2. Infectious diseases
They are clinically evident diseases with the potential of
transmission from one person or species to another.
They result from the presence of pathogenic
microorganisms (very small organisms that are
invisible to the naked eye) that are able to cause disease
in human beings.
Pathogenic micro-organisms include bacteria, viruses,
fungi, protozoa, and multicellular parasites.
Practical microbiology sessions focus on the diagnosis of
these infectious agents.
3. Laboratory diagnosis of
infectious diseases
Direct method
Indirect (serologic) method
Detection of antibodies
against the microorganism in
the patient’s serum.
Detection of:
microorganisms,
their structural components
their products
in specimens collected from the patient (e.g.
urine, blood, sputum, CSF……etc).
4.
5. I) DIRECT METHOD
1)Specimen Collection
A ’good quality’ clinical specimen.
2) Microscopic Examination:
usually before further processing of specimens
3) Microbial Detection:
a) Culture technique:
b) Non-culture technique
6. I) DIRECT METHOD
1) Specimen Collection
A ’good quality’ clinical specimen
Collecting specimens before the start of antibiotics.
Choosing the appropriate specimen (representing the infection
site).
Using sterile containers and avoiding contaminating the
specimen.
Transporting the specimen properly to the lab. as early as
possible.
8. I) DIRECT METHOD
2) Microscopic Examination:
usually before further processing of specimens
stained /unstained (wet)preparations
different types of microscopes
Staphylococci in pus
9. I) DIRECT METHOD
3) Microbial Detection:
a) Culture Technique:
Isolation of the organism in pure culture
→inoculating the specimen onto appropriate artificial culture media
followed by
Identification of the isolate by e.g.
:
microscopic examination
biochemical reactions
reaction with specific antibody (serologic identification of the organism)
DNA probes
Which of these approaches is used and in what sequence depends upon the type of specimen and
organism.
Antibiotic sensitivity
After growing the organism in pure culture.
10. I) DIRECT METHOD
3) Microbial Detection:
b) Non-Culture Technique:
I.
Identification of a specific microbial antigen such as :
a structural component (e.g. cell wall antigen, capsular polysaccharide…etc) or
a product (e.g. an exotoxin)
by reacting with specific antibody
OR:
II.
Identification of a specific gene sequence
(i.e. nucleic acid of the organism)
by the application of different molecular methods (e.g. PCR, DNA probe).
11. I) DIRECT METHOD
yield more rapid results (minutes or hours)
(do not depend on growth and multiplication of the organism)
However, antimicrobial susceptibility cannot be determined
(although the presence of resistance genes can be determined by
molecular methods).
Non-culture techniques are mainly applied if:
a rapid diagnosis is needed
The microorganism cannot be cultured on artificial
media
A slowly growing micro-organism
12.
13. II) INDIRECT (SEROLOGIC)
METHOD
Serologic diagnosis of infectious diseases involves the use of known
microbial
antigens
to
microorganism
detect antibodies
in the patient’s serum.
current (active) infection
against the
is diagnosed by the
detection of one of the following
specific IgM antibodies
rising titre of specific IgG antibodies (4-fold or greater rise)
a single high titre of IgG antibodies in certain diseases
Skin tests
N.B.
based on cell-mediated hypersensitivity may also
help in the diagnosis of certain diseases.
15. Diagnosis of infectious diseases
Diagnosis of infectious diseases
DIRECT METHOD
DIRECT METHOD
Specimen
Specimen
Microscopical ex.
Microscopical ex.
Culture technique
Culture technique
Isolation on culture media
Isolation on culture media
.Identification e.g
.Identification e.g
••microscopical.ex.
microscopical. ex.
••bioch.Reactions
bioch. Reactions
••DNAprobe
DNA probe
••serology
serology
INDIRECT METHOD
INDIRECT METHOD
(SEROLOGICAL)
(SEROLOGICAL)
Detection of antibodies in serum
Detection of antibodies in serum
••IgM
IgM
••risingtitre of IgG
rising titre of IgG
Non-culture technique
Non-culture technique
••detection of specific antigen
detection of specific antigen
(serology)
(serology)
•• detection of specific gene
detection of specific gene
sequence (mol. tech.)
sequence (mol. tech.)
Antibiotic sensitivity
Antibiotic sensitivity
28. Staining of Bacteria
Bacteria cells are almost colorless and
transparent
A staining technique is often applied to the
cells to color them →
Their shape and size can be easily
determined under the microscope.
28
32. Types of Stains
1- simple stain:
Single basic dye e.g. Methylene blue
All bacteria take the color of the dye
2- Differential stain:
Two dyes
separated by a decolorizing agent
e.g. Gram stain
& Ziehl-Neelsen stain
3- Special stain: e.g. Fontana stain
33. Differential staining
Principles of differential stain
* Application of the main stain.
• Decolourization.
*Application of the counter-stain.
e.g. Gram stain & Ziehl-Neelsen
stain