Micro 107 - Quiz 3

Basic ph

Alkaline solutions (pH 7.01 -14) have excess of OH-ions or a deficiency of protons.

Acidic ph

An acidic solution (pH 0-6.99) has an excess of protons or a deficiency of OH-

Mechanisms of Action

An antibiotic can affect the growth of bacteria in a variety of ways including: -Inhibition of cell wall synthesis -Inhibit protein synthesis -Inhibit nucleic acid synthesis -Competitive inhibition of enzymatic activity

Antibiotics

An antibiotic is a chemical (quite often produced by a microbe) that exhibits the property of selective toxicity - i.e. a chemical that is poisonous to a microbe yet harmless to the human or animal host. Antibiotics include chemicals that are toxic to the entire range of infectious agents including viruses, bacteria, protozoa and fungi.

Hypotonic

An environment in which there is a much lower concentration of solutes outside the cell than inside - this is known as a HYPOTONIC environment. In such an environment, water will flow from the outside to the interior of the cell, effectively making the cell expand like an inflated balloon. Up to a point, many microbes can survive under such conditions due to the presence of the rigid cell wall that surrounds and acts as a protective cage that encases the plasma membrane.

Hypertonic

An environment where there is a much higher concentration of solutes outside the cell than inside - this is known as a HYPERTONIC environment. In such an environment water will flow from the inside to the outside of the cell, effectively draining water out of the cell. Such a situation will lead to the plasma membrane shrinking away from the rigid cell wall - this is known as PLASMOLYSIS. This loss of water from within the cell results in the cytoplasm becoming more "syrupy" and will ultimately lead to a halt in all metabolic activity. This is the basis of food preservation through salting and sugaring of food products. It is worthwhile noting that certain microbial species are able to grow in salty environments that would kill most other microbes. The HALOTOLERANT species (e.g. the Staphylococci) can grow at low to moderate salt concentrations, while the HALOPHILES (e.g. the Halococci) actually REQUIRE a high salt environment for growth

Isotonic

An environment where there is an equal concentration of solutes inside and outside the cell - this is known as an ISOTONIC environment and there is NO net flow of water across the membrane. It is very unlikely that microbes experience this situation in nature.

Bactericidal

Bactericidal drugs, on the other hand, outright kill the organism and are usually more effective on actively growing cells - thus, organisms that replicate very slowly (e.g. Mycobacterium tuberculosis, the causative agent of tuberculosis) are difficult to treat with bactericidal drugs

Bacteriostatic

Bacteriostatic drugs will slow the growth of organisms but do not kill the organism. Thus, such agents depend on a functional and robust immune system in the host - the antibiotic inhibits growth of the pathogen to an extent that the immune system is able to cope with the bacterial load within the system.

Based on your results, do you consider that bacteria are more acid tolerant than fungi? Explain the reason for the choice you have made.

Based on my results, I consider that bacteria are not more acid tolerant than fungi because only one species had significant growth in the acidic ph 3.

Look up the name of the causative agent of gas gangrene and record it below.

Clostridium perfringens is the main causative agent of gas gangrene

Look up some of the complications of diabetes especially with regard to blood flow to the toes and feet

Diabetes can lower the amount of blood flow in your feet. Not having enough blood flowing to your legs and feet can make it hard for a sore or an infection to heal. Sometimes, a bad infection never heals. The infection might lead to gangrene.

Disinfectants

Disinfection is used to describe processes that lead to the reduction in the number of pathogens (an organism capable of causing disease) to harmless levels. It is important to note that disinfection does not imply that the object treated with a disinfectant has been sterilized, but merely no longer capable of causing disease. Such chemicals are often too harsh to use on tissues and are reserved for cleaning surfaces. They do not destroy bacterial endospores.

Microbes live in environments where cells experience a difference between the concentration of solutes (ions, nutrients, etc.) in the cytosol and that is found in their habitats.

For this reason, cells are subjected to the effect of osmosis, i.e. an attempt to balance the "concentration gradient" of solutes across the plasma membrane.

It is a relatively easy task to grow aerobic organisms in the laboratory - the atmosphere has sufficient oxygen to support the growth of such microbes.

However, the culture of anaerobes presents the challenge of having to establish an oxygen-free environment. (that's why we use the incorporation of the reducing agent, thioglycollate, into the growth media.)

Which of the species used in this exercise would you expect to be most likely to grow most rapidly in the human body?

I would expect M. luteus to be most likely to grow most rapidly in the human body because it has the highest growth average for 37 degrees Celsius, which is the internal temperature of the human body.

Which of the species used in this exercise would you expect to be most likely to cause spoilage of refrigerated food?

I would expect the Arthrobacter species to be most likely to cause the spoilage of refrigerator food because it grows best at colder temperatures.

