Nama : Dr.Sadeli Masria, dr., DMM., MS., SpMK.

Pekerjaan : Kepala Bagian Mikrobiologi

Fakultas Kedokteran Unpad
Alamat : Jl. Saturnus Tengah No. 16
Bandung
Tilp. : 022 – 7567136
HP. 08122126320
1. HISTORY OF MICROBIOLOGY
2. MICROBIAL WORD
3. MICROSCOPY AND STAINING
PROCEDURES
4. MICROBIAL ECOLOGY
 HISTORY OF MICROBIOLOGY
DEVELOPMENT OF MICROBIOLOGY
- is the branch of biology
- cannot be seen with the naked eyes
- were first seen by the Dutchman Antony van
Leeuwenhoek (1675)
- The prototype of the experiment was first
described by Louis Joblot (1718), and John
Needham (1749)
- Robert Koch in 1877 was described methods for
the easy microscopic examination of
bacteria, in 1881 devised the simple method
for isolating pure cultures of bacteria
- Robert Hooke and Anton van Leeuwenhoek are the first
investigators to use the microscope
 MICROBIAL WORD
 Organism on Earth may be catagorised either
as Eukaryotes and Prokaryotes

 Both terms are derived from the Greek word

karyon, which mean “nut”
In the case, the nut refers to the nucleus
of a cell

 Prokaryotes : are organisms whose cells lack a

nucleus
 Eukaryotes : are organisms whose cells have a
well defined nucleus
The cells of Eukaryotic organisms, contain :
- Nucleus
- Organelles, suspense in the cytoplasm
- Complex phospholipids, spingolipids, histone,
sterol

Nucleus :
- separated from the cell cytoplasm by a nucleus
membrane
- contains the genetic information of the cell
- is the largest body in the karyotic cell

The nucleolus (= a smaller body), exists within the

nucleus and play a role in the production of
ribosomal RNA and ribosomes
Organelles :
- Mitochondria
- Ribosomes
- The Endopalsmic Reticulum
- The Golgi apparatus
- Lysosomes
- The cell membrane
Many familiar organism are Eukaryotes :
- Animal, including humans
- All plants
- Protozoa
- Fungi, - may be monomorphic
- may be dimorphic : yeast or mold
- have both asexual and sexual reproduction
capabilities
- have a growth cycles that consist of both a
vegetative and a reproductive
The cells of Prokaryotic organisms
- lack a nucleus or nucleus membrane
- prokaryotic cells possess a single chromosome
composed solely of DNA
- Have no organelles, exception of ribosome
- No membrane enclosed nucleus
- No histone
- In rare cases, they contain complex phospholipid,
sphingolipids and sterol
- Have a cell wall composed of peptidoglycan
containing muramic acid
- The chromosome exists as a closed loop
- Contain ribosome
Several types of microorganism are
prokaryotes :

- The bacteria
- Mycoplasma
- Rickettsia
- Chlamydia
 BACTERIA
 The most abundant organisms on earth

 Normal flora or pathogenic in human

 Do not have a sexual growth cycles

 Most species of bacteria are hundreds of time large

than viruses

 The great of species can be seen with an ordinary light

microscope at a magnification of 1000 X
Three basic forms of bacteria
* Bacillus
e.g : - Bacillus anthracis
- Corynebacterium diphtheriae
- Clostridium tetani
Several species of bacilli that occur in chains, called
Streptobacillus

* Coccus
e.g : - Streptococcus pneumoniae
- Neisseria gonorrhoeae

* Spiral
e.g : - Vibrio cholerae
- Spirillum minus
- Treponema pallidum
VIRUSES
 The smallest microorganisms
 Composed of nucleic acid, surrounded by protein coat
 Are not cells and are not visible with the light
microscope, require electron microscope for
visualization
 Are obligate intracellular microorganisms
 Contain either RNA and DNA as genetic material
 Are called bacteriophage (or phage) if they have a
bacterial host
 Viruses do not grow, which can be accomplished only
within living cells
Viruses occur in three major shape :

- The icosahedron
- The helix (or coil)
- The complex form
Despite their structural simplicity, viruses are
responsible for several important human
diseases including influenza, hepatitis,
chicken pox, acquired immune deficiency
syndrome (AIDS)
CYANOBACTERIA
- formerly called blue-green algae, but now called Cyanobacteria
- are microorganisms that process pigments which function in the
solar energy tripping process of photosynthesis
- may occur in celled (unicellular) or multicellular filamentous
forms
- They inhabit fresh water and marine environtment, sometimes
giving a “pea soup” color to the water

ALGAE
- like cyanobacteria, have pigment, whith their cells and are
considered microorganisms
- The pigment of this algae are more like those of plants, however,
and have a complexity typical of plant cells
- Two type of microscopic algae are important in microbiology
+ The diatoms
+ The dino flagellates
FUNGI
- are complex microorganisms subdivided into two groups the
MOLD and YEASTS
- many mold prefer acidic environment such as citric fruits,
cheeses and bread
- Yeasts are single celled fungi
- important in bread production and fermentation of juices to
produce wine

PROTOZOA
are microorganism with complex cellular features

VIROIDS
- are not cells and are not visible with LM
- are obligate intracellular parasites

PRIONs
are infections particles associated with subacute progressive,
degenerative disease of the CNS, e.g. Creutzfeldt-Jacob disease
MICROSCOPY AND STAINING PROCEDURES
MICROSCOPE
1. LIGHT MICROSCOPE

