CFU
What are some ways you could make water safe to drink?
Controlling
Microbial GrowthPhysical Measures
Oliver Yuan
BIOL 240
Learning Objectives
Be able to order vocabulary in terms of most clean to least
Calculate D-value from a microbial death curve
Vocabulary
Sepsis: microbial contamination
Asepsis: the absence of significant contamination
Aseptic surgery techniques prevent the microbial contamination of
wounds
Antisepsis: destroying harmful microorganisms from living tissue
Sanitization: lowering microbial counts to safe levels for public health
Commercial sterilization: removal of foodborne pathogens while
maintaining food quality
Killing Clostridium botulinum endospores from canned goods
How Clean Does it Need to Be?
Item
Cleaning Procedure
Critical item
Sterilization: removing and
destroying ALL microbial life,
including endospores and viruses
Semicritical item
Thorough disinfection: destruction
of most harmful microorganisms
from a fomite
Noncritical item
Degerming: mechanical removal
of microbes from a limited area
Example
Controlling Bacteria
Biocide (germicide): treatments that kill microbes
Bactericidal agents: Chemicals result in bacterial death in the
culture, no growth
Subculture: no bacteria grows
Bacteriostatic agents: Chemicals cause temporary inhibition of
growth, no growth
Subculture: bacteria grow
Effectiveness of Treatment
Depends on:
Number of microbes
Environment (organic matter, temperature, biofilms)
Time of exposure
Microbial characteristics
Microbial Death Curve
D-value (decimal
reduction time):
time it takes to
reduce microbe
population by
one magnitude
(90%)
CFU
What is the Dvalue of this
agent?
Physical Methods of Microbial
Control
Heat
Filtration
Low Temperatures
High pressure
Desiccation
Osmotic pressure
Heat
Thermal death time (TDT): shortest time during which all cells in a
culture are killed at a given temperature
Thermal death point (TDP): lowest temperature at which all cells in a
culture are killed in 10 min
Moist Heat Sterilization
Moist heat denatures proteins
Boiling
Autoclave: steam under pressure
that kills all organisms and
endospores
121°C at 15 psi for 20 min
Steam must contact the item's
surface
Pasteurization
Reduces spoilage organisms and
pathogens
Equivalent treatments
63°C for 30 min
High-temperature short-time (HTST):
72°C for 15 sec
Ultra-high-temperature (UHT): 138°C
for 2-4 sec
Dry Heat Sterilization
Kills by oxidation
Flaming
Incineration
Hot-air sterilization
Filtration
Passage of substance through a
screen-like material
Used for heat-sensitive materials
High-efficiency particulate air (HEPA)
filters: remove microbes >0.3 μm
Membrane filters remove microbes
>0.22 μm
Physical Methods of Microbial
Control
Low temperature inhibits microbial growth
Refrigeration
Deep-freezing
Lyophilization: freezing dry
High pressure denatures proteins
Desiccation prevents metabolism
Osmotic pressure causes plasmolysis
Radiation damages DNA
Controlling Microbial
Growth- Cleaning Agents
and Antimicrobial Drugs
Oliver Yuan
BIOL 240
Learning Objectives
Compare and contrast different kinds of cleaning agents, their
mechanisms, and their evaluation of effectiveness
Chemical Methods of Microbial
Control
Phenol and Phenolics
Bisphenols
Alcohols
Halogens
Heavy Metals
Surfactants
Phenol and Phenolic Compounds
Disrupt plasma membranes and denaturing proteins
a) Phenol b) o-Phenylphenol c) hexachlorophene (pHisoHex)
Triclosan: another bisphenol
Inhibits an enzyme needed for the biosynthesis of fatty acids
Effective against gram-positive, certain gram-negative and yeasts
FDA banned in 2016 from soaps and body washes
Alcohols
Denature proteins and disrupting membranes
No effect on endospores and nonenveloped viruses
70% works better than 100% since the latter coagulates proteins too
fast
Halogens and Heavy Metals
Halogens
Iodine: Alters protein synthesis and
membranes
Chlorine
Bleach
Oxidizing agents
Heavy Metals
Ag, Hg, and Cu
Bind to and denature proteins
Oligodynamic: very effective at very
low concentrations
For a disk diffusion assay, the zone of
inhibition is:
ⓘ Start presenting to display the poll results on this slide.
CFU
For a disk diffusion assay, the zone of inhibition is:
A.
How far the antimicrobial agent spread into the agar
B.
The growth around a disk with an antimicrobial agent
C.
Measured in mm
D.
The same for all antimicrobial agents for a given species
CFU
Why is 70% ethanol a more effective antimicrobial agent than 100%
ethanol?
Surfactants
Soap: salts with hydrophobic tails; only degerming
Quaternary ammonium salts (quats): Disrupt membrane integrity
Effective against fungi, protozoans, and enveloped viruses, but
endospores are unaffected
Chemical Food Preservatives
Sulfur dioxide (SO2) prevents wine spoilage
Organic acids
Inhibit metabolism
Sorbic acid, benzoic acid, and calcium propionate prevent molds in
acidic foods
Nitrites and nitrates prevent endospore germination in processed
meat
Creates carcinogenic nitrosamines when heated
Evaluating a Disinfectant
Use-Dilution Tests
Metal cylinders are dipped in test
bacteria and dried
Cylinders are placed in disinfectant
for 10 min at 20°C
Cylinders are transferred to fresh
culture media without disinfectant to
determine whether the bacteria
survived treatment
Disk Diffusion Method
Filter paper disks are soaked in a chemical and placed on a culture
Look for zone of inhibition around disks
Antimicrobial Drugs
Compound produced by microorganisms that inhibit growth of
other microorganisms
Selective toxicity: specifically inhibits or kills microbial targets without
harm to the host
Actions of antimicrobial drugs
Alteration of membrane permeability
Damage to proteins (enzymes)
Damage to nucleic acids
Damage to cell walls
Alteration of metabolic pathways
Targets of Antibacterial Drugs
Evaluating Antibacterial Drugs
Kirby-Bauer Disk Diffusion Test
Filter paper disks are soaked in antimicrobial drug and placed on
Mueller-Hinton agar
Look for zone of inhibition around disks
Variability in Disk Diffusion Test
Water solubility of agent
Ability to diffuse into agar
Concentration of agent
Concentration of agar
Type of bacteria selected
Evenness of agar and of bacterial lawn
Other Antimicrobial Drugs
More difficult to treat, either because they are eukaryotes or because
they use much of the host’s own machinery
Antifungal
Antiprotozoan
Quinoline: chloroquine thought to disrupt heme breakdown in malaria and
Entamoeba histolytica
Antihelminth
Imidazole: Disrupts cell membrane by targeting ergosterol in Candida
vaginal yeast, athlete’s foot, etc.
Praziquantel: thought to cause influx of Ca2+ into tapeworms and flukes
(Schistostoma), paralyzing them
Antiviral
Remdesivir: adenosine analog that binds to RdRP and initiates termination;
hepatitis C, previous emergency treatment for COVID-19
0
