MIC 205: Chapter 9 - Control of microbial growth

Describe the number of living microbes over time with the environmental conditions being temperature at 45c and 20c

At 45c, the number of living microbes rapidly declines in ~1 minutes
At 20c, the number of living microbes slowly declines in ~8 minutes
This shows that higher temperature causes a decrease in living microbes in a short amount of time

What are the 4 mode of action for antimicrobial agents (ways antimicrobial agents cause damage)

damage to cell wall (peptidoglycan), damaged cell membrane, damaged proteins cause misfolding or to denature, and damaged nucleic acid

What is damaged to the cell wall (peptidoglycan) in mode of action: antimicrobial agents

Damaged to the cell wall weaken the cells rigidity (rigidity decreases), thus the cell will burst or lysis due to osmotic effects

What is damaged cell membrane in mode of action: antimicrobial agents

Membrane controls permeability and damage to cell membrane means poking holes in membrane.
- Damaged cell membrane will prevent the control passage of molecules in and out of the cytoplasm.
- Loss of proton gradient & electron transport chain will not function = no ATP production
- Some viruses are encased in envelope (phospholipid bilayer) necessary for attachment to target cells; damaged envelope interrupts viral replication
Summary: no membrane passage control, no ATP or ETC, and damage to nuclear envelope

What is damaged proteins cause misfolding or to denature in mode of action: antimicrobial agents

If enzymes are unable to function (due to denaturing), this will inhibit glycolysis, TCA, ETC, DNA replication, TXC, TXL…
So, no ATP or proteins are created for binary fission

What is damaged nucleic acids in mode of action: antimicrobial agents

Alterations to the DNA/RNA will inhibit DNA replication, TXC, TXL…
So, no new proteins or DNA will be created for binary fission
(Since nucleic acids are damaged, DNA can’t replicate and occur so no binary fission will happen)

What is a test to evaluate disinfectants and antiseptics and what is its purpose

Diffusion susceptibility tests is a method used for testing the effectiveness of antimicrobial agents on specific microbes

What are the steps to the diffusion susceptibility test

- Swabs taken from objects before and after application of disinfectant or antiseptic
- Swabs inoculated into growth medium and incubated
- Medium monitored for growth
- Accurate determination of proper strength and application procedure for each specific situation

Why does rubbing alcohol not show on a diffusion susceptibility test

It evaporates

What are the physical methods of microbial control

Exposure to extremes of heat (either dry or moist heat) and cold, desiccation, filtration, and radiation (either nonionizing or ionizing)

What are the effects of high temperatures on microbes in hear-related methods for microbial control

Denaturation of proteins
Interference with the integrity of cell walls
Interference with the integrity of membranes
Disruption of structure and function of nucleic acids

What is moist heat sterilization

Moist heat is more effective than dry heat because water is a better conductor of heat than air.

What is autoclaving and its conditions (part of moist heat sterilization)

Process applies pressure to boiling water to create moist heat conditions. Autoclave conditions: 121 degrees C, 15 psi, time depends on load
Autoclaving will cause instruments to become dull

What happens with the method of sterilization for media and lab equipment with moist heat sterilization

Material will leave the autoclave with condensation, such as glassware. Liquids will not lose water (volume) during sterilization

What is dry heat sterilization

Dry heat sterilization involves 2 hours at more than 180 degrees C (longer hours at higher temperatures)
Dry heat is non-toxic and does not harm the environment
It is easy to install and is cheap
It is noncorrosive for metal and sharp instruments
It is a time consuming method because of a slow rate of heat penetration and microbial killing

What are examples of why dry heat is not suitable for most materials and how much time/temperature do you need

Plastic and rubber items cannot be dry-heat sterilized because temperatures used (160-170c) are too high for these materials.
Time and temperature required will vary for different substances, and overexposure may ruin some substances.

What is boiling used to kill in heat sterilization

It kills vegetative cells of bacteria, fungi, protozoa, and most viruses within 10 minutes at sea level (100c), Denver (95c), and Flagstaff (92.7c)
(10 mins is suffice to kill most things)

Do different elevation levels change boiling temperature and does boiling kill everything

Yes elevation changes boiling temperature.
boiling does not kill everything → endospores, protozoan cysts, and some viruses survive boiling

What is pasteurization (this is not a form of sterilization)

This disinfect food-releated products. It is a common method of killing pathogenic microbes in milk, ice cream, yogurt, and fruit juices.
Pasteurization is not sterilization as heat-tolerant and heat-loving microbes do survive.

What is incineration

A mean of sterilization to kill all microbes, even prions (a specialized microbe). Incineration is the only way to remove a prion.
Prions cause mad cow disease and the only way to get rid of it is to incinerate the cows.
Incineration is the ultimate means of sterilization!!

