Microbio - Module 6: Control of Microorganisms in the Environment

8.1 Microbial Growth and Replication: Targets for Control

Common Microbial Control Methods

8.1 Microbial Growth and Replication: Targets for Control

whats biocide?

Antimicrobial agents that control microorganisms.

• Physical, chemical, mechanical or biological

8.1 Microbial Growth and Replication: Targets for Control

Define these Frequently Used Terms:

1. Sterilization

2. Disinfection

   1. Disinfectants

3. Sanitization

4. Antisepsis

   1. Antiseptics

Sterilization—process by which all living cells, spores, and acellular entities are destroyed or removed from an object.

• Sterilant is the chemical agent.

Disinfection—killing, inhibition, or removal of disease-causing microorganisms.

• Disinfectants—agents, usually chemical, used for disinfection, usually used on inanimate objects.

• Does not always sterilize as spores or a few organisms can remain.

Sanitization—reduction of microbial population to levels deemed safe by public health standards.

Antisepsis—destruction of microbes on living tissue.

• Antiseptics—chemical agents applied to tissue to kill or inhibit growth of pathogen.

8.1 Microbial Growth and Replication: Targets for Control

Antimicrobial Agents

Chemotherapy—generic term that describes application of

chemicals to kill microorganisms.

Cidal agents kill (-cide suffix indicates agent that kills).

• May be especially effective against a specific group.

• Include bactericides, fungicides, and viricides.

Static agents inhibit growth (-static suffix indicates growth

inhibiting agent).

• May be especially effective against a specific group.

• Include bacteriostatic and fungistatic

8.1 Microbial Growth and Replication: Targets for Control

Resistance to Biocides

Resistance to antimicrobial biocides has been increasing

similar to the rate of resistance to antibiotics.

Mechanisms of resistance similar to antibiotic resistance.

• Efflux pumps

• Alter membrane permeability

• Modify the target

• Specific resistance genes

8.1 Microbial Growth and Replication: Targets for Control

The Pattern of Microbial Death Mirrors the Pattern of

Microbial Growth

Microorganisms are not killed instantly when exposed to lethal agent.

Population death usually occurs exponentially.

• Reduced by the same fraction at constant intervals.

Decimal reduction time (D value): time required to kill 90%

of microorganisms.

• D value is time required to drop by 10-fold.

• The D-value, which denotes the decimal reduction time, is the time required at a specific temperature and under specified conditions to reduce a microbial population by one decimal. The decimal reduction time is dependent on the temperature, the type of microorganism and the composition of the medium containing the microorganism. Thus, after an organism is reduced by 1 D, only 10% of the original organisms remain. The population number has been reduced by one decimal place in the counting scheme. When referring to D values it is proper to give the temperature as a subscript to the D. For example,

a hypothetical organism is reduced by 90% after exposure to temperatures of

300ºF for 2 minutes, Thus the D-value would be written as D300ºF = 2 minutes.

8.1 Microbial Growth and Replication: Targets for Control

The Z-value

• The Z-value is the increase or decrease in temperature required to reduce or increase the decimal reduction time by one decimal.

• It is a measure of the change in death rate with a change in temperature. The number of degrees Fahrenheit or Centigrade required for a thermal death time curve to traverse 1 log cycle.

• The z-value gives an indication of the relative impact of different temperatures on a microorganism, with smaller values indicating greater sensitivity to increasing heat.

• Z value is the temperature change that decreases the microbial population by 90%

8.1 Microbial Growth and Replication: Targets for Control

D and Z Values

8.1 Microbial Growth and Replication: Targets for Control

Impact of Biocide Exposure

Three possible population reduction curves from different

biocides.

8.2 Microbes Can Be Controlled by Physical Means

Filtration

Reduces microbial population in heat-

sensitive materials by removing

microorganisms.

Also used to reduce microbial

populations in air.

