Parasitology: (Protozoa and Helminthes) - Mcst-1

Microbiology

Definition:


Microbiology is the science that deals with microorganisms that can not be seen

by the
naked eye
.


The Importance
of
Microbiology
:


1
-
The Environment
:
Microbes
are responsible for the cycling of carbon, nitrogen and


phosphorus
.


2
-

Medicine:

Production
of antibiotics.


3
-

Food:

Wine
and bread making, cheese and milk products production
.


4
-

Biotechnology:
Some
microorganisms have been used to synthesize many chemicals


such
as acetone and acetic acid.


5
-

Research:
Microbes
have been used as
model
organisms for the biochemical and


medical
investigations
.

What
is a Microbe?

A microbe or microorganism is:



A
-

A living creature which can not be seen except by microscope.



B
-

It is simple in structure.



C
-

Usually unicellular.



D
-

It can be either prokaryotic cell or a eukaryotic cell.




Microbiology

Nature

Range of size


Microbe


Cellular


20
-
300 nm

Viruses


Prokaryote


0.1
-
10
µm

Bacteria


Eukaryote

2 µm
-

1 meter

Fungi


Eukaryote

2
-

1000

µm

Protozoa


Prokaryotes and
Eukaryotes:

M
icrobes

Prokaryotes


Eukaryotes

Cell Structure

Simple structure

Complex structure

Nucleus

Primitive nucleus

True nucleus

Nuclear Membrane

Absent

Present

Nucleoli

Absent

Present

Mitochondria

Absent

Present

Enzyme System

Simple

Complex

Multiplication

Binary Fission

Mitosis

Examples

Bacteria, Rickettsia

Fungi, Protozoa

Microscopy:

It is the technical field of using a
microscope

to view
samples and objects
that can
not

be seen by unaided eye ( objects that aren't within the resolution range of normal
eye).


A
microscope

is the most

important
piece of

equipment
used
in clinical


microbiology.

Microscopy
forms 70
-
90% of

the
work
.






Microscopy

Working principle of a microscope:


A microscope is a magnifying instrument.



The magnified image of the object (specimen) is first produced by a lens close to
the object called
the
objective lens
.


Objective lens magnification = 4 X to 100 X


A
second lens near the eye called the
eyepiece

enlarges the primary image,
converting it into one that can enter the pupil of the eye
.


Eyepiece
magnification =
8 X to 12 X typically.





(but 10 X is most common)


Total
magnification = objective magnification* eyepiece magnification



(4, 10, 40, 100 X) * ( 10 X
)

So, final magnification ranges from 40 X up to 1000 X

Microscopy



Resolution of Microscope:


It is the ability to distinguish two
very small
and
very closed
objects as
separate

entities.

Resolution is
best

when the distance separating the two tiny objects is
small.


So, Resolution can be defined in a different way as:

It is the
smallest

distance that separate two sources of light points reflected from two
particles close together on object.


Resolution is determined by certain physical parameters that include the
wavelength
of light
, and the
light
-
gathering power
of the objective & condenser lenses.





Resolution
= light wavelength / 2


Example:

yellow light of a wavelength of 0.4
μm

give a resolution of
0.2
μm
.

Microscopy


Types
of microscopy:


1
-

The light microscope: (simple and compound).




A
-

The
student
(
Brightfield
) microscope





B
-

Phase contrast microscope.




C
-

Darkfield

illumination microscope.



2
-

The Fluorescence microscope
.



3
-

The Electron microscope
.

1
-

The Light Microscope:



-

Brieghtfield

microscope
is most likely found in classrooms or labs.



-

Better equipped classrooms and labs have
darkfield

&/or
phase contrast microscope
.




Microscopy



A
-
The
student
microscope:

Image quality is based largely on your ability to use the microscope properly.

Uses:

Brightfield

illumination microscope is mainly used to examine
stained
smear
Or

naturally
pigmented specimens.



Microscopy

Light path consists of:


Transillumination

light source
, commonly a halogen lamp in the
microscope
stand.


Condenser lens
which focuses light from the light source onto the sample.


Objective lens
which collects the light from the sample and magnifies the image.


Oculars

and /or a camera to view the sample image.


Objects seen in the light path because:


-

Natural pigmentations or stains absorb light differentially.

Or

-

They are thick enough to absorb a significant amount of light despite

being colorless.


Microscopy

B
-
Phase contrast microscope
:


This is needed to visualize
transparent

microorganisms
suspended
in a fluid.


(does
not
require staining to view the slide)


The phase microscope takes advantage of the fact that light waves passing
through
transparent slide
or objects (cells) emerge in different phases (different times).


A special optical system
converts

difference in phase into difference


in intensity. So that some

structure appear darker

than others
.

This microscope made it possible to

study
cell cycle.


Microscopy

C
-
Dark field illumination microscope
:

This method is used for visualizing organisms
suspended in fluid
and can
not
be
stained by Gram’s stain.

Both the
structure

and the
motility

of the organisms can be seen
.

Used for
: Unstained biological samples such as a smear from a tissue culture.

In this
method:
the light enters the special
condenser which
has a central
blacked
-
out
area
, so that light can not
pass
directly through it to
enter
the objective
.


Instead,
the light is reflected to pass

through the outer rim of the

condenser at a wide angle which


illuminates the microorganism


by a ring of light surrounding them.


Microscopy

Micrcoccus

species

examined by
phase contrast microscope

(
left
) and
Leptospira

examined by
Darkfield

microscope

(
Right
):

Microscopy

2
-

The
Fluorescence Microscope:

In this method, microorganisms must be stained with a
fluorescent dye
such

as
rhodamine
.


A fluorescent lamp emits visible light which is filtered off
using
optical
filters
.
(Ultraviolet (UV) wavelengths will not be filtered off).


A fluorescent dye will absorb this UV
-
light and change it to visible (yellow or
orange)
light
.


Microscopy

3
-
The
Electron Microscope:

-

Electron
microscope is an instrument that
magnify very small
objects which can
not be seen by light microscopy.



-

Resolution
of this microscope is higher than the light one.


-

The
energy source of this kind is the
electron beam
while light waves are used in
the light microscope.


-

Electromagnetic
lenses
are used instead of the glass lenses of light microscope
.



-

Visible
light of 500 nm wavelength give a resolution of 250 nm.

Electron
beam of 0.001 nm wavelength give a resolution of 0.0005 nm.


-
Viruses
with a diameter of 0.01
-
0.2 micrometer can be easily seen
.


-
Magnification of electron microscope is up to about 10.000.000 X .



Microscopy

Microscopy

Microscopy

Light waves and Electron beam: