Methods to Control Microbial Growth
Lab. 11
• 1. Physical
• 2. Chemical
The effect of disinfectant and antiseptics
on the growth of bacteria
• Objective: To understand the principle of disk
diffusion test.
• To be able to read a disk diffusion test result.
• To understand the concept of zone of
inhibition
• Disinfectant vs. Antiseptic
• Disinfectant
– antimicrobial agent used on inanimate objects
• Antiseptic
– antimicrobial agent used on living tissue
Controlling Bacteria
Antiseptic vs. Disinfectants
Antiseptic- chemicals that kill
bacteria on living things
•means – “against infection”
Examples: iodine, hydrogen peroxide,
alcohol, soap, mouthwash
Disinfectants- stronger chemicals that
destroy bacteria on objects or nonliving
things
Disinfectants
Example:
• Dettol
• Clorox
• Iodine
• Alcohol
• Flash
• Jif
• vim
Antiseptics
Example:
• Dettol
• Alcohol
• Iodine
• Tooth past
• Mouth wash
Procedure
1.
2.
3.
4.
Take sterile nutrient agar plate and label
Surface streak the plate with the given culture
Make four holes in the agar using a cork borer
Add 2-3 drops of the chemical agent in the
labeled hole
5. Incubate the plate at 37C for 24-48 hours
Cloro
Dettol
D
C
A
M
Cork borer
Alcohol
Mouth wa
Result
The bacteria will start to grow ,then it will
stop growing when it is effected by the
disinfectants, and will have a clear zone
These results depend on :
1- the kind of bacteria .
2- the kind of disinfectant or antiseptic.
• Conclusion:
 The larger the zone ,the stronger the effect of
disinfectant
 The smaller the zone ,the weaker the effect of
disinfectant
 No zone indicates no effect.
Antibiotic Disk-Diffusion Method
• Also called Kirby-Bauer method.
Purpose:
• It is used to detect the susceptibility and
resistance of test bacteria to a given antibiotic
with known concentration.
Procedure
• The suspension of tested bacteria is swabbed
on the surface of a solid culture media, and
disks of different antibiotics which contain
known concentration of these antibiotics are
placed on the swabbed culture and incubated
18-24hr.
• The antibiotics will diffuse from the disk into
the media and according to the sensitivity of
the bacteria to a given antibiotic.
The effect of antibiotics on the growth of
bacteria
• There are different antibiotics that have different
effects on the growth of bacteria
• This experiment also depends on:
– The kind of bacteria
– The kind of antibiotic
• Example: penicillin ,Amoxilline ,Streptomycin
,chloramphenicol,Tetracycline
The bacteria will start to grow ,but around the
antibiotic disks , there is no growth making clear zones
with different diameters in length.
Results
• If the bacteria is susceptible to the antibiotic (sensitive):
the antibiotic kills or inhibits their growth and a zone of
inhibition (no growth of the bacteria) will formed around
the disk.
• If the bacteria is resistant to the antibiotic: no zone of
inhibition will formed around the disk (there will be a
growth of bacteria all over around the disk).
• The diameter of the zone is measured by roller, and the
reading will be compared with the reference range of this
bacteria to this antibiotic and then the one can judge
wither the bacteria is susceptible or resistant
conclusion
 The larger the zone the stronger the effect of
antibiotics
 The smaller the zone the weaker the effect of
antibiotic
 No zone indicates no effect
Physical methods for controlling microbial growth
Controlling microbial
growth
Heat
Filtration
cold
Heat
Protein denaturation
By Heat
Moist heat
Pasteurization
72c for 15 Sec.
Dry heat
Boiling or steam flow
&
Autoclaving at 121c
For 15 min.
Milk, cream,&
Alcoholic drinks
Direct flaming
Incineration
Hot air 170c,2hr.
Pasteurization
Dry Heat
• Direct flaming: for loops& needles
Filtration
• Separation of bacteria from the suspension or
liquid.
• For sterilizing Enzymes ,Vaccines ,heat
sensitive media
Filtration
Cold: for controlling &preserving microbial growth
cold
Refrigeration
Of food culture
& drugs
Deep freezing
-50 to-95
Food, culture,&
drug
Lypholization
Food, drugs,&
culture
Most effective for
Short preservation.
Most effective for
Long preservation
Refrigeration
Deep Freeze
Lypholization
• Is the removal of water by high
vacuum under very low
temperature.