Depth filter—fibrous materials that

have been bonded into a think layer

filled with narrow, twisting channels.

• Solution with microbes is sucked

through the layer under vacuum

and microbes adsorb to the surface

of the filter material

8.2 Microbes Can Be Controlled by Physical Means

Filtering Liquids

describe membrane filters

Membrane filters

• Porous membranes with defined

pore sizes that remove

microorganisms.

• Often used after a depth filter.

8.2 Microbes Can Be Controlled by Physical Means

Filtering air

Surgical masks

• N95 disposable mask

• Exclude 95% of particles.

High-efficiency particulate air (HEPA) filters

• Used in laminar flow biological safety cabinets.

• Exclude 99.97% of particles.

• During the COVID-19 pandemic, airlines showed this filter to be effective.

8.2 Microbes Can Be Controlled by Physical Means

Moist Heat

Destroys viruses, fungi, and bacteria by degrading nucleic

acids, denaturing proteins, and disrupting cell membranes.

Boiling will not destroy endospores and does not sterilize.

• Can be used to disinfect drinking water.

Average conditions for moist heat inactivation: (look at picture)

8.2 Microbes Can Be Controlled by Physical Means

Steam Sterilization

Autoclave—device used in steam

sterilization.

Carried out above 100oC which

requires saturated steam under

pressure.

Effective against all types of

microorganisms, including spores.

Quality control

• Geobacillus stearothermophilus

strips or color changing tape

8.2 Microbes Can Be Controlled by Physical Means

Pasteurization

• Controlled heating at temperatures well below boiling.

• Used for milk, wine, and other beverages.

• Process does not sterilize, but does kill pathogens and

slows spoilage by reducing the total load of organisms

present

8.2 Microbes Can Be Controlled by Physical Means

Tyndallization

For materials that cannot withstand the high temperature of

the autoclave.

Intermittent sterilization

• 30 to 60 minutes of steam exposure with 23 to 24 hours

between incubations.

• 2 or 3 times this is repeated

8.2 Microbes Can Be Controlled by Physical Means

Dry Heat Sterilization

Less effective than moist heat sterilization, requiring higher

temperatures and longer exposure times.

• Items subjected to 160 to 170oC for 2 to 3 hours.

Oxidizes cell constituents and denatures proteins which

causes microbial death.

Not suitable for heat-sensitive materials (that is, plastic)

Advantage

• Does not corrode glassware and metal instruments

8.2 Microbes Can Be Controlled by Physical Means

Ultraviolet (UV) Radiation

which wavelength is the most lethal?

what does it cause, and this ultimately prevents?

its limitations?

used for what treatment?

• Wavelength of 260 nm is most lethal.

• Causes thymine dimers preventing replication and

transcription.

• UV limited to surface sterilization because it does

not penetrate glass, dirt films,

water, and other substances.

• Has been used for water treatment

8.2 Microbes Can Be Controlled by Physical Means

Cold Plasma

what state of matter?

how is it made?

What is RONS?

what are its key benefits?

QUESTION FR HERE

Fourth state of matter.

Made of ionized gases operating near room temperature (35-40 deg)

where the atoms and/or molecules are stripped of outer-shell electrons.

Generated by applying high-voltage electric fields to gases like helium or

argon, producing reactive oxygen and nitrogen species (RONS) that effectively

kill bacteria, viruses, and fungi by damaging their cell wall and DNA.

• Damages nucleic acids

• Oxidizes nucleic acids, proteins, and lipids.

• Safe for direct contact with skin and biological tissues.

• Key benefits include accelerated wound healing, acne treatment, skin

regeneration, and sterilization without thermal damage.

8.2 Microbes Can Be Controlled by Physical Means

Ionizing Radiation

Explain how gamma radiation penetrates deep and what is it used for?

Gamma radiation penetrates deep into objects.

• Dislodges electrons from atoms or molecules, producing

chemically reactive free radicals.

