Transcript Unit 1: History and Scope of Microbiology

Chapter 7
Control of Microbial
Growth
Terminology
• Sepsis refers to microbial contamination.
• Asepsis is the absence of significant
contamination.
• Antisepsis: Removal of pathogens from living
tissue
• Degerming: Removal of microbes from a
limited area
• Sanitization: Lower microbial counts on eating
utensils
Terminology cont’d
• Decontaminated
- item that has been treated to reduce # of
disease causing organisms
• Preservation
- delaying spoilage of foods
• Sterilization, Disinfection, Antiseptic,
Bacteriocidal, Bacteriostatic
Terms used:
Sterilization vs. Disinfection
• Sterilization: destroying all forms of life
• Disinfection: destroying pathogens or
unwanted organisms
Disinfectant vs. Antiseptic
 Disinfectant: antimicrobial agent used on inanimate
objects
 Antiseptic : antimicrobial agent used on living tissue
cidal vs. static
• Biocide/Germicide: Kills microbes
• Bacteriostasis: Inhibiting, not killing, microbes
• Examples:
- Bactericidal - kills bacteria
- Bacteriostasis (Bacteriostatic)- inhibits
bacterial
growth
- Fungicidal
- Fungistatic
- Algacidal
- Algastatic
Methods of control
• Physical or chemical?
– physical control includes heat, irradiation,
filtration and mechanical removal
– Chemical control involves the use of microbial
chemicals
– Depends on the situation
– degree of control required
Methods of control cont’d
• Daily life
- Cooking
- refrigeration
- cleaning
- soap water
mechanical
Mechanical and
chemical
Methods of control cont’d
• Hospitals
– Important to minimize nosocomial infection
(hospital acquired infection) due to
- weakened patients’ condition
- breaching of intact skin
- high concentration of pathogens from patients
and workers
– Sterile condition
Methods of control cont’d
• Microbiology lab utilizes
– Sterile equipment
– Aseptic technique
– And possesses workers who takes care of the
nature of of microbiologists (GLP)
Methods of control cont’d
• Foods/food production industry
- physical removal
- adding chemicals
- may result in toxicity
- clean surface/ machinery
Selection of Control Method
• Antimicrobial procedure used for control of
microbial growth is based on
– Types of microbe
– Extent of contamination
– Environmental conditions
– Potential risk
Selection of Control Method cont’d
• Types of microorganism
- some organisms are more resistant and
require stronger measures for control
- endospores require chemical treatment for
10 hours
- Mycobacterium’s waxy cells are resistant
to chemicals
Mycobacterial cell wall: 1-outer
lipids, 2-mycolic acid, 3polysaccharides (arabinogalactan),
4-peptidoglycan, 5-plasma
membrane, 6-lipoarabinomannan
(LAM), 7-phosphatidylinositol
mannoside, 8-cell wall skeleton
Selection of Control Method cont’d
• Extent of microbial population
- larger population take more time to destroy
- usually 90% of the population is destroyed in
a given period
e.g if in 1st 3 minutes 90% of the
population is destroyed, then 90% of the
remaining population gets destroyed in the
next 3 minutes and so on
Selection of Control Method cont’d
• Environmental conditions
- pH, temperature
- presence of
- organics: blood
Must be cleaned first, then
- dirt
controlled
- grease
- the potential risk of transmitting infectious agents
- critical items
- semicritical items
- non-critical items
Selection of Control cont’d
• Critical items have
- indirect contact with body tissues
- needles, scalpels
• Semicritical items have contact with
- mucous membranes but it does not penetrate
endoscopes, endotrachial tubes
Selection of Control cont’d
• Non-critical items have
- indirect contact with unbroken skin
- countertops, stethoscopes
Methods to Control Microbial Growth
Dry Heat
Incineration
Heat Sterilization
Boiling
Moist
Heat
Physical
Methods
Filtration
Air
Liquid
UV Light
Radiation
Ionizing Radiation
Autoclaving
Pasteurization
Physical Microbial Controls: Heat
• Heat as a microbial control
- fast, reliable, inexpensive
- does not introduce potential toxic substances
• Types of heat control include
- moist heat
- pasteurization
- pressurized steam
- dry heat
Physical Microbial Controls: Heat
