: 0.015 - 0.2 μm
• Does not grow on artificial laboratory media
• Electron microscopy required to see viruses
Bacteria
– Procaryotic
– Unicellular procaryotic organisms or simple
associations of similar cells.
– Reproduction asexual: Cell multiplication is
usually by binary fission.
– Grows on artificial laboratory media:
characteristically by simple cell division
– Range: 0.2 by 100 μm
Protozoa
– Unicellular eukaryotic organisms.
– Differentiated on the basis of morphological,
nutritional, and physiological characteristics.
– Range: 2 - 200 μm
– Some are intracellular parasites
– Reproduction by asexual & sexual processes
Fungi
– Eukaryotic lower plants devoid of chlorophyll.
– Usually multicellular but are not differentiated into
roots, stems, and leaves.
– Single-celled microscopic yeasts to Molds to giant
multicellular mushrooms and puffballs.
– Composed of filaments and masses of cells which
make up the body of the organism, known as a
mycelium.
– Reproduce by fission, budding, or by means of
spores.
Yeasts
• Range: 5 -10 μm
• Eukaryotic; unicellular.
• Reproduction by asexual cell division,
budding, or sexual processes
• Production of alcoholic beverages; also
used as food supplements
Molds
• Range: 2 -10 μm by several mm
• Eukaryotic; multicellular
• Reproduction by asexual and sexual
processes
• Responsible for decomposition
(deterioration) of many materials;
• Useful for industrial production of penicillin
Algae
– Range: 1.0 μm to many feet
– Eucaryotic; unicellular and multicellular:
reproduction by asexual and sexual
processes
– Other algae, such as large brown Kelp,
have a complex structure with cell types
specialized for particular functions.
– All algae contain chlorophyll & are
capable of photosynthesis.
–Found most commonly in aquatic
environments or in damp soil.
–Important to the production of food
in aquatic environments:
–Used as food supplement and in
pharmaceutical preparations
–Source of agar for microbiological
media; some produce toxins.
MICROSCOPE
• Roger Bacon (1220-1292) postulated that disease is
produced by invisible living creatures.
• Girolamo Fracastoro (1483-1553) & Anton von Plenciz in
1762, but without proof.
• In 1658, Athanasius Kircher (1601-1680) referred to
“worms” invisible to the naked eye in decaying bodies,
meat, milk, and diarrheal secretions.
• In 1665, Robert Hooke’s description of cells in a piece of
cork
• Anton van Leeuwenhoek (1632 – 1723) was the first to
report his observations with accurate descriptions and
drawings.
• “wee animalcules” which we recognize as free-living
protozoa.
SPONTANEOUS GENERATION
vs BIOGENESIS
• Aristotle: “Animals might originate spontaneously
from the soil, plants, or other unlike animals”
• An accepted fact: Maggots could be produced
by exposing meat to warmth and air.
• Francesco Redi (1626-1697)
– Placed meat in a jar covered with gauze.
– Attracted by the odor of the meat, flies laid eggs on
the covering, and
– From the eggs maggots developed.
– Hence the experiment established the fact that the
origin of the maggots was the flies and not the “meat”
• John Needham
– Meat exposed to hot ashes, observed the
appearance of organisms not present at the
start of the experiment and concluded that the
bacteria originated from the meat.
• Lazaro Spallanzani
– Boiled beef broth for 1 hour & then sealed
flasks.
– No microbes appeared following incubation.
• Needham argued that air was essential to the
spontaneous production of microscopic beings
and by sealing …..
• Franz Schuize & Theodor Schwann
– Schuize passed air through strong acid solutions into
boiled infusions
– Schwann passed air into his flasks through red-hot
tubes
– In neither case did microbes appear.
• But still,
– “Acid and heat altered the air so that it would not
support growth”
• In 1850, H. Schroder and T. von Dusch
– Passed air through cotton into flasks
containing heated broth.
– Thus the microbes were filtered out of the air
by the cotton fibers so that,
• Growth did not occur
– A basic technique of plugging bacterial culture
tubes with cotton stoppers was initiated.
A last attempt…..by Felix Archimede
Pouchet, who published a report in 1859
“proving”
Spontaneous Generation…
The final answer…
By the Father of Microbiology
• Louis Pasteur
– Prepared a flask with a long, narrow
gooseneck opening
– Nutrient solutions were heated in the flask
– Air - untreated and unfiltered - could pass in
or out: but the germs settled in the gooseneck
– No microbes appeared in the solution
A lasting assault…!!!
