P1 | Sterilization and Disinfection (LECTURE)

Question 1

Refers to the destruction of all forms of life, including bacterial spores

• Originated more than 100 years ago
• Complete removal of microorganisms including spores
• “All or nothing” process

Answer 1

Sterilization

Question 2

What methods are used in Sterilization?

Answer 2

• Physical method
• Chemical method

Question 3

Refers to a process that eliminates a defined scope of microorganisms, including some spores

• Reduces the number of microorganisms
• Physical or chemical methods may be used, but most disinfectants are chemical agents applied to inanimate objects

Answer 3

Disinfection

Question 4

Substance applied to the skin for the purpose of eliminating or reducing the number of bacteria present

Do not kill spores and cannot be used as disinfectants

• destroy/inhibit the growth of microorganisms
• Joseph Lister used:

Answer 4

Antiseptic

E.g., Alcohol

• Carbolic acid

Question 5

Factors that Affect the Degree of Killing of Microorganisms

(T.O, N.O, C.D.A, P.O.M, N.S.D, C.T, T, pH, BF, CoD)

Answer 5

1. Types of Organisms
2. Number of Organisms
3. Concentration of Disinfecting Agent
4. Presence of Organic Material
5. Nature of Surface to be Disinfected
6. Contact Time
7. Temperature
8. pH
9. Biofilms
10. Compatibility of Disinfectants

Question 6

Different organisms have varying ability in withstanding and chemical and physical treatment due to the different biochemical composition of these organisms and various mechanisms that they use to protect themselves

Answer 6

Types of Organisms

Question 7

Types of Organisms

(SFB, MB spp., BFB, P)

Answer 7

1. Spore forming Bacteria
2. Mycobacterium spp.
3. Biofilm forming Bacteria
4. Prions

Question 8

Types of Organisms

Spores are coated with proteins, lipids and carbohydrates as well as dipicolinic acid calcium

SFB

• able to resist heat

Answer 8

Spore forming Bacteria

Question 9

Types of Organisms

cell wall is high in lipid which enables them to become resistant to most environmental stress such as desiccation

MB spp.

Answer 9

Mycobacterium spp.

Question 10

Types of Organisms

certain bacteria can aggregate into communities of bacteria which makes then resistant to chemical and physical means of destruction

(BFB)

Answer 10

Biofilm forming Bacteria

• Biofilm: microorganism living in community

Question 11

Types of Organisms

naked pieces of proteins, similar to viruses but without the nucleic acid, that is most resistant to the action of heat, radiation, and chemicals

• agent able to cause the generative diseases of our nervous system

Answer 11

Prions

Question 12

this factor basically refers to the amounts of organisms present in the object to be treated referred to as microbial load (bioburden)

• the higher the number of organisms, the longer the exposure time needed to eliminate 99.9% of the microorganisms

Answer 12

Number of Organisms

Question 13

The amount needed to destroy microorganisms varies based on the agent to be used.

concentrated agents do not necessarily mean that it would work better

• It is therefore important to follow, the correct preparation and dilution as prescribed by the manufacturer.
• 1:10 bleach (common disinfectant)

Answer 13

Concentration of Disinfecting Agent

Question 14

Blood, puss, and mucus are examples of organic materials that may prevent the full contact of the agent to the organisms, hence limiting its action

POM

• Example: is bleach (sodium hypochlorite) that is easily inactivated by organic material.

Answer 14

Presence of Organic Material

Question 15

Some instrument that we use in the laboratory sometimes are made up of biomaterial which exempts them to disinfection or sterilization due to possible damage

NSD

• Example: is endoscopic instruments which can’t be autoclaved

Answer 15

Nature of Surface to be Disinfected

Question 16

It is critical to observe proper (___) of the agent and the object to be disinfected or sterilized.

• In principle, (____) may be affected by all previous factors already mentioned as well as temperature

Answer 16

Contact Time

Question 17

Alcohol and betadine has to be in contact for about (____) to work properly

Answer 17

1-2 minutes

• Spore forms may need more contact time than its vegetative counterpart

Question 18

their activity may increase at a certain degree by a corresponding increase in temperature or may decrease when temperature is decreased

Answer 18

Temperature

Question 19

Temperature

disinfectants are usually used at room temperature (____)

Answer 19

20C to 220C

Question 20

Temperature

Too high or low temperature may?

Answer 20

inactivate disinfectants and sterilants

Question 21

It is also important to consider the (____) of the material to be treated and he agent itself.

