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Sterilization &

Disinfection

By

Eman A.Abd AlrahmanAss. Lecturer of Microbiology

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Sterilization is the killing of all living forms

of microbes including their spores.

Disinfection is the destruction of most but not

necessarily all pathogenic microbes or their

spores

Decontamination is a term applied to any

procedure that reduces pathogenic

microorganisms to a level where items are safe

for handling, for use or disposal. It can be done

by cleaning, disinfection or sterilization.

.

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Cleaning is a process that removes foreign

material (e.g. dirt, organic material, and some

microorganisms). Cleaning must precede

disinfection and sterilization. It is usually done

with soap and water, detergents or enzymatic

products.

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Method of Sterilization

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A- Sterilization by Heat

1-Dry heat:- kills by destructive oxidation of

essential cell constituents. It is less efficient than

moist heat. However, it is less expensive and is not

corrosive

a) Red heat : bacteriological loop & forceps

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b) Flaming : used for the mouth of tubes and

glass slides.

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c) Incineration: used for contaminated

materials by burning in a special incinerator.

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d) Infra red radiation : used for surgical

instrument, temp: 180 °C.

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e) Hot air oven•Composition :isolated double

walled chamber electrically

heated, provided with

thermotstate and a fan to

ensure rapid and controlled

heating .

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•Tempreture:160 °C

•Holding time :1.1/2 hour (after temp reach 160

°C

•Used for: sterilization of glass ware, ointments,

powders, oils and metallic instruments.

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Moist heatUnder 100 oC• Pasteurization•Inspissation

At 100 oC• Boiling• Tyndallisation.

Over 100 oC •Autoclaving

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Moist heatkills bacteria by protein denaturation.

a)Temperature below 100 °C

1- Pasteurization: as pasteurization of milk

by heating at 63oC for 30 min. or at 72 °C

foe 20 sec. and immediately cool below 10 °C

• This kills non spore forming milk borne

pathogens e.g. M.tuberculosis, Br.abortus,

Salmonella and Coxiella burnetti.

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2-Inspissation: Heating at 85 ̊C for 30 min,

for 3 successive days.

•Used for sterilization of media containing

egg.

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b) Temperature of 100 oC1-Boiling:Used for disinfection of

surgical and medical equipment

when sterility is not required.

Boiling occurs at 100 °C for

20 min. and enhanced by

adding 2% Na bicarbonate.

Kills all vegetative bacteria

and HBV but not spores.

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2-Sterilization by tyndalization (heating at

100oC for 30 min, for 3 successive days)

• At first day vegetative bacteria are killed and

spores germinate to vegetative which will be

killed in 2nd and 3rd days.

• Used for sterilization of media containing

sugars or gelatin.

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c)Temperature above 100°C (Steam sterilization or autoclaving):Principle: When heating water in a closed vessel,

boiling point rises above 100°C.

Conditions of sterilization: Heating at 121°C under

2 atmospheric pressure for 20-30 min. or at 134°C

under 3 atmospheric pressure for 3-6 min. (flash

autoclave)

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Autoclaving is the most efficient method of sterilization.Why?

High temp.

High penetration power of steam under

pressure.

Condensed steam liberates large amount of

latent heat.

Not toxic, inexpensive and rapid.

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Monitoring of autoclaves:1- Mechanical methods: monitoring time, temperature

and pressure of sterilization cycle.

2- Chemical methods:

Bowie Dick tape is applied to articles being autoclaved. If

the process has been satisfactory, dark brown stripes will

appear across the tape. Pale brown stripes are suggestive

of poor heat penetration, and an unsatisfactory

sterilization process.

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3-Biological indicator:

Geobacillus stearothermophilus spores strips may be

placed inside the autoclave at the start of its cycle.

After running, the autoclave the strip is recovered

and cultured. Absence of growth after a suitable

period indicates a successful run.

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Types of Autoclave

Automated autoclave Simple autoclave

Flash autoclave

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Sterilization by radiation

• Ultraviolet rays

• Poor penetration power

• Source: mercury vapor lamp

• Sterilization of air and water

• Can cause eye damage

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• Ionizing radiation

• High penetration power

• Source: radioactive isotope as cobalt 60, β

rays and gamma rays

• Sterilization of materials not stand heat as plastic

syringes, gloves, catheters.

• Monitoring: By spore indicator (B. pumilus)

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Filtration• It is a mechanical method for exclusion of organisms from biological fluids which are destroyed by heat e.g. serum, plasma, hormones, vitamins and fluid culture media. Also sterilization of air in operation rooms, drug filling cubicles.• It depends on use of filters with pore size smaller than that of bacteria so retain them.

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1)Vacuum filter(Seitz):

Used for sterilization of

large volumes of fluids.

suction is applied to side

tube to force fluid to pass

through the filter.

Types of filters:

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3-Millipore filter:

Suitable for sterilization of

small amounts of fluids.

4- Air filters:

HEPA (high efficiency particle arrester)

filter.

Used to decontaminate air input to

operation rooms and air withdrawn

from areas of dangerous pathogens.

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Gaseous sterilization

a)Ethylene oxide (EO)

• Ethylene oxide bactericidal & sporicidal

• capable of killing spores rapidly by interfering with

the normal metabolism of protein and reproductive

processes, resulting in death of cells.

• Sterilization of heat sensitive devices e.g. plastic

and rubber articles, heart lung machines.

Chemical sterilization

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Sterilized articles exposed to gas at 55 °C for 3 –6 h then aerated for 8-12 h.• Bacillus atrophaeus spores are used for monitoring EO

Disadvanteges

Toxic , explosive and carcinogenic requiring safety

precautions .

Colorless gas ,very toxic and flammable .

Requires special equipment.

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Materials must be well aerated after sterilization .

takes longer time than steam sterilization.

Long-term exposure to EO is potential

occupational carcinogen causing leukemia.

Over exposure cause nasal and throat irritation

Prolonged exposure may result in nausea, vomiting

dizziness, difficulty breathing and peripheral paralysis.

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Sterilization by ethylene oxide (EO)

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Gas plasma• Plasma: any gas consists of electrons, ions or neutral

particles

• Hydrogen peroxide alone or with peracetic acid

are injected into sterilization chamber under

pressure to evaporate and surround items to be

sterilized then radiofrequency energy is applied

which excites Hydrogen peroxide and produce

plasma containing free radicals which are lethal to

bacteria.

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Uses: Sterilization of surgical instruments with long narrow lumen as laparoscopes

Advantages of Gas plasma

1. The process is dry and nontoxic; items do not need

to be dried or aerated.

2. Low temperature allows safe sterilization of some

heat sensitive items, including endoscopes and fiber

optics.

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4. Plasma has significantly less effect on metal

than steam sterilization corrosion does not

occur.

5. The sterilizer is simple in design, and

connects to standard

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Gas Plasma Sterilizer

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OthersUsed for Examples

Clinical thermometer& skin antiseptic Ethyl alcohol 70%

Skin antiseptic Iodine (tinc.iodine or betadine)

Water and swimming pool cholrine

Wound antiseptic Hydrogen peroxide

endoscope Glutraldehyde

Wound infection Salts of heavy metalsmercucrome

Culture decontamination Phenol

Room disinfection Formaldehyde