Introduction of microbiology kbk

INTRODUCTION OFINTRODUCTION OF
MICROBIOLOGYMICROBIOLOGY
Laboratory of MicrobiologyLaboratory of Microbiology
Faculty of MedicineFaculty of Medicine
Brawijaya UniversityBrawijaya University

microbiologymicrobiology
The branch of biology that deals withThe branch of biology that deals with
microorganisms and their effects on other livingmicroorganisms and their effects on other living
organismsorganisms
Many branches of Microbiology :Many branches of Microbiology :
- Environmental Microbiology- Environmental Microbiology
- Space Microbiology- Space Microbiology
- Marine Microbiology- Marine Microbiology
- Agricultural Microbiology- Agricultural Microbiology
- Food Microbiology ……. etc.- Food Microbiology ……. etc.
-- Medical MicrobiologyMedical Microbiology

First SessionFirst Session
A BRIEF HISTORY OFA BRIEF HISTORY OF
MICROSCOPEMICROSCOPE

A BRIEF HISTORY OFA BRIEF HISTORY OF

THE FIRST OBSERVATION
THE DEBATE OVER SPONTANEOUS
GENERATION
THE GOLDEN AGE OF MICROBIOLOGY
THE BIRTH OF MODERN CHEMOTHERAPY :
DREAMS OF A “MAGIC BULLET”
MODERN DEVELOPMENTS IN MICROBIOLOGY

THE FIRST OBSERVATION
 1665 – An Englishman, Robert Hooke,
reported to the world that life’s smallest structural units were
“little boxes” or “cells”
Using his improving version of a compound microscope 
he was able to see individual cells  beginning of the cell theory 
that all living things are composed of cells
 1673 – 1723, Antony van Leeuwenhoek
The Dutch merchant and amateur scientist was the first to actually
observe live microorganism through his simple single lens microscope
 the “animalcules”  in rainwater, in liquid in which peppercorn had
soaked, and in material scraped from his teeth
He drew the basic form of bacteria as coccus, rods and spiral

THE DEBATE OVER SPONTANEOUS GENERATION
 Until the second half of the 19th
century many scientists and philosophers
(Aristoteles, Samson, Virgil) believed that some form of life could arise
spontaneously from nonliving matter  spontaneous generation
 People commonly believed that toads, snakes, and mice could be born
of moist soil; that flies could emerge from manure; and that maggots
could arise from decaying corpses
Evidence Pro and Con
 1668 – Fransesco Redi, the Italian physician, a strong opponent of
spontaneous generation  demonstrate that maggots do not arise
spontaneously from decaying meat
Redi filled three jars with decaying meat and sealed them tightly.
Then he arranged three other jars similarly but left them open. Maggots
appeared in the open jars.Sealed containers showed no signs of maggots
 Redi’s antagonists were not convinced; they claimed that fresh air was
needed for spontaneous generation

 Redi set up a second experiment  three jars were covered with a fine
net instead of being sealed. No larvae appeared in the gauze-covered jars
 Many scientists still believed that small organisms (van Leeuwenhoek’s
animalcules) were simple enough to be generated from nonliving materials
 1745 : spontaneous generation seem to be strengthened, when
John Needham, an Englishman, found that even after he heated nutrient
fluid before pouring them into covered flasks, the cooled solution were
soon teeming with microorganisms.
Needham claimed that microbes developed spontaneously from the fluid
 Twenty years later : Lazzaro Spallanzani, an Italian scientist, suggested
that microorganisms from the air probably had entered Needham’s
solutions after they were boiled. Spallanzani showed that nutrient fluids
heated after being sealed in a flask did not develop microbial growth
 Needham responded by claiming the “vital force” necessary for sponta-
neous generation had been destroyed by the heat and was kept out of
the flasks by the sealed