• Manufacturers usually optimize this factor to achieve maximum activity

Answer 21

pH

Question 22

Certain bacteria have to ability to form communities of layers of bacteria with protective shield which is called as?

• community of bacteria
• Inanimate and animate objects

Answer 22

Biofilms

Question 23

It is important to consider that (____) formation may require longer contact time or increase in the concentration of the agent

• E.g., Hospital – Catheter (lots of bacteria)

Answer 23

Biofilm

Question 24

Some disinfectants may inactivate the action of another hence it is also important to consider the (____)

• bleach and quaternary ammonium compounds which may negate each other

Answer 24

Compatibility of Disinfectants

Question 25

**METHODS OF STERILIZATION**

Answer 25

**Physical methods:** * Moist Heat * Dry Heat * Ionizing Radiation * Filtration **Chemical Methods:** * Peracetic Acid * Formaldehyde Vapor/ Vapor Phase H2O2 * Glutaraldehyde * Ethylene Oxide (ETO) Gas

Question 26

# ***Physical Methods*** coagulation of bacterial proteins including bacterial enzymes

Answer 26

Moist Heat

Question 27

Simplest means of sterilizing a material:

Answer 27

Heat

Question 28

Types of moist heat: | A.T.I

Answer 28

* Autoclave * Tyndallization * Inspissation

Question 29

# **Physical Methods:** *moist heat* operates based on the principle of steam under pressure

Answer 29

Autoclave

Question 30

# **Physical Methods:** *moist heat* **Effective indication:** * **Sterilization:** * **Decontamination:** * **Biological indicator:** | Autoclave

Answer 30

* 121C to 15lbs/ in2 for 15 minutes * 135C for 30lbs in2 for 30 minutes * *Bacillus stearothermophilus*

Question 31

# **Physical Methods:** *moist heat* fractional discontinuous sterilization

Answer 31

Tyndallization

Question 32

# **Physical Methods:** *moist heat* * **Effective indication:** * **Instrument:** | Tyndallization

Answer 32

* 100C for 30-60 minutes * Arnold’s Sterilizer

Question 33

# **Physical Methods:** *moist heat* thickening through evaporation

Answer 33

Inspissation

Question 34

# **Physical Methods:** *moist heat* * **Effective indication:** * **Instrument:** | Inspissation

Answer 34

* 75C to 80C for 2 hours * Inspissator

Question 35

**Control of Microorganisms using Heat Methods** | BW(S), AC (SUP), PBM, PFM, O (DH)

Answer 35

**Methods:** * Boiling Water (steam) * Autoclave (steam under pressure) * Pasteurization Batch Method * Pasteurization Flash Method * Over (Dry Heat)

Question 36

# **Control of Microorganisms using Heat Methods** * Boiling Water (steam) * Autoclave (steam under pressure) * Pasteurization Batch Method * Pasteurization Flash Method * Over (Dry Heat) | *Temperature*

Answer 36

* 100 C * 121.6 C * 63 C * 72 C * 160-180 C | *Temperature*

Question 37

# **Control of Microorganisms using Heat Methods** * Boiling Water (steam) * Autoclave (steam under pressure) * Pasteurization Batch Method * Pasteurization Flash Method * Over (Dry Heat) | *Time required*

Answer 37

* 15min * 15min at 15psi * 30min * 15s * 1.5-3hrs | *Time required*

Question 38

# **Control of Microorganisms using Heat Methods** * Boiling Water (steam) * Autoclave (steam under pressure) * Pasteurization Batch Method * Pasteurization Flash Method * Over (Dry Heat) | *Applications*

Answer 38

* Kills microbial vegetative forms; endospores survive * Sterilizes and kills endospores * Disinfects and kills milk-borne pathogen and vegetable forms; endospores survive * Same, but shorter time at higher temp. * Sterilizes; keeps materials dry | *Applications*

Question 39

# **METHODS OF STERILIZATION:** Physical Methods oxidation of bacterial components | DH ## Footnote * requires longer exposure time