• Used for sterilization and pasteurization of antibiotics,

hormones, sutures, plastic disposable supplies, and food.

8.3 Microorganisms Are Controlled with Chemical Agents

Chemical Control Agents and Their Requirements

explain disinfection and Antisepsis

Disinfection

• Ideally the biocide is effective against wide variety of

infectious agents at low concentrations and in the

presence of organic matter.

• Balance between effectiveness and low toxicity.

Antisepsis

• Reduce number of pathogens on human tissues to

prevent infection.

8.3 Microorganisms Are Controlled with Chemical Agents

Phenolics

what are they commonly used for?

How do they act?

Name an example of this in real world

• Commonly used as laboratory and hospital disinfectants.

• Act by denaturing proteins and disrupting cell membranes.

• Tuberculocidal, effective in presence of organic material, and

long lasting.

• Lysol®, a commercial disinfectant, is a mixture of phenolics

8.3 Microorganisms Are Controlled with Chemical Agents

Alcohols

what are the two most common alcohols?

They are not effective for which ones—bactericidal,fungicidal, sporicidal

how do they act?

• Among the most widely used disinfectants, antiseptics,

and sanitizers.

• Two most common are ethanol and isopropanol.

• Bactericidal, fungicidal, but not sporicidal.

• Inactivate some viruses.

• Act by denaturing proteins and dissolving membrane

lipids.

8.3 Microorganisms Are Controlled with Chemical Agents

Halogens—Iodine

• what kind of antiseptic?

• how does it act?

• what can it cause?

• What is iodophor?

Skin antiseptic.

Acts by oxidizing cell constituents and iodinating proteins.

At high concentrations may kill endospores.

May cause skin damage, staining, and allergies can be a

problem.

Iodophor

• Iodine complexed with organic carrier.

• Released slowly to minimize skin burns

8.3 Microorganisms Are Controlled with Chemical Agents

Halogens—Chlorine

how does it act?

important in disinfecting what?

Why’s it so well-loved as a household disinfectant?

• Acts by oxidizing cellular materials and destroys

vegetative bacteria and fungi.

• Important in disinfection of water supplies, swimming

pools, and used in dairy and food industries.

• Chlorine is a household disinfectant as well because it is

inexpensive and effective

8.3 Microorganisms Are Controlled with Chemical Agents

Metals

• Ions of mercury, silver, arsenic, zinc, and copper were used for many years as ___.

• whats used topically on burns?

• Copper sulfate is an effective algicide for what?

Ions of mercury, silver, arsenic, zinc, and copper were used

for many years as germicides.

• Silver and copper only used now.

Silver sulfadiazine is used topically on burns.

Copper sulfate is an effective algicide in lakes and swimming

pools.

Act by inactivating proteins, often via sulfhydryl groups

8.3 Microorganisms Are Controlled with Chemical Agents

Quaternary Ammonium Compounds

1. detergents that have broad spectrum antimicrobial activity are ____ disinfectants

2. what are detergents?

3. ___ r detergents are effective disinfectants?

4. does it kill bacteria and endospores?

ON EXAM

Detergents that have broad spectrum antimicrobial activity and are

effective disinfectants.

• Detergents—amphipathic organic cleansing agents.

Cationic detergents are effective disinfectants.

• Kill most bacteria, but not M. tuberculosis or endospores.

• Stable and nontoxic, but inactivated by hard water and soap

8.3 Microorganisms Are Controlled with Chemical Agents

Aldehydes

• Commonly used agents are formaldehyde and

glutaraldehyde.

• Highly reactive molecules that act by inactivating nucleic

acids and proteins.

• Sporicidal and can be used as chemical sterilants.

8.3 Microorganisms Are Controlled with Chemical Agents

Effects of Glutaraldehyde

ON EXAM

8.3 Microorganisms Are Controlled with Chemical Agents

Sterilizing Gases

1. sterilizes what materials?

2. it can kill what?

