1. LECTURE: Physical Methods of Control
MICROBIOLOGY – ALCAMO
LECTURE:
Physical Methods of Control

2. Incident at the Pool It was July, 1988 in Los Angeles, Ca
A person unintentionally defecated in a pool
The pool was later used by a high school water polo team, a SCUBA diving class, and an elementary school field trip

3. Incident at the Pool
In early August, pool users began to go to their doctor complaining of watery diarrhea, stomach cramps and fever
11 patients were tested for MO’s in their stool and it was found that they had the protozoan Cryptosporidium
CDC investigators found
that although chlorination
was OK, the filters being
used were not functioning

4. Incident at the Pool
This shows how MO’s can spread if proper water purification techniques are not used
Sterilization is the destruction or removal of all life forms
Something can not be partially sterilized, it is all or nothing
It is considered contaminated until sterilized
Even if something is sterilized, it can still be harmful if the MO gave off a toxin

5. How Does Sterilization Kill Cells??
Denaturation
Breaks H bonds in RNA, DNA, enzymes
Coagulation of proteins (cooked eggs)
Irreversible

6. How Does Sterilization Kill Cells??
Oxidation
Large molecules break into small molecules, loss of electrons
Cell walls, membranes hit first

7. How Does Sterilization Kill Cells??
Antimetabolite
Interferes with normal cell reactions
It is a molecule that is similar enough to a natural molecule used in an organism to interfere with the molecule’s function
Antibiotics are antimetabolites as well as cold temperatures

9. Principal Methods of Sterilization
Physical Agents:
heat
radiation
filtration
Aim of sterilization is destruction of bacterial spore
most resistant forms of life
carry on life processes at minimum rate
possess enzymes to transform from dormant state

10. Control With Heat Heat has a killing effect on MO’s
It is fast, reliable, inexpensive and does not introduce any chemicals
MO’s have an optimal growth temperature
Growth slows down both
above and below this
temperature

12. Effects of Heat
Biochemical changes in organic molecules result in cell death
Enzyme structure altered (can’t perform basic life functions)
Structural molecule breakdown (cell membranes fall apart)
Water loss can result in cell death
Organisms depend on water
Excessive heat dehydrates them

13. Heat Killing Rate Function of both time and temperature Each MO has:
Thermal death time – time necessary for killing it at a specific temperature
Thermal Death Point – the temperature at which it dies in a given time

14. How to Determine Time and Temperature
Type of organism to be killed:
To sterilize materials – direct method at bacterial spores
To prepare milk – no need to sterilize, just heat to kill off vegetative cells
Material to be treated
Powders – use dry heat
Solutions – use moist heat

15. Temperature High - 100˚C and above
Direct flame: Loop in a Bunsen burner
Incineration: Fast oxidation, kills all, +/- 1,000˚F long enough to burn up
Example: disposable hospital gowns, cows that died of anthrax
Cautions: Aerosols & Completeness
Hot air oven: Slow oxidation, kills all, dry heat, 160˚C for 2 hours
Example: powders, glass pipettes and syringes
Non-burnables only

16. Temperature High - 100˚C and above Boiling Water:
moist heat penetrates materials more quickly than dry heat
lower temperatures and less time required
works by denaturing proteins
Not considered a sterilizing agent because can’t always be sure spores and viruses are inactivated
Minimum exposure of 30 minutes

17. Autoclave Moist heat – pressurized steam
Most dependable method for destruction of life forms
When the pressure of a gas (steam) increases, the temperature of the gas increases proportionally (under 15 lbs pressure, the water temp goes up to ˚C in 15 minutes)
Reduces cooking time in a pressure cooker and sterilizing time in an autoclave
Broadest professional application: Food, medical, labs, research

19. Autoclave
Moist heat denatures proteins

20. Temperature Medium - Below 100˚C, selective kill, not sterile
Pasteurization:
Aimed at pathogens
Reduces # of MO in a liquid and destroys organisms that could cause spoilage or disease
Spores are not affected by pasteurization
Fractional Sterilization (Tyndallization)- Repeated heating 3X with time for spore generation in between
Holding Method - 63˚C/30 minutes
Flash Pasteurization - 72˚C/15 seconds
Ultrapasteurization - 82˚C/3 seconds

21. Heat sensitive liquids:
Milk
Wine
Beer

22. . Low Temperature Refrigeration: Antimetabolite
Static Measure Only – does
not kill MO’s
Warm up and growth recurs
Food Spoilage implications
1-7˚C, Lowers MO’s enzyme reaction
rates (watch out for psychrophiles)
Freezing: 0˚C and Below, same as refrigeration

23. 2. Water No water = no chemical reactions Antimetabolite
Cidal or Static – Time & Organism Mycobacterium (TB) – Months Treponema (Syphilis) – Minutes Spores – Years, Decades

24. 2. Water Desiccation = Water deprivation by drying out
Osmosis = Water deprivation by chemical means

25. 3. Radiation Electromagnetic waves that denature and kill
Short: “X rays” & Gamma rays
Also called ionizing radiation
Penetrate objects well
Human cells very sensitive
Drugs, medical & dental supplies
No apparent residual effects

26. 3. Radiation

27. 3. Radiation Long: Ultraviolet (100-400 nm)
No penetration, surface only
Can damage retina of eye
Sterile culture rooms, O.R.
No one around or special goggles

28. 4. Sound Ultrasonic vibrations “Sonication” Limited use:
High frequency sound waves
When used in fluids – causes formation of microscopic bubbles or cavities and water appears to boil
These cavities easily collapse and send out shock waves
MO’s in fluid are disintegrated by external pressures
Limited use:
Dental – clean instruments
Research – breaking open cells

30. 5. Filtration Mechanical device for removing MO’s from a liquid
MO’s get stuck in pores of filter and rest of fluid is decontaminated or sterile
Size of unwanted particles must be known
Used to purify IV solutions, Bacterial media, pharmaceuticals, beverages
Types of Filters:
Inorganic – porcelain or glass
Organic – diatomaceous earth
Membrane – cellulose
HEPA – air filters

32. Terms Cidal – kills Static – controls
Disinfectant – applied to inanimate object
Antiseptic – applied to living tissue
Sterilization – killing of “all” mo’s
Toxin may exist