 Laurant Lavoisier showed the importance of oxygen to life.
 Spallanzani’s observations were criticized on the grounds that there was
not enough oxygen in the sealed flasks to support microbial life
The Theory of Biogenesis
 Rudolf Virchow challenged spontaneous generation with the concept
of biogenesis, he claim that living cells can arise only from preexisting
living cells
 Arguments about spontaneous generation continued until 1861, when the
issue was resolved by the French scientist Louis Pasteur
 With a series of ingenious experiments, Pasteur demonstrate that
microorganisms are present in the air and can contaminate sterile
solutions, but air itself does not create microbes

Pasteur’s experiments disproving the theory of
spontaneous generation
(1) Pasteur first poured beef broth into a long-necked flask. (2) Next he heated the neck of the flask
and bent it into an S-shaped curve; then he boiled the broth for several minutes. (3)
Microorganisms did not appear in the cooled solution, even after long periods

 Pasteur showed that microorganisms can be present in
nonliving matter  on solids, in liquids, and in the air
 He demonstrated that microbial life can be destroyed
by heat  form the basis of aseptic techniques
 The debate of Spontaneous Generation disproved
 However still have a problem of “spores”  resistant to
heat  John Tyndall (1820-1893)  TYNDALLIZATION
 Spontaneous Generation Theory totally finished

The Golden Age Of MicrobiologyThe Golden Age Of Microbiology
1857 - 19141857 - 1914
Pasteur & Robert Koch, led to the establishmentPasteur & Robert Koch, led to the establishment
of microbiology asof microbiology as a sciencea science
Discoveries during these years included both theDiscoveries during these years included both the
agents of many diseases and the role ofagents of many diseases and the role of
immunity in the prevention and cure of diseaseimmunity in the prevention and cure of disease
Fermentation and PasteurizationFermentation and Pasteurization
The Germ Theory of DiseaseThe Germ Theory of Disease
VaccinationVaccination

Fermentation & PasteurizationFermentation & Pasteurization
A group of French merchants asked Pasteur to find out whyA group of French merchants asked Pasteur to find out why
wine and beer souredwine and beer soured  a method that would preventa method that would prevent
spoilage ?spoilage ?
Many scientist believed that air converted the sugars inMany scientist believed that air converted the sugars in
these fluids into alcoholthese fluids into alcohol
Pasteur found instead that microorganisms calledPasteur found instead that microorganisms called yeastsyeasts
convert the sugars to alcohol in the absence of air. Thisconvert the sugars to alcohol in the absence of air. This
process, calledprocess, called fermentationfermentation, is used to make wine and, is used to make wine and
beerbeer
Souring and spoilage are caused by different micro-Souring and spoilage are caused by different micro-
organisms calledorganisms called bacteriabacteria. In the presence of air, bacteria. In the presence of air, bacteria
change the alcohol in the beverage into vinegar (acetic acid)change the alcohol in the beverage into vinegar (acetic acid)

Before the time of Pasteur, effective treatments for manyBefore the time of Pasteur, effective treatments for many
diseases were discovered by trial and error, but the causesdiseases were discovered by trial and error, but the causes
of the disease were unknownof the disease were unknown
The realization that yeasts play a crucial role inThe realization that yeasts play a crucial role in
fermentation was the first link between the activity of afermentation was the first link between the activity of a
microorganism and physical and chemical changes inmicroorganism and physical and chemical changes in
organic materialsorganic materials
This discovery alerted scientists to the possibility thatThis discovery alerted scientists to the possibility that
microorganisms might have similar relationships with plantsmicroorganisms might have similar relationships with plants
animals - specifically, that microorganisms might causeanimals - specifically, that microorganisms might cause
disease. This idea was known asdisease. This idea was known as the germ theory ofthe germ theory of
diseasedisease

