Transcript Microbiology

Chapter 7
The
Control of
Microbial
Growth
SLOs
 Define sterilization, disinfection, antisepsis, sanitization, biocide, germicide,
bacteriostasis, and asepsis.
 Describe the microbial death curve.
 Describe the effects of microbial control agents on cellular structures.
 Compare effectiveness of moist heat (autoclaving, pasteurization) vs .dry
heat.
 Describe how filtration, low temperature, high pressure, desiccation, and
osmotic pressure suppress microbial growth.
 Explain how radiation kills cells.
 List the factors related to effective disinfection.
 Interpret results the disk-diffusion test.
 Identify some methods of action and preferred uses of chemical disinfectant.
 Differentiate between two halogens used as antiseptics and disinfectants.
 List the advantage of glutaraldehyde and ethylene oxide over other chemical
disinfectants.
 Identify the method of sterilizing plastic labware.
 Explain how microbial control is affected by the type of microbe.
SLOs cont.: Check Your Understanding
 How is it possible that a solution containing a million bacteria would
take longer to sterilize than one containing a half-million bacteria?
 Would a chemical microbial control agent that affected plasma
membranes affect humans?
 How is microbial growth in canned foods prevented?
 What is the connection between the killing effect of radiation and
hydroxyl radical forms of oxygen?
 If you wanted to disinfect a surface contaminated by vomit and a
surface contaminated by a sneeze, why would your choice of
disinfectant make a difference?
 Why is alcohol effective against some viruses and not others?
 Is Betadine an antiseptic or a disinfectant when it is used on skin?
 What chemicals are used to sterilize?
 The presence or absence of endospores has an obvious effect on
microbial control, but why are gram-negative bacteria more resistant
to chemical biocides than gram-positive bacteria?
Terminology
Sepsis: microbial
contamination.
Asepsis: absence of significant contamination.
Aseptic surgery techniques prevent microbial
contamination of wounds.
Antimicrobial chemicals, expected to destroy
pathogens but not to achieve sterilization
 Disinfectant: used on objects
 Antiseptic: used on living tissue
Nosocomial
see
. . . More Terminology
Sterilization: Removal of all
microbial life (heat, filtration)
For food: Commercial sterilization to kill
C. botulinum endospores
Sanitization: reduces microbial numbers to safe
levels (e.g.: eating utensils)
Bacteriostatic: Inhibits bacterial
reproduction
Bactericidal: Kills bacteria
Fungicide, sporicide, germicide,
biocide
Remember Semmelweis,
Pasteur, and Lister from
Ch 1
Single most effective
measure!
Rate of Microbial Death
Bacterial populations subjected to heat or
antimicrobial chemicals die at a constant rate.
Microbial Death
Curve, plotted
logarithmically,
shows this
constant death
rate as a straight
line.
Rate: 90% / min
Foundation Fig 7.1
 How is it possible that a solution containing a million bacteria would
take longer to sterilize than one containing a half-million bacteria?
Foundation Fig 7.1 cont.
Effectiveness of Antimicrobial Treatment
Time it takes to kill a microbial population is
 to number of microbes.
Different microbial species and life cycle
phases (e.g.:_____________) have
different susceptibilities to physical and
chemical controls.
Organic matter may interfere.
Temperature determines exposure time:
Longer exposure to lower heat produces
same effect as shorter time at higher heat.
Actions of Microbial Control Agents
Alteration of membrane permeability
Damage to proteins
Damage to nucleic acids
Check your understanding:
Would a chemical microbial agent that affects
plasma membranes affect humans?
Physical Methods of Microbial Control
Moist Heat Sterilization
 ___________proteins
 Autoclave: Steam under pressure
 Most dependable sterilization method
 Steam must directly contact material to be sterilized.
 Pressurized steam reaches
higher temperatures.
 Normal autoclave conditions:
_____C for ___ min.
 Prion destruction:
132C for 4.5 hours
Fig 7.2
 Limitations of the autoclave
Pasteurization
Significant number reduction (esp. spoilage and
pathogenic organisms)  does not sterilize!
Historical goal: destruction of M. tuberculosis
Classic holding method: 63C for 30 min
Flash pasteurization (HTST): 72C for 15 sec.
Most common in US.
Thermoduric organisms survive
Ultra High Temperature (UHT):
140C for < 1 sec.
Technically not pasteurization because
it sterilizes.
Dry Heat Sterilization Kills by Oxidation
Flaming of loop
Incineration of carcasses
 Anthrax
 Foot and mouth disease
 Bird flu
Hot-air sterilization
Equivalent
treatments
Hot-air
Autoclave
170˚C, 2 hr
121˚C, 15
min
Filtration
Air filtration using high efficiency particulate
air (HEPA) filters. Effective to 0.3 m
Membrane filters for fluids.
 Pore size for bacteria: 0.2 – 0.4 m
 Pore size for viruses: 0.01 m
Compare to Fig 7.4
Low Temperature
 Slows enzymatic reactions  inhibits microbial
growth
 Refrigeration (watch out for _______________!
 Freezing forms ice crystals that damage microbial
cells
 Deep freezing and lyophilization
Various Other Methods