3. name an example of one. and used in what equipment?

• Used to sterilize heat-

sensitive materials.

• Microbicidal and sporicidal. (kills them)

• Ethylene oxide sterilization

is carried out in equipment

resembling an autoclave.

• Vaporized hydrogen

peroxide can also be used.

8.4 Antimicrobial Agents Must Be Evaluated for Effectiveness

Evaluation of Antimicrobial Agent Effectiveness

1. what is this?

2. who regulates disinfectants?

3. who regulates agents used on humans and animals and establishes guidleines under which of these agents are used and agent effectiveness is measured?

Complex process regulated by two U.S. federal agencies:

• Environmental Protection Agency regulates disinfectants.

• Food and Drug Administration (FDA) regulates agents used on humans and animals.

Establish guidelines under which these agents are used and agent effectiveness is measured.

8.4 Antimicrobial Agents Must Be Evaluated for Effectiveness

Conditions Influencing the Effectiveness of Antimicrobial Agent Activity

1. what are they? (6)

Population size

• Larger populations take longer to kill than smaller

populations.

Population composition

• Microorganisms differ markedly in their sensitivity to

antimicrobial agents (endospores more resistant).

Concentration or intensity of an antimicrobial agent

• Usually higher concentrations kill more rapidly.

• Not a linear relationship, small changes can have large effect

Contact time

• The longer the exposure, the more organisms killed.

Temperature

• Higher temperatures enhance chemical activity.

Local environment

• Population to be controlled is not isolated but surrounded

by environmental factors that can either prevent or aid in

destruction.

• pH, viscosity, and concentration of organic matter.

• Organisms in biofilms are less susceptible to many antimicrobial

agents.

8.4 Antimicrobial Agents Must Be Evaluated for Effectiveness

Efficiency Evaluation of Chemical Agents

1. three tests

Phenol coefficient test

• Potency of a disinfectant is compared to that of phenol.

• Useful for initial screening but may be misleading.

• test not 100% correct so u need another test to add on to it

Use dilution test

• Determines rate at which selected bacteria are destroyed

by various chemical agents.

Normal in-use testing

• Testing done using conditions that approximate normal

use of disinfectant

8.5 Microorganisms Can Be Controlled by Biological Methods

Biological Control of Microorganisms

1. what is this?

2. what are bacteriophages?

3. what are enzybiotics?

Emerging field showing great promise.

basically abt using natural organisms or substances to kill harmful microorganisms, especially bacteria istead of using traditional antibiotics or chemicals

Natural control mechanisms:

• Predation by Bdellovibrio.

• Bacteriophages—A spray format recently approved by the FDA can be used on food

products.

• Enzybiotics—Proteins purified from bacteriophage that cause host cell lysis.

• Toxin-mediated killing using bacteriocins

8.5 Microorganisms Can Be Controlled by Biological Methods

Enzybiotics

1. what are they?

2. What are their advantages over Conventional Antibiotics?

ON EXAM

Enzybiotics are a promising class of protein-based, antibacterial, and antifungal agents derived from bacteriophage-encoded enzymes (endolysins) or microorganisms that degrade bacterial cell walls.

• proteins/enzymes tht kill bacteria by damaging their cell wall

Advantages over Conventional Antibiotics:

• High Specificity: They target specific pathogens without destroying beneficial microbiota.

• Low Resistance: Due to their mechanism, bacteria find it difficult to develop resistance.

• Effective against Persisters: They can kill metabolically inactive ("persister") bacteria that conventional antibiotics often miss.

• Biofilm Disruption: Some, such as polysaccharide depolymerases, can break down protective biofilms.

Applications: Primarily under development for treating infections (e.g., Staphylococcus aureus, Pseudomonas aeruginosa), with some in clinical trials. They are also researched for food bio-preservation to reduce pathogens.

Limitations: Their proteinaceous nature can limit stability, and they can be challenging to

produce and purify.