Answer 39

Dry Heat

Question 40

# **Physical Methods:** Dry heat direct application of flame in aseptic technique

Answer 40

Direct Flame

Question 41

# **Physical Methods:** Dry heat used in the sterilization of heat resistant materials

Answer 41

Dry/Hot Air Oven

Question 42

# **Physical Methods:** Dry heat * **Effective indication:** * **Biological indicator:** | *Dry/Hot Air Oven*

Answer 42

* 160-180C for 1.5 to 2 hours * *Bacillus subtilis var. niger*

Question 43

# **Physical Methods:** Dry heat burns materials into ashes; used in the disposable of biological wastes

Answer 43

Incineration

Question 44

# **Physical Methods:** Dry heat * **Effective indication:** | Incineration

Answer 44

* 870-980C for 2 seconds

Question 45

# **Physical Methods** works by alkylation of nucleic acid of bacteria using high energy short wavelength deep penetrating gamma rays; used for heat sensitive materials ## Footnote * sterilization of recycled materials

Answer 45

Ionizing Radiation

Question 46

# **Physical Methods:** Ionizing Radiation * **Biological indicator:**

Answer 46

* *Bacillus pumilis*

Question 47

# **Physical Methods** * Based on membrane gradient by differences in particle size * Used for the sterilization of heat sensitive materials

Answer 47

Filtration

Question 48

# **Physical Methods:** Filtration **Types of Filtration:**

Answer 48

* Water/ Liquid Solutions/ Antibiotics/ Vaccines * Air: High Efficiency Particulate Air Filter (HEPA)

Question 49

# **Physical Methods:** Filtration usually uses a thin membrane filter of cellulose acetate with different pore size depending on the intended purpose

Answer 49

Water/ Liquid Solutions/ Antibiotics/ Vaccines

Question 50

# **Physical Methods:** Filtration most bacteria, yeasts, and molds are retained but may allow passage of Pseudomonas- like organisms | *Water/ Liquid Solutions/ Antibiotics/ Vaccines* ## Footnote (size)

Answer 50

0.45 – 0.80um

Question 51

# **Physical Methods:** Filtration used to filter Pseudomonas- like organisms; used for critical sterilization of parenteral solutions | *Water/ Liquid Solutions/ Antibiotics/ Vaccines* ## Footnote (size)

Answer 51

0.22um

Question 52

# **Physical Methods:** Filtration able to retain small viruses | *Water/ Liquid Solutions/ Antibiotics/ Vaccines* ## Footnote (size)

Answer 52

0.01um

Question 53

# **Physical Methods:** Filtration Has a pore size of 0.3 μm; usually used in Biological Safety Cabinet (BSC) and rooms of immunocompromised patients

Answer 53

Air: High Efficiency Particulate Air Filter (HEPA)

Question 54

# **Chemical Methods** for surgical instruments

Answer 54

Peracetic Acid

Question 55

# **Chemical Methods** for HEPA filters and large spaces

Answer 55

Formaldehyde Vapor/ Vapor Phase H2O2

Question 56

# **Chemical Methods** for medical instruments (e.g., bronchoscopes, etc.)

Answer 56

Glutaraldehyde

Question 57

# **Chemical Methods** This method is also used extensively by the manufacturing industry for the sterilization of low-cost thermoplastic products ## Footnote * For large spaces and spaceships and other heat sensitive materials

Answer 57

Ethylene Oxide (ETO) Gas

Question 58

# **Chemical Methods:** Ethylene Oxide (ETO) Gas * The recommended concentration of ethylene oxide is (____) * per liter of chamber space at (____)

Answer 58

* 450 to 700mg * 55C to 60C for 2 hours

Question 59

# **Chemical Methods:** Ethylene Oxide (ETO) Gas * **Biological indicator:**

Answer 59

* *Bacillus subtilis var. globijii*

Question 60

**METHODS OF DISINFECTION**

Answer 60

**Physical Methods** * Boiling * Pasteurization * Non-Ionizing Radiation **Chemical Methods** * Alcohol * Halogens * Heavy Metals * Quaternary Ammonium Compounds (QUATS) * Phenol/Phenolic Compounds/Bisphenols

Question 61

# **Physical Methods** Destroys vegetative cells of bacteria but not their spores

Answer 61

Boiling

Question 62

# **Physical Methods:** Boiling * **Effective indication:**

Answer 62

* 100C for 15-30 minutes

Question 63

# **Physical Methods** Used for the preservation of alcoholic beverages such as beers, wines, and also dairy products such as milks and yogurt

Answer 63

Pasteurization

Question 64

# **Physical Methods:** Pasteurization * **Batch:** * **Flash:** * **Ultra-High Temperature (UHT):**

Answer 64

* 62.5C for 30 minutes * 72C for 15 seconds * 72C to 110C for 5 seconds

Question 65

# **Physical Methods** Uses low energy long wavelength ultraviolet rays to disinfect heat sensitive materials as well as large spaces