What is refrigeration and what type of process is it

- It inhibits growth by decreasing rates of metabolism and reproduction
- It is only a -static process (inhibits growth) not -cidal (kills)
- Chemical reactions occur slower at lower temperatures as enzymes are less functional.
- Liquid water is less readily available

What is resistant to refrigeration

Bacteria because they are unicellular

What is freezing

Inhibits growth by decreasing rates of metabolism and reproduction
Chemical reactions occur slower at lower temperatures
Liquid water is NOT readily available

What can freezing lead to and an example with lettuce

Freezing can lead to cell death.
Ex: Ice crystal formation between and within cells punches holes in walls and membranes of lettuce and the cells burst/lysis upon thawing
Effects depend on the speed of cooling and the presence/absence of cryoprotectants
Organisms vary in susceptibility to freezing-induced death. this is problematic in multi-cellular organisms

What is lyophilization

Freeze-drying and can prevent ice crystal formation by removing water at cold temperatures. This is used for long-term preservation of microbial cultures
Lyophilization is a static process and by adding water to yeast it activates it

What is desiccation

Inhibits growth due to removal of water, due to hypertonic environment (cell shrink)
(basically removing water out of microbe/cell)

What type of process is desiccation and why

Only a -static process, not -cidal because:
Tough bacterial cell walls and capsules prevent complete desiccation and preserve cell structure.
Dehydrated microbes and microbial structures (endospores, cysts) can be rehydrated and brought back to life

What is salt induced desiccation

Osmotic pressures can cause desiccation when the environment is hypertonic (cell shrinkage → water moves out) with respect to microbial cytoplasm

What is filtration and when is it useful

An effective and reasonable economical method of sterilization used to sterilize heat-sensitive fluids and air
It is particularly useful for solutions containing toxins, enzymes, drug, serum, and sugars

What are filters: Air (HEPA)

High efficiency particulate air filters (like the hood in science labs and filters in biological safety cabinets)

What is filtration: mask

Pores on the mask that allows less spread of viruses coming from an individual

What are viruses measured in

Nanometer

What are the 2 forms of radiation

Ionizing and nonionizing radiation

What does radiation effect and what is it commonly employed for

Radiations forms will affect on cellular structure of the cells.
Radiation is commonly employed for sterilizing heat sensitive materials like disposable plastic products and materials that cannot withstand moisture

What is ionizing radiation

Wavelengths shorter than 1nm like X-rays that break phosphodiester bonds
The energy creates ions that disrupts hydrogen bonding, oxidize covalent bonds, DNA, and creates hydroxyl radicals
Examples: electron beams, gamma rays, some x-rays

What is nonionizing radiation

Wavelengths greater than 1nm
Excites electrons, causing them to make new covalent bonds → affects nucleic acid by causing thymine dimers and denaturing proteins
Suitable for disinfecting air, transparent fluids, and surfaces of objects
Example: UV light absorbed by DNA and cause thymine dimers

What are chemical methods of microbial control (what are they more effective against, affect and effect)

Tends to be more effective against enveloped viruses and vegetative cells of bacteria, fungi, and protozoa
It affects microbes cell walls, cytoplasmic membranes, proteins, or DNA
Effects varies with temperature, pH, length of exposure, amount of organic matter, and concentration and age of chemical
3 things to consider: how long, at what concentration, and how much organic matter

What are phenols and phenolics (part of chemical methods of microbial control)

They are good at denaturing proteins and disrupt cell membranes.
Effective in the presence of organic matter and remain active for a prolonged time
Commonly used in healthcare setting, labs, and homes

What are alcohols (part of chemical methods of microbial control)

Denature proteins and disrupt cell membranes by removing water from cell
Evaporate rapidly → both advantageous and disadvantageous, but have high penetration and low contact time
Swabbing of skin with 70% alcohol prior to injections removes most microbes but is not effect against fungal spores or bacterial endospores
*washing hands is better than sanitier

What is halogens (part of chemical methods of microbial control)

Damage enzymes by denaturation of proteins
1 of the most diverse and broadly applied chemical groups
Example: Cl, F, Br, I (what u use before surgery)

What is heavy metals (part of chemical methods of microbial control)

Denature proteins that alter the 3D shape of proteins, inhibiting or eliminating their function
Specific application: 1% silver nitrate prevents blindness, mercury as preservative in vaccines, & copper for controlling algal growth in reservoirs, fish tanks, pools, water storage tanks

What is surfactants (part of chemical methods of microbial control)

- Surface active chemicals that reduce surface tension of solvents to make them more effective at dissolving solutes
- Surfactant is -static as it has no killing abilities
- Ex: soaps and detergents → soaps have hydrophilic and hydrophobic ends as they are a good degerming agent but not antimicrobial & detergents are positively charged organic surfactants
*removes surface tension to get washes away

What is quaternary ammonium compounds (quats)

This is still surfactants but it disrupts cellular membranes and is ideals for many medical and industrial applications

What is oxidizing agents (part of chemical methods of microbial control)

Toxic forms of O2 and it disrupts cellular membrane
Kill by oxidation of microbial enzymes and cell membrane lipids which causes denaturing of the proteins enzymes in high-level disinfectants and antiseptics
Example: H2O2, Ozone, Peroxides, and peracetic acid

What can hydrogen peroxide be used for in oxidizing agents (part of chemical methods of microbial control)

Can disinfect and sterilize surfaces. It is less useful for treating open wounds due to catalase activity (catalase will be on exam!!)