1840 –1840 – Ignaz SemmelweisIgnaz Semmelweis a Hungarian physician, hada Hungarian physician, had
demonstrated that physician, who at the time did notdemonstrated that physician, who at the time did not
disinfect their hands, routinely transmitted infectionsdisinfect their hands, routinely transmitted infections
(puerperal, or childbirth fever) from one obstetrical patient to(puerperal, or childbirth fever) from one obstetrical patient to
anotheranother
1860 –1860 – Joseph ListerJoseph Lister an English surgeon, applied the german English surgeon, applied the germ
theory to medical procedure. Disinfectants were not used attheory to medical procedure. Disinfectants were not used at
the time, but Lister knew that phenol (carbolic acid) killsthe time, but Lister knew that phenol (carbolic acid) kills
bacteria, so he began treating surgical wounds with abacteria, so he began treating surgical wounds with a
phenol solutionphenol solution
1876 –1876 – Robert KochRobert Koch, a German physician, proved that, a German physician, proved that
bacteria actually cause diseasebacteria actually cause disease  discovered rod-shapeddiscovered rod-shaped
bacteria now known asbacteria now known as Bacillus anthracisBacillus anthracis in the blood ofin the blood of
cattle that had died of anthraxcattle that had died of anthrax

Koch’s research provides a framework for the study of theKoch’s research provides a framework for the study of the
etiology of any infectious diseaseetiology of any infectious disease  Koch Postulates :Koch Postulates :
1. The same pathogen must be present in every case of1. The same pathogen must be present in every case of
diseasedisease
2. The pathogen must be isolated from the diseased host2. The pathogen must be isolated from the diseased host
and grown in pure cultureand grown in pure culture
3. The pathogen from the pure culture must cause the3. The pathogen from the pure culture must cause the
disease when it is inoculated into a healthy, susceptibledisease when it is inoculated into a healthy, susceptible
laboratory animallaboratory animal
4. The pathogen must be isolated from the inoculated4. The pathogen must be isolated from the inoculated
animal and must be shown to be the original organismanimal and must be shown to be the original organism

VACCINATIONVACCINATION
1796 –1796 – Edward JennerEdward Jenner, a young British physician, embarked, a young British physician, embarked
on an experiment to find a way to protect people fromon an experiment to find a way to protect people from
smallpox :smallpox :
- A young milkmaid informed Jenner that she couldn’t get- A young milkmaid informed Jenner that she couldn’t get
smallpox because she already had been sick from cowpoxsmallpox because she already had been sick from cowpox
- First Jenner collected scrapings from cowpox blisters. Then- First Jenner collected scrapings from cowpox blisters. Then
he inoculated a healthy 8-year-old volunteer with thehe inoculated a healthy 8-year-old volunteer with the
cowpox material by scratching the person’s arm with acowpox material by scratching the person’s arm with a
pox-contaminated needle. In a few days, the volunteerpox-contaminated needle. In a few days, the volunteer
became mildly sick but recovered and never againbecame mildly sick but recovered and never again
contracted either cowpox or smallpox. The process wascontracted either cowpox or smallpox. The process was
calledcalled vaccinationvaccination (vacca=cow)(vacca=cow)
- The protection from disease provided by vaccination is- The protection from disease provided by vaccination is
calledcalled immunityimmunity

VACCINATIONVACCINATION
1880 – Pasteur discovered why vaccination work. He found1880 – Pasteur discovered why vaccination work. He found
that the bacteria that causesthat the bacteria that causes fowl cholerafowl cholera lost its abilitylost its ability
to cause disease after it was grown in the laboratory forto cause disease after it was grown in the laboratory for
long periods. However it – and other microorganisms withlong periods. However it – and other microorganisms with
decreased virulence – was able to induce immunity againstdecreased virulence – was able to induce immunity against
subsequent infections by its virulent counterpart.subsequent infections by its virulent counterpart.
Some vaccines are still produced from avirulent microbialSome vaccines are still produced from avirulent microbial
strains that stimulate immunity to the related virulent strain.strains that stimulate immunity to the related virulent strain.
Other vaccines are made from killed virulence microbes,Other vaccines are made from killed virulence microbes,
from isolated components of virulent micoorganisms, or byfrom isolated components of virulent micoorganisms, or by
genetic engineering techniquesgenetic engineering techniques