cont’d
• Moist heat
- causes irreversible coagulation of proteins
found in microorganisms
- 10 minutes of boiling
- most microbes and viruses will be
destroyed except endospores and few
others which can survives hours of boiling
Physical Microbial Controls: Heat
cont’d
• Pasteurization
- reduces number of heat sensitive organisms
- widely used in milk and juices
increases shelf life and does not alter
quality
- original pasteurization was 62ºC, 30 mins
- now: UHT-shorter time 72ºC, 15 secs
Physical Microbial Controls: Heat
cont’d
• Pressurized steam
- pressure cooker or autoclave
- higher air pressure increases the temperature
at which steam forms
- 15 psi (lbs/square inch) at 121ºC for 15 mins
-effective to kill endospores
The autoclave: Moist heat and pressure
• 15psi, 121ºC, 15 minutes
• Thermal death point (TDP): Lowest
temperature at which all cells in a culture get
killed in 10 mins
• Thermal death time (TDT): time to kill all
cells in a culture
• Decimal reduction time (DRT): Minutes to
kill 90% of a population at a given temperature
The autoclaving machine
Temperature of steam and Pressure at
sea level
The autoclaving machine
Physical Microbial Controls: Heat
cont’d
• Dry heat
- without moisture e.g. flaming
- burns cell constituents
- object is oxidized to ash
- irreversibly denatures proteins
- takes longer (200ºC, 1.5 hrs dry=121ºC, 15 min moist)
- advantages are for powders, does not corrode metals
and blunt sharps
- e.g flasks, tubes, pipettes in microbiological
laboratories.
Physical Microbial Controls: Filtration
• Used for heat sensitive fluids
• air
Physical Microbial Controls: Filtration
cont’d
• Fluids
– solutions of antibiotics, vitamins,
tissue extracts, animal serum, etc.
– Depth filters
–able to retain microorganisms
while allowing fluids to pass
through
– Membrane Filter
- The use of graded pore size 0.20.4µm
Physical Microbial Controls: Filtration
cont’d
• Air
- HEPA (High Efficient Particulate Air) filter and
laminar air flow are commonly used
- filter incoming air and outgoing air respectively
- HEPA filter prevents the income of 0.3µm and large
size particles to enter.
Physical Microbial Controls: Radiation
• “Cold sterilization“ for disposable materials
made up of plastics, wool, cotton, etc without
altering the material.
• Radiation damages DNA
• Ionizing radiation (X-rays, gamma rays, electron
beams)
• Non-ionizing radiation
- UV
- Microwaves kill by heat not especially
antimicrobial
Physical Microbial Controls: Radiation
cont’d
• Gamma irradiation
- penetrate deeply
- for heat sensitive materials
- causes biological damage to microorganisms
- does not alter food flavor (meat)
Physical Microbial Controls: Radiation
cont’d
• UV light
– damages the structure and function of nucleic acids
– Penetrate poorly- cannot penetrate even into liquid.
– Used to disinfect surfaces
– Can cause damage to human cells
– Germicidal lamps -kill or reduce the number of
viable microorganisms to sterilize microbiological
laboratories hospital operating rooms, and specific
filling rooms in various industries
Physical Microbial Controls: Radiation
cont’d
• Microwave
– Kills by heat
– Does not affect microorganisms directly
Physical Methods used to control Microbial growth
Chemical Microbial Control
Chemical
Method
Gas sterilants
Ethylene oxide
Antiseptics and
disinfectants
Germicidal
chemical
Chemical Microbial Control cont’d
• Grouped according to potency
– Sterilants
– High-level
– Intermediate level
– Low level
Chemical Control
sterilants
• destroy microorganisms, endospores and
viruses
• used for critical equipment-scalpels
chemical control cont'd
• high level
- destroy viruses and vegetative microorganisms (no
endospores)
- used for semicritical equipment: endoscopes
• intermediate level
- destroy vegetative microorganisms, some viruses
- used for non-critical equipment: sthetoscopes
• low level
-destroy fungi, vegetative microorganisms
- used for general purpose disinfectants
Selecting Germicidal Chemical
• Germicide: An agent capable of killing pathogens and nonpathogens but not necessarily endospores
• toxicity to human or environment?