UV light occupies a region of the electromagnetic spectrum that resides between 100 - 400 nm (visible light resides at wavelengths between ~ 400-700 nm).

In addition to being more energetic than visible light, UV is particularly damaging to cells because DNA absorbs light of this wavelength. DNA exposed to UV light (especially at the shorter wavelengths), is especially susceptible to a type of mutation known as a THYMINE DIMER.

Biologists can take advantage of the effect of UV light on cells as a means to reduce the microbial population in tissue culture hoods and lab benches

It is important to note that UV light is NOT very penetrating and as such its effects are limited to surfaces that are directly exposed to the radiation. For instance, the visible portion of sunlight readily penetrates glass or plastic while UV light is blocked.

Kirby-Bauer Disk Diffusion Assay

Microbial sensitivity testing to antibiotics

What is the difference between the optimal and maximal growth temperatures?

Optimal growth temperatures are the temperatures at which an organism is able to best grow, at which the organism grows the fastest and spreads the easiest. Maximal growth temperatures refer to the highest temperatures at which an organism can survive. It can grow, but that does not mean it can grow very successfully at its maximal growth temperature.

What species was most resistant to the effect of high concentrations of salt?

Penicillium was most resistant to the effect of high concentrations of salt due to still having a relatively high growth compared to the other species.

What distinguishes a psychrophile from a thermophile?

Psychrophiles are cold-loving microbes, while thermophiles are heat-loving microbes. A psychrophile grows best at colder temperatures between 0 and 15 degrees Celsius, while a thermophile grows best at hotter temperatures between 40 and 80 degrees Celsius.

Thioglycollate media

Sodium thioglycollate is a reducing agent that very effectively removes oxygen from the environment. It accomplishes this task by reacting with and sequestering the oxygen in the media. Within a thioglycollate broth tube, an oxygen gradient is established: the portion at the interface between the air and the medium is somewhat aerobic while the deeper regions that are at a distance from the atmosphere become increasingly anaerobic. In fact, we can track the amount of oxygen along the depth of the culture tube because the medium also contains the oxygen indicator RESAZARIN - it is colorless in anaerobic conditions and will turn pink as it becomes exposed to oxygen.

Sterilant

Sterilization is a term used to describe the destruction of all organisms in or on something. This includes all organisms on a surface or in a fluid

What concentration of salt was most effective at inhibiting the growth of microbes?

The 15% concentration of salt was most effective at inhibiting the growth of microbes as evident from the results showing low growth at 15%.

The effectiveness of an antibiotic can be described in terms of its SPECTRUM as well as the SENSITIVITY/RESISTANCE of the bacterial pathogen to a particular antibiotic.

The Antibiotic Spectrum describes the range of species that are sensitive to the antibiotic. Thus, there are "broad" and "narrow" range antibiotics.

Faculative microbes

The Facultative microbes are a very adaptable class of microbes in that they can grow either in the presence or absence of oxygen. In the presence of oxygen, they grow by aerobic respiration; in the absence of oxygen, they are quite content to grow by anaerobic respiration or fermentation.

Based of your results from this exercise, explain why the causative agent of gas gangrene is such a danger to diabetics

The causative agent of gas gangrene, clostridium perfringens, is such a danger to diabetes because it can grow in the feet and will not heal effectively due to the lower amount of blood flow caused by the diabetes.

The compartment with the higher concentration of water is also the side that has the lower concentration of solutes.

The compartment with the lower concentration of water will also be the compartment that has the higher concentration of solutes.

Osmotic pressure

The force that drives this movement of water molecules across the membrane in osmosis

ph

The pH of a solution is a quantitative indicator of its acidity or alkalinity.

ph scale

The pH scale is a logarithmic or an exponential scale and ranges from 0 -14: a change of 1 pH unit is equivalent to a 10-fold increase or decrease in the number of protons.

Which of the species are likely to grow in a jar of canned tomatoes (canned tomatoes are quite acidic)?

The species that is likely to grow in a jar of canned tomatoes is S. cerevisiae, considering that canned tomatoes are quite acidic and S. cerevisiae grew the best in ph 3 compared to the other species.

Plasmolysis

This happens when a cell shrinks inside its cell wall while the cell wall remains intact due to lack of water (occurs in hypertonic environments)

Microbes occupy habitats where they are exposed to variations in the pH of the environment.

Thus, different microbial species have evolved to survive within a defined pH range.

The Sensitivity/Resistance is a feature of the bacterium - i.e. whether or not a particular bacterium can be inhibited or killed by the drug.

Thus, effective antibiotic treatment requires the bacterium to be sensitive to and also within the range of an antibiotic.

Thymine dimers are formed when UV leads to a covalent linkage between adjacent thymine residues on the same DNA strand. This leads to a distortion in the shape of the double helix and ultimately results in errors during replication. These errors are MUTATIONS.