The Birth of Modern Chemotherapy :The Birth of Modern Chemotherapy :
Dreams of a “ Magic Bullet “Dreams of a “ Magic Bullet “
After the relationship between microorganisms andAfter the relationship between microorganisms and
disease was established, medical microbiologists nextdisease was established, medical microbiologists next
focused on the search for substances that could destroyfocused on the search for substances that could destroy
pathogenic microorganisms without damaging thepathogenic microorganisms without damaging the
infected animal or humaninfected animal or human
Treatment of disease by using chemical substances isTreatment of disease by using chemical substances is
calledcalled chemotherapychemotherapy
Chemotherapeutic agents prepared from chemicals inChemotherapeutic agents prepared from chemicals in
the laboratory are calledthe laboratory are called synthetic drugssynthetic drugs
Chemicals produced naturally by bacteria or fungi to actChemicals produced naturally by bacteria or fungi to act
against other microorganisms are calledagainst other microorganisms are called antibioticsantibiotics

The First Synthetic DrugsThe First Synthetic Drugs
Paul EhrlichPaul Ehrlich, a German physician, was the imaginative, a German physician, was the imaginative
thinker who fired the first shot in the chemotherapythinker who fired the first shot in the chemotherapy
revolution. Ehrlich speculated about a “magic bullet” thatrevolution. Ehrlich speculated about a “magic bullet” that
could destroy pathogen without harming the infected hostcould destroy pathogen without harming the infected host
In 1910, he found a chemotherapeutic agent calledIn 1910, he found a chemotherapeutic agent called
salvarsansalvarsan, an arsenic derivative effective against syphylis, an arsenic derivative effective against syphylis
By the late 1930s, researchers had developed several otherBy the late 1930s, researchers had developed several other
synthetic drugs, mostly were derivatives of dyes that couldsynthetic drugs, mostly were derivatives of dyes that could
destroy microorganismsdestroy microorganisms
In addition,In addition, Domagk (1935)Domagk (1935) discovered thatdiscovered that prontosilprontosil hadhad
dramatically effect against streptococcal infectionsdramatically effect against streptococcal infections  in thein the
body was changed intobody was changed into sulfanilamidesulfanilamide that analog withthat analog with
PABAPABA

A Fortunate Accident - AntibioticsA Fortunate Accident - Antibiotics
The first antibiotic was discovered by accidentThe first antibiotic was discovered by accident
1928 –1928 – Alexander FlemingAlexander Fleming, a Scottish physician and, a Scottish physician and
bacteriologist, looked at the curious pattern of growth on thebacteriologist, looked at the curious pattern of growth on the
contaminated plates. There was a clear area around thecontaminated plates. There was a clear area around the
mold where the bacterial culture had stopped growingmold where the bacterial culture had stopped growing
The mold was later identified asThe mold was later identified as Penicillium notatumPenicillium notatum, and, and
Fleming named the mold’s active inhibitor asFleming named the mold’s active inhibitor as penicillinpenicillin..
The enormous usefulness of penicillin was not apparentThe enormous usefulness of penicillin was not apparent
until the 1940suntil the 1940s  Florey & ChainFlorey & Chain
1939 –1939 – ReneRene´´ Dubos,Dubos, a French microbiologist, discovereda French microbiologist, discovered
two antibiotics calledtwo antibiotics called gramicidingramicidin andand tyrocidine.tyrocidine. Both wereBoth were
produced by a bacterium,produced by a bacterium, Bacillus brevisBacillus brevis, cultured from soil., cultured from soil.

