Controlling Microbial Growth
What factors limit microbial growth? In what situations are large microbial numbers undesirable?
•
Concept of Microbial Control
•
Factors Which Affect Control
•
•
•
Temp., species type and status, environment
Physical Control Methods
•
Heat: Moist vs. Dry
•
Autoclaving, pasteurization
•
Filtration
•
Cold
•
Desiccation & high osmotic pressure
•
Radiation (UV, gamma rays)
Chemical Control Methods
•
Factors which influence effectiveness
•
•
Dilution, time, pH, organic matter
Types of Disinfectants/Antiseptics
•
Phenol and phenolics (e.g.. amphyl)
•
Halogens (Chlorine, iodine, bromine)
•
Alcohols (e.g.. isopropyl alcohol)
•
Heavy metals (Ag, Hg, Cu, Zn)
•
Surface active agents (soaps & detergents)
•
Quaternary Ammonium Cmpds (quats)
•
Aldehydes (e.g.. glutaraldehyde)
Microbial growth can be inhibited by temperature, moisture control or by antimicrobial chemicals
Terminology
• Sterilization: Removal of all microbial life
• Commercial Sterilization: Killing C. botulinum
endospores
• Disinfection: Removal of pathogens
• Antisepsis: Removal of pathogens from living tissue
• Degerming: Removal of microbes from a limited area
• Sanitization: Lower microbial counts on eating utensils
• Biocide/Germicide: Kills microbes
• Bacteriostasis: Inhibiting, not killing, microbes
• Bacterial populations die at a constant logarithmic rate.
Figure 7.1a
Effectiveness of antimicrobial treatment depends on:
• Number of microbes
• Environment
(organic matter,
temperature,
concentration,
biofilms)
• Time of exposure
• Microbial
characteristics (e.g.
glycocalyx,cell wall,
resistance)
Figure 7.1b
Controlling Microbial Growth
What factors limit microbial growth? In what situations are large microbial numbers undesirable?
•
Concept of Microbial Control
•
Factors Which Affect Control
•
•
•
Temp., species type and status, environment
Physical Control Methods
•
Heat: Moist vs. Dry
•
Autoclaving, pasteurization
•
Filtration
•
Cold
•
Desiccation & high osmotic pressure
•
Radiation (UV, gamma rays)
Chemical Control Methods
•
Factors which influence effectiveness
•
•
Dilution, time, pH, organic matter
Types of Disinfectants/Antiseptics
•
Phenol and phenolics (e.g.. amphyl)
•
Halogens (Chlorine, iodine, bromine)
•
Alcohols (e.g.. isopropyl alcohol)
•
Heavy metals (Ag, Hg, Cu, Zn)
•
Surface active agents (soaps & detergents)
•
Quaternary Ammonium Cmpds (quats)
•
Aldehydes (e.g.. glutaraldehyde)
Microbial growth can be inhibited by temperature, moisture control or by antimicrobial chemicals
Physical Methods of Microbial Control
• Heat
• Thermal death point (TDP): Lowest temperature at
which all cells in a culture are killed in 10 min.
• 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
Autoclaving
• Moist heat
denatures
proteins
• Autoclave:
Steam
under
pressure
• 15 min at
121oC at
15 psi
Figure 7.2
Pasteurization
63oC for 30 minutes
72oC for 15 seconds
140oC for 1 second
Pasteurization reduces spoilage organisms and pathogens
Physical Methods of Microbial Control
• Dry Heat Sterilization kills by oxidation
• Flaming
• Incineration
• Hot-air sterilization
Equivalent treatments
Hot-air
Autoclave
170˚C, 2 hr
121˚C, 15 min
Controlling Microbial Growth
What factors limit microbial growth? In what situations are large microbial numbers undesirable?