Answer 65

Non-Ionizing Radiation

Question 66

# **Chemical Methods:** Alcohol * **MOA:** * **Minimum Contact Time:** ## Footnote * 70% Alcohol not 90% * Ethyl and Isopropyl alcohol kills (____)

Answer 66

* Dehydration, Lipid dissolution and Protein denaturation * 1-2 minutes or until completely evaporated ## Footnote * *Mycobacterium tuberculosis*

Question 67

# **Chemical Methods:** Halogens * **MOA:** * **Chloride (Cl) IN NaOCl:** ## Footnote * commonly used in household

Answer 67

* inhibits protein function and acts as strong oxidizing agents * used as disinfecting agents in many laboratory and hospitals spaces, surfaces, and also in treating water for portability Iodine (I2) in Betadine used as a household antiseptics and surgical antiseptics ## Footnote * Sodium hydrochloride

Question 68

# **Chemical Methods:** Heavy Metals * **MOA:** * **Mercury (Hg):** * **Cooper (Cu):** * **Silver (Ag):**

Answer 68

* Denaturation of enzymes and other essential bacterial proteins * active ingredient or merthiolate but this is already banned in the market due to its known toxicity * CuSO4 crystals are used as algaecide in swimming pools and aquarium * 1% AgNO3 – used as prophylactic agent in Crede’s Prophylaxis in suspected cases of Ophthalmia neonatorum (caused by *Neisseria gonorrhoeae*) (replaced with erythromycin)

Question 69

# **Chemical Methods:** Quaternary Ammonium Compounds (QUATS) * **MOA:** * **Zephiran:** * **Cepacol:** ## Footnote * enzyme inhibition, protein denaturation, and disruption of plasma membrane

Answer 69

* enzyme inhibition, protein denaturation, and disruption of plasma membrane * Benzalkonium chloride * Cetylpyridium chloride

Question 70

# **Chemical Methods:** Phenol/Phenolic Compounds/Bisphenols * **MOA:** ## Footnote * *Germicidal soaps*

Answer 70

* Plasma membrane destruction and enzyme denaturation

Question 71

# Chemical Agents Commonly used as Disinfectants and Antiseptics (SUMMARY) TYPES:

Answer 71

* Alcohols (50%-70%) * Aldehydes (in solution) * Halogens * Heavy Metals * Detergents * Phenolics * Gases

Question 72

# Chemical Agents Commonly used as Disinfectants and Antiseptics (SUMMARY) * Alcohols (50%-70%) * Aldehydes (in solution) * Halogens * Heavy Metals * Detergents * Phenolics * Gases | *Agent/s*

Answer 72

* Ethanol, isopropanol, benzyl alcohol * Formaldehyde (8%), glutaraldehyde (2%) * Tincture of iodine (2% in 70% alcohol) Chlorine and chlorine compounds * Silver nitrate (AgNO3) & Mercuric chloride (HgCl2) * Quaternary ammonium compounds * Phenol, carbolic acid, Lysol, hexachlorophene * Ethylene oxide

Question 73

# Chemical Agents Commonly used as Disinfectants and Antiseptics (SUMMARY) * Alcohols (50%-70%) * Aldehydes (in solution) * Halogens * Heavy Metals * Detergents * Phenolics * Gases | *Action*

Answer 73

* Denature proteins; make lipids soluble * React with NH2+, -SH and -COOH groups * (Inactive proteins) Reacts with water to form hypochlorous acid (HClO); oxidizing agent * Precipitates proteins Reacts with -SH groups; lyses cell membrane * Disrupt cell membranes * Denature proteins; disrupt cell membranes * Alkylating agent

Question 74

# Chemical Agents Commonly used as Disinfectants and Antiseptics (SUMMARY) * Alcohols (50%-70%) * Aldehydes (in solution) * Halogens * Heavy Metals * Detergents * Phenolics * Gases | *Applications and Precautions*

Answer 74

* Skin antiseptics * Disinfectants; kill endospores; toxic to humans * (Skin disinfectants) Used to disinfect drinking water; surface disinfectants * Eye drop (1% solution) Disinfectant; toxic at high concentrations * Skin antiseptics; disinfectants * Disinfectants at high concentrations; used in soaps at low concentration * Sterilization of heat-sensitive objects