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

63oCfor30minutes
72oCfor15seconds
140oCfor1second
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

170C, 2 hr

121C, 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 manufacturers instructions) for
min at 20C
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

Ophenylphenol
(Lysol,Amphyl)

Hexachlorophene

Thymol
PineSol/pine oil
(turpineol)

Halogens
Bromineandchlorineproducesarestrongoxidizers
Iodinedenaturesproteinsbybreakingdisulfidebonds
Tinctureof
Iodine(I2);
iodine
denatures
proteinsby
breaking
disulfidebonds

Sodium
hypochlorite
(NaOCl)in
bleachisa
strongoxidizing
agent.

Povidoneiodines(I
linkedtoorganic
molecule)asinBetadine;
denaturesproteinsby
breakingdisulfidebonds

Brominetablets
(BCDMH
bromochloro
dimethyl
hydantoin)usedin
hottubs,pools,
killsbyoxidation

Chlorinegas
usedinswim
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

Denatureproteinsbybreakingdisulfidebonds

Mercurachrome

antisepticforwounds

Coppersulfate(CuSO4)

algicide

Zincchloride(ZnCl2)

mouthwashes

Silvernitrate(AgNO3)

antisepticforeyes,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

Benzalkoniumchloride
(akaZephiran)
Roccal:labdisinfectant
Cationic detergents

Cetylpyridiniumchloride(Cepacol)

Types of Disinfectants:Aldehydes
Aldehydes
Inactivate proteins by cross-linking with functional
groups (NH2, OH, COOH, SH)
Glutaraldehyde, formaldehyde

Disinfectionofhospitalequipment,
especiallyinrespiratorytherapy,for
kidneydialysismachines,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