•
Concept of Microbial Control
•
Factors Which Affect Control
•
•
•
Temp., species type and status, environment
Physical Control Methods
•
Heat: Moist vs. Dry
•
Autoclaving, pasteurization
•
Filtration
•
Cold
•
Desiccation & high osmotic pressure
•
Radiation (UV, gamma rays)
Chemical Control Methods
•
Factors which influence effectiveness
• Dilution, time, pH, organic matter
•
Types of Disinfectants/Antiseptics
• Phenol and phenolics (e.g.. amphyl)
• Halogens (Chlorine, iodine, bromine)
• Alcohols (e.g.. isopropyl alcohol)
• Heavy metals (Ag, Hg, Cu, Zn)
• Surface active agents (soaps & detergents)
• Quaternary Ammonium Cmpds (quats)
• Aldehydes (e.g.. glutaraldehyde)
Microbial growth can be inhibited by temperature, moisture control or by antimicrobial chemicals
Physical Methods of Microbial Control
• Filtration removes microbes by trapping them in filter
• Low temperature inhibits microbial growth
• Refrigeration
• Deep freezing (-20oC or -80oC)
• Lyophilization
• High pressure denatures proteins
• Desiccation prevents metabolism
• Osmotic pressure causes plasmolysis (shrinkage of
cytoplasm)
Physical Methods of Microbial Control
• Radiation damages DNA
• Ionizing radiation (X rays, gamma rays, electron
beams)
• Nonionizing radiation (UV)- surface sterilization only
• (Microwaves kill by heat; not especially antimicrobial)
Controlling Microbial Growth
What factors limit microbial growth? In what situations are large microbial numbers undesirable?
•
Concept of Microbial Control
•
Factors Which Affect Control
•
•
•
Temp., species type and status, environment
Physical Control Methods
•
Heat: Moist vs. Dry
•
Autoclaving, pasteurization
•
Filtration
•
Cold
•
Desiccation & high osmotic pressure
•
Radiation (UV, gamma rays)
Chemical Control Methods
•
Factors which influence effectiveness
• Dilution, time, pH, organic matter
•
Types of Disinfectants/Antiseptics
• Phenol and phenolics (e.g.. amphyl)
• Halogens (Chlorine, iodine, bromine)
• Alcohols (e.g.. isopropyl alcohol)
• Heavy metals (Ag, Hg, Cu, Zn)
• Surface active agents (soaps & detergents)
• Quaternary Ammonium Cmpds (quats)
• Aldehydes (e.g.. glutaraldehyde)
Microbial growth can be inhibited by temperature, moisture control or by antimicrobial chemicals
Chemical Methods of Microbial Control
• Principles of effective disinfection
• Concentration of disinfectant
• Organic matter
• pH
• Time
Chemical Methods of Microbial Control
• Evaluating a disinfectant
• Use-dilution test
1.
Metal rings dipped in test bacteria are dried
2.
Dried cultures placed in diluted disinfectant
(according to manufacturer’s instructions) for
min at 20°C
3.
Rings transferred to culture media to
determine whether bacteria survived
treatment
Chemical Methods of Microbial Control
• Evaluating a disinfectant
• Disk-diffusion method
• Particular species are evaluated in each test
• Zone of inhibition must be at or beyond a certain
diameter
Figure 7.6
Controlling Microbial Growth
What factors limit microbial growth? In what situations are large microbial numbers undesirable?
•
Concept of Microbial Control
•
Factors Which Affect Control
•
•
•
Temp., species type and status, environment
Physical Control Methods
•
Heat: Moist vs. Dry
•
Autoclaving, pasteurization
•
Filtration
•
Cold
•
Desiccation & high osmotic pressure
•
Radiation (UV, gamma rays)
Chemical Control Methods
•
Factors which influence effectiveness
• Dilution, time, pH, organic matter
•
Types of Disinfectants/Antiseptics
• Phenol and phenolics (e.g.. amphyl)
• Halogens (Chlorine, iodine, bromine)
• Alcohols (e.g.. isopropyl alcohol)
• Heavy metals (Ag, Hg, Cu, Zn)
• Surface active agents (soaps & detergents)
• Quaternary Ammonium Cmpds (quats)
• Aldehydes (e.g.. glutaraldehyde)
Microbial growth can be inhibited by temperature, moisture control or by antimicrobial chemicals
Phenolics: Disruption of the plasma membrane
Phenol
O-phenylphenol
(Lysol, Amphyl)
Hexachlorophene
Thymol
PineSol/pine oil
(turpineol)
Halogens
Bromine and chlorine produces are strong oxidizers
Iodine denatures proteins by breaking disulfide bonds
Tincture of
Iodine (I2 );
iodine denatures
proteins by
breaking
disulfide bonds
Sodium
hypochlorite
(NaOCl) in
bleach is a
strong oxidizing
agent.
Povidone iodines (I
linked to organic
molecule) as in Betadine;
denatures proteins by
breaking disulfide bonds
Bromine tablets
(BCDMH bromo-chlorodimethylhydantoin)used in
hot tubs, pools,
kills by oxidation
Chlorine gas
used in swim
pools oxidizing
agent.