Modern Development in MicrobiologyModern Development in Microbiology
BacteriologyBacteriology
MycologyMycology
ParasitologyParasitology
ImmunologyImmunology
VirologyVirology
Recombinant DNA technologyRecombinant DNA technology

VIROLOGYVIROLOGY
The study of virus, actually originated during the Golden Age ofThe study of virus, actually originated during the Golden Age of
MicrobiologyMicrobiology
1892 –1892 – Dmitri IwanowskiDmitri Iwanowski reported that the organism that causereported that the organism that cause
mosaic disease of tobacco was so small that it passed through filtermosaic disease of tobacco was so small that it passed through filter
fine enough to stop all known bacteriafine enough to stop all known bacteria
1935 –1935 – Wendell StanleyWendell Stanley demonstrated that the organism, calleddemonstrated that the organism, called
tobacco mosaic virus (TMV)tobacco mosaic virus (TMV), was fundamentally different from other, was fundamentally different from other
microbes and so simple and homogeneous that it could be cristallizedmicrobes and so simple and homogeneous that it could be cristallized
like chemical compoundlike chemical compound
1940 – Since the development of the electron microscope,1940 – Since the development of the electron microscope,
microbiologists have been able to observe the structure of viruses inmicrobiologists have been able to observe the structure of viruses in
detail.detail.

Recombinant DNA TechnologyRecombinant DNA Technology
Until the 1930s, all genetic research was based on the study of plant andUntil the 1930s, all genetic research was based on the study of plant and
animal cellsanimal cells
In the 1940s, the scientists turned to unicellular organisms, primarilyIn the 1940s, the scientists turned to unicellular organisms, primarily
bacteriabacteria
In 1941 –In 1941 – George W.BeadleGeorge W.Beadle andand Edward L.TatumEdward L.Tatum demonstrated thedemonstrated the
relationship between genes and enzymesrelationship between genes and enzymes
1944 –1944 – Oswald Avery, Colin McLeodOswald Avery, Colin McLeod, and, and Maclyn McCartyMaclyn McCarty  DNA wasDNA was
established as the hereditary materialestablished as the hereditary material
1946 –1946 – Joshua LederbergJoshua Lederberg andand Edward L.TatumEdward L.Tatum discovered that geneticdiscovered that genetic
material could be transferred from one bacterium to another by a processmaterial could be transferred from one bacterium to another by a process
called conjugationcalled conjugation
1953 –1953 – James WatsonJames Watson andand Francis CrickFrancis Crick proposed a model for theproposed a model for the
structure and replication of DNAstructure and replication of DNA

MICROSCOPEMICROSCOPE
Compound Light MicroscopyCompound Light Microscopy
Darkfield MicroscopyDarkfield Microscopy
Phase-contrast MicroscopyPhase-contrast Microscopy
Fluorescence MicroscopyFluorescence Microscopy
Confocal MicroscopyConfocal Microscopy
Electron MicroscopyElectron Microscopy

Compound Light MicroscopeCompound Light Microscope
Has a series of lenses and use visible light as its sourceHas a series of lenses and use visible light as its source
of illuminationof illumination
A series of finely ground lenses forms a clearly focusedA series of finely ground lenses forms a clearly focused
image that is many time larger than the specimens itselfimage that is many time larger than the specimens itself
This magnification is achieved when light rays from anThis magnification is achieved when light rays from an
illuminatorilluminator  condensorcondensor  specimenspecimen  objectiveobjective
lenseslenses  ocular lensocular lens
Total magnification = objective lens magnification xTotal magnification = objective lens magnification x
ocular lens magnificationocular lens magnification
Objective lens : 10 x (low power), 40 x (high power), andObjective lens : 10 x (low power), 40 x (high power), and
100 x (oil immersion)100 x (oil immersion)
Ocular lens : 10 xOcular lens : 10 x

The total magnifications would be 100 x for low power,The total magnifications would be 100 x for low power,
400 x for high power , and 1000 x for oil immersion.400 x for high power , and 1000 x for oil immersion.
Some compound light microscopes can achieve aSome compound light microscopes can achieve a
magnification of 2000 x with the oil immersion lens.magnification of 2000 x with the oil immersion lens.
The oil immersion has the same refractive index asThe oil immersion has the same refractive index as
glass, so the oil becomes part of the optics of the glassglass, so the oil becomes part of the optics of the glass
of the microscope.of the microscope.
Unless immersion oil is used, light rays are refracted asUnless immersion oil is used, light rays are refracted as
they enter the air from the slidethey enter the air from the slide