When many of these errors accumulate in a genome, mutated proteins accumulate and ultimately lead to the death of the organism. It is worthwhile noting that the same type of mutation occurs in skin cells that are overexposed to sunlight, ultimately increasing the risk of skin cancer.

Antiseptics

a chemical that is used externally on tissues to destroy OR inhibit the growth of microorganisms. An antiseptic like Hydrogen Peroxide will effectively clean a cut but it will also cause moderate tissue damage.

Sensitivity vs. Resistance

a feature of antibiotics

Ultimately, a bigger difference in the concentration on the opposite sides of a membrane results in

a larger osmotic pressure.

Phenol coefficient

a quantitative assessment of the effectiveness of a compound as a disinfectant

Resazarin

a redox-sensitive dye (an oxygen indicator) -Colorless in anaerobic conditions -Pink in aerobic conditions

Osmosis

a special example of diffusion; water will flow from the compartment with a higher concentration to the side with the lower concentration On the other hand, the movement of water from an area of low solute concentration to an area of high solute concentration in order to achieve equilibrium.

Which chemical was least effective against each of the bacteria used?

a. E. coli: 0.5% Lysol b. B. megatherium: 0.5% Lysol

Which chemical was most effective against each of the bacteria used?

a. E. coli: 70% Ethanol b. B. megatherium: 5% Lysol, Mouthwash, 70% Ethanol and 10% Chlorox

Strict anaerobes

an organism that cannot survive in an atmosphere of oxygen; e.g. the Clostridium species. For some of these organisms, exposure to oxygen is fatal. They depend on anaerobic respiration and/or fermentation for growth.

Strict aerobes

can only grow in oxygen; e.g. the Micrococcus species, have an absolute requirement for oxygen and depend on aerobic respiration for growth.

Phenol coefficient

compares a chemical's antimicrobic properties to those of phenol; This is a quantitative assessment of the effectiveness of a compound as a disinfectant. The anti-microbial activity of the compound is compared with that of Phenol on two tester strains: the Gram-negative bacterium Salmonella typhi and the Gram-positive bacterium Staphylococcus aureus. These organisms are grown under standardized conditions and a set concentration of the chemical is tested against a set concentration of phenol.

The Temperature Growth Range

describes the values between the minimal and maximal growth temperatures that can support growth and metabolism. At the lowest end of the range is the MINIMUM growth temperature; at the upper end is the MAXIMUM growth temperature. Somewhere between the minimum and maximum is the OPTIMAL growth temperature. At lower end of the growth range, the metabolic activity of cells slow down to a point that is incompatible with survival. As the temperature increases from the minimum, metabolic activity initially increases to a point where cells are most comfortable as evidenced by their growth rate (how often they divide per unit time) - this is the OPTMIMAL growth temperature. However, beyond this point, metabolic activity begins a steady decline due to the effect that higher temperatures have on the cells' macromolecules (e.g. proteins, the lipid membrane, DNA, etc.), i.e. the DENATURATION of the macromolecules.

Antiseptics

destroy OR inhibit the growth of microorganisms , use on tissue

Kirby-Bauer Disk Diffusion Assay

determine the sensitivity of a microbe to a set of antibiotics. The assay involves the inoculation of a Mueller-Hinton (M-H) agar plate with a lawn of bacteria. M-H agar is a rich medium that supports the growth of many human pathogens. A set of antibiotic impregnated disks is then placed on the bacterial lawn, and the antibiotic diffuses in a circle away form the disk into the surrounding agar. If a bacterial strain is sensitive to a particular antibiotic, a circular zone of clearing and inhibition is observed surrounding the disk. However, if the strain is resistant, the bacterial lawn may grow up to the edge of the disk.

Oxidizing agents

e.g. Hydrogen Peroxide - Oxidation of biological macromolecules.

Halogens

e.g. Iodine and Chlorine. Denature proteins.

Phenolics

e.g. Lysol, pHisoHex hand wash. Dissolve the plasma membrane and denature proteins.

Alcohols

e.g. isopropanol (rubbing alcohol) and ethanol. Work primarily by dissolving the plasma membrane can denature proteins.

Acidophiles

grow in acidic environments, low phs

Alkaliphiles

grow in alkaline environments, high phs

Erythromycin

inhibits bacterial protein synthesis

Tetracycline

inhibits bacterial protein synthesis

Ampicillin

inhibits peptidoglycan synthesis

Cephalothin

inhibits peptidoglycan synthesis

Penicillin

inhibits peptidoglycan synthesis

Vancomyocin

inhibits peptidoglycan synthesis

Neutraphiles

live at near neutral environments

Halotolerant

organisms that can tolerate some reduction in water activity of environment but generally grow best in the absence of the added solute; can grow at low to moderate salt concentraions

-cidal agents

permanent destruction of a cell

Sodium thioglycollate

reducing agent (removes oxygen)

Disinfectant

reduction of pathogen load, use on utensils

Sanitization

removing microorganisms to reduce contamination to safe levels,generally used in food industry & public health

Growth range

represents the range of conditions under which a bacterium can grow; minimum, maximum, and optimum growth temperatures

-static agents

slows or stops growth but if the agent is removed growth may resume

Cytolysis

the bursting of a cell from internal water pressure

Sterilant

the destruction of all organisms in or on something

Osmotic pressure

the force that drives this movement of water molecules across the semi-permeable membrane

Diffusion

the movement of particles (atoms, ions or molecules) from a region in which they are in higher concentration to regions of lower concentration.