Types of Disinfectants: Alcohols
• Alcohols. Ethanol,
isopropanol
• Denatures proteins,
dissolve lipids
Table 7.6
Controlling Microbial Growth
What factors limit microbial growth? In what situations are large microbial numbers undesirable?
•
Concept of Microbial Control
•
Factors Which Affect Control
•
•
•
Temp., species type and status, environment
Physical Control Methods
•
Heat: Moist vs. Dry
•
Autoclaving, pasteurization
•
Filtration
•
Cold
•
Desiccation & high osmotic pressure
•
Radiation (UV, gamma rays)
Chemical Control Methods
•
Factors which influence effectiveness
•
•
Dilution, time, pH, organic matter
Types of Disinfectants/Antiseptics
•
Phenol and phenolics (e.g.. amphyl)
•
Halogens (Chlorine, iodine, bromine)
•
Alcohols (e.g.. isopropyl alcohol)
•
Heavy metals (Ag, Hg, Cu, Zn)
•
Surface active agents (soaps & detergents)
•
Quaternary Ammonium Cmpds (quats)
•
Aldehydes (e.g.. glutaraldehyde)
•
Oxidizing Agents
Microbial growth can be inhibited by temperature, moisture control or by antimicrobial chemicals
Heavy metals
Denature proteins by breaking disulfide bonds
Mercurachrome
antiseptic for wounds
Copper sulfate (CuSO4)
algicide
Zinc chloride (ZnCl2)
mouthwashes
Silver nitrate (AgNO3)
antiseptic for eyes, wounds
Types of Disinfectants: Surface Active Agents
• Surface-Active Agents or Surfactants (Soaps and
Detergents)
Soap
Degerming
Acid-anionic detergents
Sanitizing
Quaternary Ammonium Compounds (Quats)
Disruption of plasma membrane, denaturation of proteins
Benzalkonium chloride
(aka Zephiran)
Roccal: lab disinfectant
Cationic detergents
Cetylpyridinium chloride (Cepacol)
Types of Disinfectants:Aldehydes
• Aldehydes
• Inactivate proteins by cross-linking with functional
groups (–NH2, –OH, –COOH, —SH)
• Glutaraldehyde, formaldehyde
Disinfection of hospital equipment,
especially in respiratory therapy, for
kidney dialysis machines, endoscopes
glutaraldehyde
Acids and Bases: Denaturation of Proteins, Dissolving Membranes
NaOH (sodium hydroxide) in
some floor cleaners
NH4OH in ammonia, esp.good
at breaking down lipids
Benzoic acid in mouthwashes;
benzoic acid, sorbic acid, citric
acid, and ascorbic acid, in food
products
Gas Sterilants and Oxidizing Agents
• Gaseous Sterilants
• Denature proteins
• Ethylene oxide to sterilize hospital
equipment, disposable lab plasticware
• Other oxidizing Agents
• O3 (swim pools), H2O2(skin), Cl2 (swim pools), peracetic
acid (dialysis equipment), bleach (sodium hypochlorite),
bromine
• Steals electrons from biomolecules, causing breakdown
Microbial Characteristics and Microbial Control
Figure 7.11
Microbial Characteristics and Microbial Control
Chemical agent
Effectiveness against
Endospores
Mycobacteria
Phenolics
Poor
Good
Quats
None
None
Chlorines
Fair
Fair
Alcohols
Poor
Good
Glutaraldehyde
Fair
Good
Controlling Microbial Growth
What factors limit microbial growth? In what situations are large microbial numbers undesirable?
•
Concept of Microbial Control
•
Factors Which Affect Control
•
•
•
Temp., species type and status, environment
Physical Control Methods
•
Heat: Moist vs. Dry
•
Autoclaving, pasteurization
•
Filtration
•
Cold
•
Desiccation & high osmotic pressure
•
Radiation (UV, gamma rays)
Chemical Control Methods
•
Factors which influence effectiveness
• Dilution, time, pH, organic matter
•
Types of Disinfectants/Antiseptics
• Phenol and phenolics (e.g.. amphyl)
• Halogens (Chlorine, iodine, bromine)
• Alcohols (e.g.. isopropyl alcohol)
• Heavy metals (Ag, Hg, Cu, Zn)
• Surface active agents (soaps & detergents)
• Quaternary Ammonium Cmpds (quats)
• Aldehydes (e.g.. glutaraldehyde)
Microbial growth can be inhibited by temperature, moisture control or by antimicrobial chemicals