• In 1877, English physicist John Tyndall
– Demonstrated that dust did indeed carry
germs and
– if dust was absent, broth remained sterile
even if directly exposed to air.
THE GERM THEORY OF DISEASE
• Fracastoro of Verona suggested that
diseases might be due to invisible
organisms transmitted from one person to
another
• Von Plenciz stated that living agents are
the cause of disease and suspected that
different germs were responsible for
different diseases.
• Robert Koch (1843-1910)
– The father of Medical Microbiology
– Anthrax problem
– Discovered the typical bacilli with squarish
ends in the blood of cattle that had died of
anthrax.
– Grew these bacteria in cultures in his
laboratory
– Examined them microscopically to be sure he
had only one kind present
• Injected them into other animals
• These became infected and developed clinical
symptoms of anthrax.
• From these experimentally infected animals, he
isolated microbes like those he had originally
found in sheep that died of anthrax.
• The first time, a bacterium had been proved to
be the cause of an animal disease.
• This series of observations led to the
establishment of …..
KOCH’s POSTULATES
•
•
•
•
Provided guidelines to identify the causative
agent of an infectious disease.
(1) A specific organism can always be found in
association with a given disease.
(2) The organism can be isolated and grown in
pure culture in the laboratory.
(3) The pure culture will produce the disease
when inoculated into a susceptible animal.
(4) It is possible to recover the organism in pure
culture from the experimentally infected animal.
Pure Cultures
• The growth of a mass of cells of the same
species in a laboratory vessel (such as a
test tube) is called a pure culture.
• Laboratory
procedures
to
isolate
microorganisms
representing
each
species and to grow (cultivate) each of the
species separately.
Pasteur’s exptl demo of Koch’s Postulates
PUZZLED !!!???
After few weeks,
• Inoculated healthy chickens with his pure cultures, but
chickens failed to die…!!!!
• Found that he had accidentally used cultures several
weeks old
• Repeated the experiment, using 2 groups of chickens.
– One group inoculated at the first demonstration with the old
cultures that had proved ineffective, and
– the second had not been previously exposed.
•
•
•
•
Both groups received bacteria from fresh young cultures.
First group remained healthy….!!!!
Second group got sick and died,
Explanation:
– Bacteria still retained their capacity for stimulating the host to
produce substances, i.e. antibodies, that protect against
subsequent exposure to virulent organisms.
Edward Jenner
• To test his theory, drawn from folklore of the countryside
– “Milkmaids who suffered the mild disease of cowpox never
contracted smallpox”
• In 1796, the famous experiment on 8-year-old James
Phipps.
• Pus taken from a cowpox pustule from a dairymaid, on
May 14, 1796.
• Injected it into an incision on the boy's arm.
• Subsequent inoculation with smallpox
• Demonstrated the boy's immunity to the disease.
• Jenner's first vaccination of a human patient.
Sarah Nelmes, a milkmaid
• Proved that having been inoculated with
cowpox, Phipps was immune to smallpox.
• Submitted a paper to the Royal Society in
1797
• Was told that his ideas were too
revolutionary and that he needed more
proof…!!!
• Experimented on more children, including his
own 11-month-old son.
• In 1798, published results
• Coined the word vaccine from the Latin 'vacca'
for cow.
• Was widely ridiculed; Claimed it was “ungodly”
to inoculate someone with material from a
diseased animal.
• Soon, the advantages of vaccination and the
protection it provided won out, and vaccination
soon became widespread.
• The last endemic smallpox infection was
recorded in 1977
• The last known case of death due to
smallpox in 1978.
• The
WHO
declared
the
disease
completely eradicated two years later.
Back on stage… Pasteur
• To make a vaccine for hydrophobia, or
rabies,
• A disease transmitted to people by bites of
dogs, cats, etc..
• Joseph Meister bitten by a mad wolf.
• Disease could be produced in rabbits by
inoculating them with saliva from mad dogs.
• Brain and spinal cord could be removed from the
infected rabbits, dried, pulverized, and mixed
with glycerin.
• Injected this mixture into dogs protected them
against rabies
• Also, Injected this mixture into Joseph Meister
• Did not die…..!!!! Recovered