- weigh the benefits vs the risks
• presence of organic material
- hypochlotrite is inactivated by the presence of organic matter
• compatibility
-electrical equipment with a liquid??
• residue
- some have to be rinsed with sterile water
• cost and availability
Selecting germicidal Chemical cont'd
• storage and stability
- may come in concentrated form for ease in
storing
- those have to be mixed
• environmental risk
- is neutralization necessary before disposal?
Classes of Germicidal Chemicals
•
•
•
•
•
alcohols alcohols
aldehydes
biguanides
ethylene oxide
halogens: oxidize proteins
Classes of Germicidal Chemicals
• alcohols
- coagulated enzymes and proteins
- damage lipid membranes
- on-toxic
- inexpensive
- no residue
Classes of Germicidal Chemicals
cont’d
• aldehydes
- inactivate proteins and
nucleic acids
- toxic to humans
• Peroxygens
- oxidizing agents
- hydrogen peroxide
- leaves no residue
Classes of Germicidal Chemicals cont’d
• biguanides
- extensive antiseptic use
- adheres and persists on skin, mucous membranes
- low toxicity
• Phenolic compounds
– Hitorically important
– Irritant, unpleasant odor
– Destroy cytoplasmic membrane and denatures protein
Classes of Germicidal Chemicals
cont’d
• ethylene oxide
- reacts with proteins
- gas: penetrable
- mutagenic
• Metal compounds
– Interfere protein function
– Toxic
– pollutants
Classes of Germicidal Chemicals
cont’d
• halogens: oxidize proteins
-chlorine
-irritating to skin
-organic compounds consume free chlorine
-iodine
-tincture
- Iodophore
Effect of germicidal activity on
Microbes
Cytoplasmic
membrane
• Biguanides
• Phenolics
• Quats
DNA
• Ethylene
oxides
• Aldehydes
Proteins
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•
•
•
•
Alcohols
Aldehydes
Halogens
Metals
Ozone
Peroxygens
Phenolics
Effect of
germicidal activity
on Microbes
Chemical methods of microbial
control
• Evaluating a disinfectant
- Disk diffusion method
Preservation of Perishables
• Extends shelf life
– Slow or halts microbial growth thus delaying
spoilage
Preservation of Perishables
• Chemical preservatives
– Some chemical preservatives are used in non-food
items
– Food preservatives must be non-toxic to humans
• Benzoic acid, propionic acids, nitrate are commonly
used
• Nitrate
– Inhibits germination of C. botulinum endospores
Preservation of Perishables
• Low temperature storage
– Temperature dependent
• most microorganisms do not reproduce in ordinary
refrigerator (0-7ºC)
– Freezing
• ice crystals can cause irreversible damage to many
microorganisms (kills up to 50% growth)
• Freezing stops all growth, but may start to reproduce
again once food is thawed
Preservation of Perishables
• Reducing water
– Salt/sugar
– Draw water out of cell
– Less available for microorganisms
• Drying
– Desiccation
• Removing water such as milk powder
Salt cured meat
Preservation of Perishables
• Lyophilization
– Freeze drying
• Freeze food first
• followed by putting in vacuum
Factors that influence effectiveness
•
•
•
•
Number of microbes
Environmental influences
Time of exposure
Microbial characteristics
Microbial death curve
Limitations: Microbial Characteristics
Actions of Microbial Control Agents
• Alteration of membrane
permeability
• Damage to proteins
• Damage to nucleic acids
Questions????
Sand rich in salts n nitrates preserved mummy