2. 1 DEFINITION OF MICROIOLOGY
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DEFINITION OF MICROORGANISM
CLASSIFICATION OF MICROORGANISM
STRUCTURE OF BACTERIA & VIRUS
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ISOLATION OF PURE CULTURE
STAINING OF BACTERIA
TYPES OF STAINING
3. DEFINITION OF MICROBIOLOGY
The word Microbiology is derived from-
Micros – Small
Bios – Life
Logos – To study or science
Microbiology is the branch of science deals with the study of living micro-organisms.
Eg., Bacterial, Virus, Fungi etc.
The study includes structure, function, reproduction and multiplication of micro-organisms.
4. DEFINITION OF MICRO-ORGANISMS
These are the small groups of living organisms which can not be seen by
naked eyes and studied under microscope.
Eg., Bacteria like streptococcal, pneumococcal, salmonella typhi.
Virus like DNA or RNA virus, HIV.
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6. BACTERIA
These are the member of a large group of unicellular
(Prokaryotic) microorganisms which have cell wall
but lack of cell organelle like Golgi apparatus,
Mitochondria etc.
Size: - 1 – 5 micron.
Shape: - Cocci, Spherical, Rod-shaped, Spiral, Thread
shaped…..
7. VIRUS
These are non cellular, ultramicroscopic highly infectious agent and posses only one type
of nucleic acid either DNA or RNA surrounded by protein (protective) coat called, Capsid.
Size- 0.02 to 0.2 micron.
Shape – As in image
9. Isolation of Pure Culture
Growth of microbes on laboratory medium is known as Culture. A culture which may contains only one
species of microbe is called a pure culture and one which consist of several species is called mixed
culture.
It is very difficult to obtain pure culture of bacterias in nature because they exist as mixed culture. To obtain
organisms in pure culture various techniques are used-
1. Streak plate method
2. Pour plate method
3. Spread plate method
4. Micromanipulator
5. Roll tube method
10. Staining Methods
Stains are the organic dyes used for staining the micro-organisms.
For ex, Crystal violet, methylene blue, safranin etc.
Purpose of Staining: -
1. For greater visualization of cells.
2. For study of their structures.
3. To differentiate the cells.
4. To inhibit the growth of some organism so the others can be visualized.
12. Simple Staining
It is also called as Monochrome technique. In this method only one stain is used. It used to study morphology i.e.
size, shape and arrangement of microbes.
Prepare a smear and fixed on slide
Add stain for 30 sec to 3 min using
methylene blue
Wash with cool water
Air dry and examine under oil immersion
lens
13. Differential Staining
Gram’s Staining: -
A differential staining technique used to classify bacteria i.e. gram positive or gram negative and their specific
structure.
Gram staining was discovered by a Danish Physician “Hans Christian Gram” while working in Berlin in
1883 and later procedure published in 1884. Hence, it is called Gram’s staining.
Requirements: - Staining reagents like
1. Crystal violet- Primary Stain
2. Gram’s Iodine- Mordant - fixative agent
3. Acetone 95% or Alcohol- Decolorizer
4. Saffranine / dilute carbol fuchsin counter stain
14. Gram’s Staining
Crystal Violet: - All bacteria takes crystal violet so all are appears violet colour.
Iodine: - Crystal violet-iodine (CV-I) complex is formed.
Acetone: - Bacteria with high lipid content loose CV-I complex and appears
colourless but bacteria with less lipid content retains CV-I complex and appears
violet.
Saffranine: - Only colourless bacteria takes saffranine and appears pink
16. Observation
If the bacteria shows violet or
purple then indicates Gram
Positive.
Eg., E. Coli, Pneumococci
If the bacteria shows pink or red
then indicates Gram Negative.
Eg., S. typhi, H. Influenzae
17. Acid fast staining technique
This technique was discovered by the scientist Zeihl & Neelson,
This technique is used to identify all the separation of Mycobacterium Group members from the others.
It means it is used to identify the acid-fast micro-organism like Mycobacterium tuberculosis,
Mycobacterium Leprae etc.
Acid –fast microorganism are characterized by wax like nearly impermeable cell walls means they
contain mycolic acid and large amount of fatty acids, waxes and complex lipids.
Acid-fast organisms are highly resistant to disinfectants because of the cell wall is so resistant to most
compounds. Acid-fast microorganism required a special staining technique i.e. called Acid-fast staining
method.