Darkfield MicroscopyDarkfield Microscopy
Is used for examining live microorganisms that either areIs used for examining live microorganisms that either are
invisible in the ordinary light microscope, cannot beinvisible in the ordinary light microscope, cannot be
stained by standard methods, or are so distorted bystained by standard methods, or are so distorted by
staining that their characteristics then cannot be identifiedstaining that their characteristics then cannot be identified
A darkfield microscope uses a darkfield condensor thatA darkfield microscope uses a darkfield condensor that
contain an opaque disccontain an opaque disc  the specimen appears lightthe specimen appears light
against a black backgroundagainst a black background
One use for darkfield microscopy is the examination ofOne use for darkfield microscopy is the examination of
very thin spirochetes, such asvery thin spirochetes, such as Treponema pallidumTreponema pallidum, the, the
causative agent of syphiliscausative agent of syphilis

Phase-Contrast MicroscopyPhase-Contrast Microscopy
Is especially useful because it permits detailed examinationIs especially useful because it permits detailed examination
of internal structures in living microorganismsof internal structures in living microorganisms
The principle of phase-contrast microscopy is based on theThe principle of phase-contrast microscopy is based on the
wave nature of light rays, and the fact that light rays can bewave nature of light rays, and the fact that light rays can be
in phasein phase (their peaks and valleys match) or(their peaks and valleys match) or out of phaseout of phase..
In phase-contrast microscopy, one set of light rays comesIn phase-contrast microscopy, one set of light rays comes
directly from the light source. The other set comes from lightdirectly from the light source. The other set comes from light
that is reflected or diffracted from particular structure in thethat is reflected or diffracted from particular structure in the
specimenspecimen
Phase-contrast microscope is provided with diffraction platePhase-contrast microscope is provided with diffraction plate

Brightfield Darkfield Phase-contrastBrightfield Darkfield Phase-contrast

Fluorescence MicroscopyFluorescence Microscopy
The object is stained with one of a group of fluorescent dyesThe object is stained with one of a group of fluorescent dyes
calledcalled fluorochromesfluorochromes ((primolin, acridine orange R,
thiazo yellow-G, auramine O, fluorescein isothiocyanate)
The light source  Ultraviolet light
The principal use of fluorescence microscopy is a diagnostic
technique called fluorescence-antibody (FA) technique, or
immunofluorescence

Confocal MicroscopyConfocal Microscopy
A fairly recent development in light microscopyA fairly recent development in light microscopy
Like fuorescent microscopy, specimens are stained withLike fuorescent microscopy, specimens are stained with
fluorochromesfluorochromes
The light sourceThe light source  laserlaser
Most confocal microscopes are used in conjunction withMost confocal microscopes are used in conjunction with
computers to construct three-dimensional imagescomputers to construct three-dimensional images
Can be used to evaluate cellular physiology byCan be used to evaluate cellular physiology by
monitoring the distributions and concentrations ofmonitoring the distributions and concentrations of
substances such as ATP and calcium ions.substances such as ATP and calcium ions.

Electron MicroscopyElectron Microscopy
Object smaller than about 0.2Object smaller than about 0.2 µm, such as virus, or theµm, such as virus, or the
internal structures of cels must be examined with aninternal structures of cels must be examined with an
electron microscopeelectron microscope
A beam of electrons is used instead of lightA beam of electrons is used instead of light
Objects are generally magnified 10,000 – 100,000 xObjects are generally magnified 10,000 – 100,000 x
Instead of using glass lenses, an electron microscopeInstead of using glass lenses, an electron microscope
uses electromagnetic lenses to focus a beam ofuses electromagnetic lenses to focus a beam of
electrons onto specimenelectrons onto specimen
There are two types of electron microscope :There are two types of electron microscope :
- The transmission electron microscope- The transmission electron microscope
- The scanning electron microscope- The scanning electron microscope

Introduction of microbiology kbk

Introduction of microbiology kbk

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