This document discusses various techniques used to test antimicrobial activity, including disk diffusion, Etest, dilution plating, and determining minimum inhibitory concentration (MIC). The disk diffusion technique involves placing disks containing antibiotics on an agar plate inoculated with bacteria, and measuring the zone of inhibition. The Etest uses strips with an exponential gradient of drugs to determine the MIC. Dilution plating reduces bacterial concentration to obtain isolated colonies. MIC testing finds the lowest drug concentration that prevents bacterial growth. These techniques help determine antibiotic effectiveness and resistance.

1. MICROBIOLOGY
ANTIMICROBIAL TECHNIQUE
MEMBERS :
IMRAN KHAN SULAIMAN
CHE ABDUL HALIM
MUHD HUZAIFAH
NURUL ATIQA IBRAHIM
NADHIRAHTUL HUSNA
NOR ZAFIFAH
ROSE AMALINA

2. What is antimicrobial ?
• kills or inhibits the growth
of microorganism such
as bacteria
, fungi, or protozoa

3. kill microbes
(microbiocidal)
prevent growth of
microbes (microbiostatic).

4. antimicrobial substances
used on non-living objects
or outside the body.

5. Why antimicrobial technique is needed ?
• Bacteria have the ability to
develop resistance following
repeated or subclinical
(insufficient) doses
• so more advanced
antibiotics and synthetic
antimicrobials are
continually required to
overcome them.

7. Technique in testing for Antibiotic
sensitivity
• The method includes several steps including :
• obtaining a bacterial sample
• identifying the type of bacteria in the bacterial
sample
• selecting a set of antibiotics based on the
identity of the bacteria in the bacterial sample
• obtaining a control sample from the bacterial
sample

8. Antimicrobial technique .
..

9. Antimicrobial agent
placed on the
surface of an agar
plate
It containing
disease agent shows medium that has been
Disk inoculated with the
how effective the disease agent being
antimicrobial agent diffusion tested, which will grow
is. and fill the disk.
size of the area Antimicrobial agent
killing some of the diffuse into medium
cleared disease agent
around where the
anitmicrobial agent
was innoculated

10. For example . . .
• Small wafers containing
antibiotics are placed
onto a plate which
bacteria are growing.
• If the bacteria are
sensitive to the
antibiotic, a clear
ring, or zone of
inhibition, is seen
around the wafer
indicating poor growth
Antibiotic sensitivity testing

11. Zone sizes differ on sensitivity pattern
• It has been determined
that zones of inhibition
of a certain diameter
(varies for antibiotic
and to a lesser
extent, bacterial
species) correlate with
sensitivity or resistance
to the antibiotic tested

12. Steps for disk difussion method

13. Epsilometer test
• to determine whether or not a specific strain
of bacterium or fungus is susceptible to the
action of a specific antibiotic.
• commonly used in the setting of
medicine, where a particular organism has
been found to infect a patient, and the
doctor treating the patient is seeking
guidance on what concentration of antibiotic
is suitable.

14. rectangular strip
that has been
impregnated with
the drug
Etest utilise by then
A lawn of bacteria
an exponential
is spread and
gradient of the
grown on an agar
drug to be tested
Epsilometer plate
test
and
producing
Etest strip is laid
into the agar
on top
Drug diffuse out

15. As a result . . .
• an exponential scale
printed on the strip .
• After 24 hours of
incubation, an elliptical
zone of inhibition is
produced
• the point at which the
ellipse meets the strip
gives a reading for
the minimun inhibitory
concentration (MIC) of
the drug.

16. Dilution plating technique
• to reduce the concentration of
microscopic organisms or cells in a
sample
• A series of cultures is tested with
various concentration pf an antibiotic
to determine minimum inhibiting
concentration of antibiotic

17. spread bacteria over a
wide area
The numbers of bacteria
count the number of
colonies that grow on
colonies that grow
each plate are counted
bacterial cell in the
original sample should
produce a single colony

18. Stokes’ Method
• In original Stokes’ method
the inoculum of the control
strain is evenly spread over
the upper and lower thirds
of a plate
• the test strain over the
central third,uninoculated
(gaps 2 – 3 mm wide are left
to test from the control
areas.

19. For example . .
• In the Stokes controlled
sensitivity test, a control
organism is inoculated on part of
a plate
• the test organism is plated on
the remainder.
• Disks are placed at the interface
and the zones of inhibition are
compared.
• The use of a sensitive control
shows that the antibiotic is active
• if the test organism grows up to
the disk it may safely be assumed
that the test organism is resistant
to that drug.

20. Testing Minimum Inhibitory
Concentration
• In alternative measure of susceptibility is to
determine the Minimum Inhibitory Concentration
(MIC) and the Minimum Bactericidal Concentration
(MBC) of a drug.
• Both are mixed with serially diluted antibiotic
solutions and a standard inoculum is applied.
• After incubation, the MIC is the first broth in which
growth of the organism has been

21. What is Minimum
Inhibitory concentration
• Minimum inhibitory concentration (MIC), in is
the lowest concentration of an antimicrobial
that will inhibit the visible growth of a micro
organism after overnight incubation.
• Minimum inhibitory concentrations are
important in diagnostic laboratories to
confirm resistance of micro organisms to an
antimicrobial agent and also to monitor the
activity of new antimicrobial agents.

22. The Antibiotics are diluted to various dilution
to test the minimum inhibitory concentration