What is aldehydes (part of chemical methods of microbial control)

Cross-link functional groups to denature proteins and inactivate nulceic acids.
It is frequently used to fix tissues, disinfect rooms and sterilize instruments
Example: formaldehyde (perservation agent), glutaraldehyde, and formalin

What is gaseous agents (part of chemical methods of microbial control)

Denature proteins and DNA by cross-linking functional groups
It can be hazardous to people and is high explosive, poisonous, and carcinogenic.
It is usually only used in closed chambers to sterilize items in hospitals and dental offices
Example: ethylene oxide

What are enzymes (part of chemical methods of microbial control)

Antimicrobial enzymes act against microorganisms
Like how human tears contain lysozyme that degrade/digest peptidoglycan cell wall bacteria

What are the 2 ways enzymes can be used to control microbes in the environment

Lysozyme used to reduce the number of bacteria in cheese
Prionzyme can remove prions on medical instruments

What are antimicrobial agents (part of chemical methods of microbial control)

Was first synthesized in 1909 in Paul Ehrlich lab as an antimicrobial chemical. There are 3 antimicrobial agents: antibiotics, semisynthetic antibiotics, and synthetic chemicals

What are antibiotics used for

The treatment of disease but some are used for antimicrobial control outside of the body. Potential for very specific, targeted uses

What is a drawback of antibiotics

Antimicrobial resistance

What makes a good antimicrobial agent

Selective toxicity: a drug that kills harmful microbes without damaging the host

What structures/enzymes are only found in microorganisms in cell wall synthesis

Cell wall synthesis → penicillins, cephalosporins, and bacitracin

What structures/enzymes are only found in microorganisms in protein synthesis

Protein synthesis → erythromycin, tetracycline, and streptomycin

What structures/enzymes are only found in microorganisms in membrane structure

Membrane structure → polymyxins

What structures/enzymes are only found in microorganisms in nucleic acid synthesis

Nucleic acid synthesis: rifampicin, novobiocin, and quinolones

What structures/enzymes are only found in microorganisms in other metabolic pathways

Other metabolic pathways → trimethoprim and sulfanilamide

What is sterilization

The complete removal of ALL microbes (including endospores and cysts)
For example, virus, bacteria, protozoa

What is aseptic

Produring sterile conditions in a lab or healthcare setting

What is disinfection/disinfectants

The removal of most but NOT all microbes - usually involve chemicals

What is antisepsis/antiseptic

Removal of microbes from human surfaces (disinfection)

What is degerming

Physical removal of microbes via scrubbing action (physical scrubbing action)

What is sanitization

Disinfection of public surfaces

What is pasteurization

Disinfection by use of gentle heating

What does the suffix -stasis/-static mean

Control growth but does not always kill microbes. it inhibits binary fission

What does the suffix -cide/-cidal mean

agents that KILL (directly kill)

What is microbial death rate

Microbial agents kill a constant percentage of cells over time, rather than simultaneously killing all cells

What is the ideal antimicrobial agent

• Inexpensive

• Fast acting

• Stable during storage

• Broad spectrum

• Harmless to humans, animals, and objects

(Bleach is a good antimicrobial agent that fits into most of the categories)

*Fewer organisms = faster sterility*

What does the expensiveness of antimicrobials depend upon

• Nature of site to be treated

• Susceptibility and number of microbes involved

• Environmental conditions during application

*Fewer organisms = faster sterility*

Where can harsh chemicals and extreme heat not be used on

Humans, animals, and fragile objects (methods and level of control must be based on-site of intended use or medical procedure)

What is an appropriate method for cleaning human skin

EtOH → alcohol

What are high-level germicides

Kill ALL pathogens, including endospores

What are intermediate-level germicides

Kill fungal spores, protozoan cysts, viruses, and pathogenic bacteria

What are low-level germicides

Kill vegetative bacteria, fungi, protozoa, and some viruses

What is the list of susceptibility of microorganisms

Most resistant → prions, bacterial endospores, mycobacteria, cysts of protozoa, active-stage protozoa (trophozoites), gram-negative bacteria, fungi, nonenveloped viruses, gram-positive bacteria, enveloped viruses ← most susceptible