Optimal growth rate

the point at which cells are most comfortable as evidenced by their growth rate

Sanitization

the process of removing microorganisms to reduce contamination to safe levels. This term is often used by public health officials, the food industry and in medicine to describe something that is "clean".

Antibiotic spectrum

the range of species that are sensitive to the antibiotic (narrow to broad) ;a feature of antibiotic

The direction of water flow is dependent on the difference in the concentration of water molecules between both sides of the membrane -

water will flow from the compartment with a higher concentration to the side with the lower concentration.

Soaps and detergents

work by dissolving the plasma membrane

Antibiotics can have one of two effects on microorganisms: a "-static" or a "-cidal" effect.

!!

The ultimate determination of resistance or sensitivity of a bacterial strain to an antibiotic depends on the size of the zone of inhibition.

!!

Halophiles

"salt-loving" archaea that live in environments that have very high salt concentrations

Osmosis

-Diffusion of water through a selectively permeable membrane -the process in which water flows from one side to the other of a semi-permeable membrane.

All living organisms, including microbes, can sense and respond to changes in environmental temperature.

-Each species has a temperature growth range between which we can identify a MINIMUM, OPTIMUM, and MAXIMUM growth temperatures. -Based on these values, bacteria can be classified as PSYCHROPHILES, MESOPHILES, or THERMOPHILES.

Pure water has a pH of 7.

-If an acid (e.g. hydrochloric acid, HCl) is added to such a water solution, the acid donates protons to the water and reduces its pH. -If an alkali (also known as a base; e.g. sodium hydroxide, NaOH) is added to water, it donates hydroxyl ions to the water and increases its pH.

Hyperthermophiles

-Organisms that grow in extremely high temperatures (90 degrees C) -can grow above 100 degrees C

Antibiotics

-Selective toxicity to microbes & no effect on hosts -Many produced by microbes to eliminates competitors -Synthetic antibiotics based on structure of natural one -can be consumed unlike disinfectants & antiseptics -Bactericidal vs. Bacteriostatic

Cidal agents vs static agents

-cidal agents kill the microbes, while -static agents can stop the growth (however the microbes can return if the agent isnt being in use)

Psychrophiles

-cold-loving microbes -grow at low temps: below 0 - 15 degrees C

Thermophiles

-heat loving microbes -grow between...40 - 80 degrees C

Microbes can experience three environments with regard to osmotic pressure.

-isotonic -hypertonic -hypotonic

Mesophiles

-moderate temperature loving microbes -grow between ~15 - 45 degrees C -human's internal temperature: 37 degrees C, therefore mesophiles are the species that grow best in humans

Mechanism of Action

1) Inhibition of cell wall synthesis e.g. penicillin, ampicillin 2) Inhibit protein synthesis e.g. tetracycline 3) Inhibit nucleic acid synthesis 4) Competitive inhibition of enzymatic activity

There are a number of variables that influence the effectiveness of UV light as a germicide. These include:

1. The wavelength used - the shorter wavelengths at about 260 nm are far more effective than longer wavelengths 2. The duration of exposure - longer is better 3. The type of organism - endospores are far more resistant than vegetative cells 4. Activation of the cellular DNA repair machinery - these pathways exist to help repair mutations when they arise. However, the DNA repair systems may be overwhelmed if too many mutations accumulate in the cell.

Neutral ph

A "neutral" solution (pH 7.0) has an equal amount of protons (H+) and hydroxyl (OH-) ions.

Light is measured as a wavelength in nanometers.

A certain range of the electromagnetic spectrum (visible light) can be detected by the human eye; however, the wavelengths that exist on either side of the visible spectrum cannot be detected by the human eye

This principle of selective toxicity exploits the differences in cellular structure and metabolic pathways that exist between the animal host and the infectious agent. The more effective agents target structures or metabolic pathways that are unique to the bacterium, e.g. the effect of penicillin on the peptidoglycan layer of the bacterial cell wall.

A key difference between antiseptics/disinfectants and antibiotics is related to this issue of selective toxicity: while all agents are toxic to bacterial and human cells, only antibiotics preferentially target bacteria. For this reason, only antibiotics can be taken orally or injected into the bloodstream to combat infections.