* Uses two sets of lenses : - Ocular lens :

- Monocular
- Binocular (paired lenses)

- Objective lens :
- Low power 10 X
- High power 40 X
- Oil immersion 100 X
* The stage
* Clips
* Condensor
2. DARK FIELD MICROSCOPE

- The condensor does not allow light to pass directly

through the specimen
- The background a dark field
- Utility for detecting certain bacteria directly in
patient specimens
- because of their thin dimensions, cannot be seen
by light microscopy
- because of their physiology are difficult to grow in
culture
- Dark field microscopy is used to detect spirochetes
e.g Treponema pallidum
3. ELECTRON MICROSCOPE
- The great technological advances of the century
- First developed and made commercially available
in the 1930 s
- uses accelerated electrons that have a wavelength
a fraction of the wavelength of light
- The result is an image magnified tens and
hundreds of thousands of times
- There are several types of electron microscope
available today for use in microbiology
*The transmission electron microscope
(TEM)
* The scanning electron microscope (SEM)
The Scanning Electron Microscope (SEM)
- was developed in the 1960 s
- is easier to operate than the TEM
- Produces a three dimensional effect permits the
scanning of whole and sectioned objects without
extensive preparation
- Permits the visible surface of the specimens to be
studied in vivid detail
- Usually results in relatively low magnification
(under !00.000) times
 The Gram Stain
 In 1884, Hans Christian Gram, a Danish doctor
working in Berlin, accidentally stumbled on a
method which still forms the basis for the
identification of bacteria
 While examining lung tissue from patients who had
died of pneumonia, he discovered that certain
stains were preferentially taken up and retained by
bacterial cells
 Over the course of the next few years, Gram developed
a staining procedure which divided almost all
bacteria into two large groups - the Gram stain
PRINCIPLES OF GRAM STAINING
The difference in composition between
Gram positive Gram negative

Cell walls Thick peptidoglycan with numerous Thin layer of peptidoglycan

teichoic acid cross linkages

Lipid Low High

concentration
ACID FAST STAINS
- Specifically designed for a subset of bacteria whose
cell walls contain long chain fatty acids.
- The classic acid fast staining method Ziehl-Neelsen
- Mycobacteria are the most commonly encountered
acid fast bacteria
- The procedure requires heat to allow the primary stain
(carbol fuchsin) to enter the wax containing cell wall.
ACID FAST STAIN PROCEDURE AND PRINCIPLES

Acid fast (+) bacteria Steps for staining Acid fast (-) bacteria

1. Cells on slide
2. Primary stain
(carbol fuchsin red)
3. Decolorizer
(HCl, alcohol)
4. Counter stain
(methylene blue)
acid fast positive organisms stain red
- Depending on the type of counterstain used (e.g methylene blue
or malachite green), other microorganisms, host
cells and debris stain a blue to blue greem color.
- The Kinyoun acid fast method, is a modification of this
procedure.
MICROBIAL ECOLOGY
PREDOMINANT NORMAL FLORA AT VARIOUS BODY SITES

Body sites Microbial flora

Skin S. epidermidis, Corynebacterium,

Propionibacterium, micrococcus, yeast
Conjunctivae S. epidermidis
Nose and nasopharynx S. epidermidis, S. aureus, Streptococcus sp.
Mouth and oropharynx S. epidermidis, S.pneumoniae, S. mitis,
S. salivarius, Neisseria, Haemophillus, Veilonella,
Bacteroides, Fusobacterium, Lactobacillus, yeast
Intestinal tract :
Small intestine Lactobacillus, Streptococcus spp., Enterococcus,
Veillonella, Actinomycetes, yeast
Large intestine Bacteroides, Clostridium, Fusobacterium,
Eubacterium, Bifidobacterium, Lactobacillus,
Peptostreptococcus, Enterococcus,
Enterobacteriaceae
Genitourynary Tract Corynebacterium, alpha and nonhemolytic
Streptococci, S. epidermidis, Enterococcus,
Lactobacillus, Mycobacterium smegmatis,
Enterobacteriaceae, Bacteroides, Fusobacterium
Blood and Cerebrospinal fluid Sterile
Tissues, Bladder, Uterus and Sterile
fallopian tubes, middle ear and paranasal sinuses
THE HOST AND MICROORGANISMS RELATIONSHIPS
- Are always bidirectional in nature.
- Humans dominate the relationship because microorganisms are used
by humans in various settings,
including :
- the food and fermentation industry,
- as biologic insecticides for agriculture,
- genetic engineering a multitude of products and
- even biodegrading our industrial wastes.
General Stages of Microbial - Host Interaction
General Stages of Microbial - Host Interaction
Physical
General Stages encounter
of Microbial - Host Interaction
between host and
Microorganism

Microorganism
colonization of host
surface (s)

Microorganisms,
entry invasion and
Dissemination

Outcome
 MICROBIAL RESERVOIRS AND MODES OF
TRANSMISSION TO HUMANS

Microorganism Sources (Reservoirs) :

Humans, Animals, Food (Plant and Animal Sources)
Water, Air , Soil

Modes of Transmission :
Direct, Transmitted by direct contact between Reservoir and Host
Indirect, Transmitted to Host via Intervening agent

Intervening Agents :
Vectors : Animals, Insects, Other humans
Vehicles : Water, Food, Air, Medical devices, etc