Desiccation prevents metabolism

Osmotic pressure causes plasmolysis
Ionizing Radiation
X-rays, -rays have short wave length
 dislodge e- from atoms  production
of free radicals and other highly reactive
molecules
Used for sterilization of heat sensitive
materials: drugs, vitamins, herbs, suture
material
Also as “cold pasteurization” of food 
Consumer fears!?
Nonionizing Radiation: UV light
 Effect:
thymine dimers
Actively dividing
organisms are
more sensitive
because thymine dimers cause ______________?
Used to fight air and surface contamination. Only
kills at close range and directly exposed
microbial agents
 E.g.: germicidal lamps in OR, cafeteria, and our lab ??
Nonionizing Radiation: Microwave
 Heats H2O
 Indirectly kills bacteria. How ?
 Solid food heats unevenly. Why?
Fig 7.5
Chemical Methods of Microbial Control
 Few chemical agents achieve sterility.
 Disinfectants regulated by EPA
 Antiseptics regulated by FDA
 Evaluating Disinfectants:
 Use-dilution test
 Disk-diffusion
method
Fig 7.6
Types of Antibacterial Chemicals
 Phenol = carbolic acid
(historic importance)
 Who used first?
 Many derivatives today:
 Phenolics, e.g.: Lysol
 Bisphenols, e.g.:
 Hexachlorophene (in
pHisoHex used in hospitals)
Fig 7.7
 Triclosan (toothpaste,
antibacerial soaps, etc.)
Phenol and derivatives disrupt plasma membranes
(lipids!) and lipid rich cell walls (??)
Remain active in presence of organic compounds
Halogens
Cl
I
Chlorine
 Oxidizing agent
 Widely used as disinfectant
 Forms bleach (hypochlorous acid) when added to water.
 Broad spectrum, not sporicidal (pools, drinking water)
Iodine
More reactive, more germicidal. Alters protein synthesis and
membranes.
Tincture of iodine (solution with alcohol)  wound
antiseptic
Iodophors: Iodine plus organic molecule. E.g.: complexed
with detergent: Betadine®. Occasional skin sensitivity.
Alcohols
 Ethyl (60 – 80% solutions)
and isopropyl alcohol
 Denature proteins, dissolve
lipids
 No activity against spores
and poorly effective against
viruses and fungi
 Easily inactivated by
organic debris
 Also used in hand sanitizers
and cosmetics
Heavy Metals
Oligodynamic action: toxic effect due to metal ions
combining with sulfhydryl (—SH) and other functinal
groups  proteins are denatured.
 Silver (1% AgNO3): Antiseptic for
eyes of newborns
 Copper against chlorophyll
containing organisms
 Algicides;
also X-gel hand sanitizer
 Zinc (ZnCl2) in mouthwashes,
ZnO as antifungal in paint
Surface Acting Ingredients / Surfactants
 Soaps and Detergents
 Major purpose of soap: Mechanical removal and use
as wetting agent
 Definition of detergents
Acidic-Anionic detergents Anion reacts with plasma membrane.
Nontoxic, non-corrosive, and fast acting. Laundry soap, dairy industry.
Cationic detergents  Quarternary ammonium compounds (Quats).
Strongly bactericidal against wide range, but esp. Gram+ bacteria
Soap
Degerming
Acid-anionic detergents
Sanitizing
Quarternary ammonium compounds
(cationic detergents)
Strongly bactericidal, denature
proteins, disrupt plasma membrane
Chemical Food Preservatives
Sodium nitrate and
nitrite prevent endospore
Prevents ES germination. Used in meats.
Conversion to nitrosamines: Carcinogenic!
Organic acids
Inhibit metabolism
E.g.: Sorbic acid,
benzoic acid, etc.
In foods and cosmetics
Sulfur dioxide
wine
Aldehydes and Chemical Sterilants
Aldehydes (alkylating agents)
 Inactivate proteins by cross-linking
with functional groups
(–NH2, –OH, –COOH, –SH)
Formaldehyde:
 Embalming  Formalin
 Virus inactivation for vaccines
 Glutaraldehyde: Liquid Sterilant
for delicate surgical instruments
(Kills S. aureus in 5, M. tuberculosis in
10 min, ES in 3 – 10h)
 Ethylene oxide: Gaseous Sterilant
Hydrogen Peroxide: Oxidizing agent
Inactivated by catalase 
Not good for open wounds
Good for inanimate objects; packaging for food
industry (containers etc.)
3% solution (higher conc. available)
Especially effective against anaerobic
bacteria (e.g.:
Effervescent action, may be useful for wound
cleansing through removal of tissue debris
Microbial
Characteristics and
Microbial Control
Fig 7.11