This document discusses various methods of sterilization and disinfection. It defines key terms like sterilization, disinfection, antisepsis, and asepsis. It describes physical methods like heat, radiation, and filtration. It provides details on moist heat sterilization methods like autoclaving, boiling, and steaming. It also discusses sterilization using gas, disinfectants, and antiseptics. Risk groups for different pathogens are defined.

1. Dr Shireen Rafiq
MBBS, M.Phil, Ph.D

2. PURPOSE
 CULTURE MEDIA
 INSTRUMENTS AND GLASSWARE
 DISPOSAL OF MATERIALS
 STERILIZATION CHECKS

3. Basic terms
 sterilization
 disinfection
 Antisepsis
 Bacteriostasis
 Asepsis

4. Sterilization
 The process of destroying all microbial
forms both pathogenic and non
pathogenic. A sterile object is one free of all
microbial forms, including bacterial spores.

5. Disinfection
 The reduction or elimination of pathogenic
microorganisms in or on materials, so they
are no longer a health hazard.

6. Antisepsis
 Use of chemical agents on skin or other
living tissue to inhibit or eliminate
microbes; no sporicidal action is implied.

7. Bacteriostasis
 Inhibits the growth of micro-organisms.

8. Asepsis
 No living microorganisms exists.

9. Risk Groups (WHO)
Risk Groups Organisms
Group I Low risk Food spoilage bacteria moulds and yeasts
Group II
Moderate risk
Staph, Strep, vibrios, Enterobacteriaceae,
adeno, polio, coxsackie V
Toxoplasma, Leishmania
Group III
High Risk
Brucella, MTB, S typhii, Plague bacteria
HIV, Rickettsiae,
Group IV
Viruses
Smallpox, Hemmorhagic fevers, Encephalitis,
Arbo Viruses

10. PhysicalPhysical ChemicalChemical
Methods of Sterilization
and Disinfection
Methods of Sterilization
and Disinfection
HeatHeat
RadiationRadiation
FiltrationFiltration
DisinfectantsDisinfectants
AntisepticsAntiseptics

11. HEATHEAT
DRYDRY
MOISTMOIST
RED HEATRED HEAT
HOT AIR
OVEN
HOT AIR
OVEN
PASTEURIZATIONPASTEURIZATION
BOILINGBOILING
STEAMINGSTEAMINGWASHING
MACHINES
WASHING
MACHINES

12. High Temperature
static action cidal action

13. DRY HEAT
>Red Heat Flaming (Metallic objects)
>Hot Air Sterilizer 160-1800
C
Glassware and instruments
Oils, Powders, Waxes

14. MOIST HEAT advantage
 Pasteurization 56C for 1 hour
(Heat below 100C)
Heat labile fluids (serum, body fluids)
Juices, milk
Washing machines (washing+heat)
(3mins at 71C or 1min at 80C
Linen, bedpans, protective clothing,

15. MOIST HEAT
Below 100
 Pasteurization
 Vaccine bath
At 100
 Boiling
 Steaming
 Tyndalization
Above 100
 Autoclave

16. MOIST HEAT
 Boiling at 100C for 5mins (water bath)
>Advantage HBV, all vegetative
bacteria, some spores
Useful in emergency
conditions
>Disadvantage Not for heat labile
/porous articles

17. MOIST HEAT cont…
 Steaming at 100C (for 5 mins)
>Advantage: prevents cracking of
glassware
Heat labile culture media
Destroys most vegetative
bacteria
>Disadvantage: Spores, thermophiles not
destroyed

18.  fractional sterilization or Tyndallization:
1) Steam heating to 100 °C for 30 min——
Vegetative cells are destroyed but endospores survive
2) Incubate at 30 °C -37 °C overnight
Most bacterial endospores germinate
3) Second heat treatment, 100 °C, 30 min
Germinated endospores are killed.
4) Second incubation at 30°C-37 °C overnight
Remaining endospores germinate
5) Third heat treatment, 100 °C, 60 min
Last remaining germinated endospores are killed

19. MOIST HEAT cont…
 AUTOCLAVES
Produces saturated steam at higher than
atmospheric pressure
Latent + sensible heat
121C for 15mins at 15lbs pressure
Denaturation of all microbial protein

21.  Penetration time
Thermocouples
 Hold time
Bacillus stearothermopilus
 Condensation of steam
Saturated steam
 1670 vol steam
=1vol condensate

22.  Uses
Culture media and other lab supplies
Fluids in sealed bottles
Glassware and reusable equipment
Waste materials
Grade 2-4 pathogens

23. Autoclave Efficiency testing
MC
 Bowie Dick Test
 Browns Indicator Tube or TST indicator

28. STERILIZATION by GAS
 Uses
Heat labile equipment
LTSF
Ethylene Oxide
Gas plasma

29. Filters
 Membrane filters
Diameters 13-239mm
Pore sizes 0.015-12um
Syringe filters
Vacuum filters
Pressure filters
Air filters HEPA

30. 

31. Air filters

32. Hepa

33. Vacuum filter

34. Sterilization by Radiation
 Ionizing Radiation
 UV Radiation

35. Disinfection
 Definition
Destruction of microorganisms but not
usually their spores
British Standards Institution
Disinfectants: Chemical agents capable of
disinfection

36. GROUPS OF DISINFECTANTS
 PHENOLICS Disruption of plasma membrane.
 Denaturation
 HALOGENS Inhibition of protein function. Strong
 oxidizing agents
 BIGUANIDES Disruption of plasma membrane.
 ALCOHOLS Protein denaturation and lipid dissolution
 QUATS Disruption of plasma membrane.

37. Use of Disinfectants
 Decontamination of surfaces
 Medical devices
 Large surface areas
 Heat labile objects

38. Activity
 Gram positive easily killed
 Gram negative more resistant (Pseudos)
 AFB even more resistant
 Viruses variable
 Spores very resistant
Disadvantages

39. Disinfection of Surfaces and
Spillages
 Body Fluids, Blood, Virus –
10,000ppm Hypochlorite
2% Phenol
2% Glutaraldehyde
Phenolics : Gram positive and negative
bacteria and Mycobacteria,
Not suitable for endospores,
or viruses

41.  Glutaraldehyde (Cidex)
Pre-cleaned surfaces
Small surfaces areas with suspected viral
contamination
Useful against Mycobacteria and spores

42.  Ethyl Alcohol
Bactericidal
Not active against spores and many
viruses
Poor activity in presence of organic material.
WIPING WITH ALCOHOL IS DANGEROUSLY INEFFECTIVE

43. Sterilization by Gas
 Ethylene Oxide Alkylation of protein, DNA and RNA
 Formaldehyde
Uses: Devices and materials that cannot stand high temp/humiditiy
Critical and semi critical items