Topic 4-L4 - Control of microbial growth

Question 1

Decontamination:

Answer 1

Neutralization or removal of microbes (general term)

Question 2

Disinfection:

Answer 2

Eliminating or removing harmful organisms

Question 3

Sterilization:

Answer 3

killing all microorganisms. If something is sterile, it is completely free of microbes. Sterility can be thought of as binary –it’s either sterile or it isn’t.

Question 4

Decimal reduction time:

Answer 4

amount of time it takes to reduce number of microbes by a factor of 10 (a log)

Question 5

Thermal death time:

Answer 5

Amount of time if takes to kill all cells at a given temperature. Depends on number of cells.

Question 6

Both decimal reduction time and thermal death time depend on a number of factors:

Answer 6

pH, salt concentration, moisture levels, presence of fats/sugars/proteins in samples (can decrease heat penetration), etc.

Question 7

small changes in temp can have a huge impact on

Answer 7

of microbes remaining after a given heating time.

Question 8

Autoclaves are widely used to

Answer 8

sterilize by heat using steam.

• Endospores (highly resistant!) require
  temperatures of 121oC for 15 minutes

Question 9

How are autoclave 121C?

Answer 9

Water (steam) is therefore pressurized to increase boiling temp - allowing it to be heated to 121oC

• High pressure steam circulated throughout autoclave. Typical cycle length ~ 30 minutes to ensure materials reach 121oC for at least 15 minutes (varies - longer for large volumes of liquids, for example)

Question 10

Pasteurization is a common methods to prevent

Answer 10

spoilage and protect against food-

borne pathogens with certain foods/beverages

Question 11

How is pasteurization done?

Answer 11

Heated for a specific amount of time to eliminate pathogens and to reduce spoiling agents (non-pathogenic bacteria, enzymes)

Question 12

Pasteurization For milk,

Answer 12

71oC for 15 seconds

• In milk, heat-resistant lactic acid bacteria survive pasteurization and are what cause the milk to spoil over time.

Question 13

Pasteurization is NOT

Answer 13

Sterilization

Question 14

Ultraviolet (UV) light (220-300 nm wavelength)

Answer 14

damages DNA and is lethal to microbes at high enough intensities

• used for sterilizing bio safety hood (poor penetrating power)

Question 15

Ionizing radiation (such as
gamma rays) have

Answer 15

improved penetrating power and also can be used to sterilize

• used for sterilizing surgical supplies,
  labware, even food!

Question 16

Heat is a great way to sterilize, heat can also cause

Answer 16

damage (e.g., inactivate proteins or other molecules)

Question 17

Filter sterilization

Answer 17

Passing liquids through filters.

• very effective, but not as reliable as autoclaving

Question 18

Sterilants:

Answer 18

Kill all microbes – e.g. formaldehyde

Question 19

Disinfectants:

Answer 19

largely for surfaces, kill many/most microbes but not all (usually not endospores) – e.g. lysol

Question 20

Sanitizers:

Answer 20

Unlike the above, less harsh to humans, but also generally less effective – e.g. soaps

Question 21

Antiseptics:

Answer 21

Kill or inhibit growth of microbes, non-toxic enough to use on tissues (external – such as wounds). E.g. ethanol.

Question 22

Antimicrobial agent:

Answer 22

Chemical that kills or inhibits the growth of microbes

Question 23

“cidal” (bactericidal, fungicidal) –

Answer 23

Agents that kill cells. Irreversible.

Question 24

“static” (bacteriostatic) –

Answer 24

Agents that do not kill, but that inhibit growth. Prevent growth, but microbes are not dead and can recover.

Question 25

“lytic” (bacrteriolytic) –

Answer 25

Agents that not only kill cells, but that cause them to lyse (pop open – no dead cells can be observed)

Question 26

The “MIC” of a compound against a particular microbe is the

Answer 26

minimal inhibitory concentration

Question 27

minimal inhibitory concentration

Answer 27

Lowest concentration of a compound that fully inhibits microbial growth

Question 28

How is minimal inhibitory concentration determined?

Answer 28

using serial dilutions of the compound (usually 2x dilutions) – measuring microbial growth.

Question 29

Measuring toxicity of chemicals

Answer 29

• MIC

- solid media, zones of inhibition

Question 30

Solid media, zones of inhibition

Answer 30

• Antimicrobial activity can also be assessed using solid media combined
  with disks that contain the compound of interest
• Disk placed on agar plate – diffuses
• Closer to disk = higher concentration
  of antimicrobial agent
• The size of the “zone of inhibition”
  – area of no microbial growth –
  indicates susceptibility to compound
• “Kirby-Bauer test”

Question 31

antibiotic typically refers to medicines (chemicals) that have

Answer 31

antimicrobial properties that are used to treat infections.

Question 32

Antibiotics use

Answer 32

specific mechanisms to tangent microbes, rather then generic and non-specific toxicities

Question 33

Topical antibiotics are antibiotics manufactured into

Answer 33

creams/ointments that are applied to the skin. Others are ingested and dont hard human cells

Question 34

Antibiotics are Generally they are

Answer 34

small molecules that target a specific aspect of the biology of the microbe that is absent or very different in humans (e.g.
bacterial cell wall)

Question 35

Fleming discovered

Answer 35

penicilin (not used for a decade after discovery) and lysozyme

Question 36

First effective, ingested antibiotic used in clinics was

Answer 36

protonsil (discov. By Bayer by screening dyes)

Question 37

Protonsil not effective in test tube, only

Answer 37

live animals – pro-drug that must be processed by the body to active form

Question 38

How do antibiotics work

Answer 38

• specific targets, disrupt microbial processes
• target essential processes (cell wall biosynthesis, translation(protein synthesis), DNA replication, essential biosynthetic processes)
• need to get into cell, permeability is an issue

Question 39

How do antibiotics work: penicillin

Answer 39

• binds and inhibits activity of penicillin binding proteins – which are proteins that catalyze transpeptidation rxn that crosslink the cell wall
• In growing bacteria, cell wall is constantly being remodeled (autolysin proteins). Digested cell wall can’t be repaired – cell wall loses integrity, cells lyse due to osmotic pressure.

Question 40

Many antibiotics are

Answer 40

natural products (or chemically modified versions of natural products) isolated from microbes

Question 41

Microbes that produce a lot of antibiotics

Answer 41

Certain Actinomycetes such as Streptomyceshave been a very rich source of antibiotics

Question 42

Streptomyces are soil-dwelling bacteria that produce large numbers of
antibiotics –

Answer 42

large genomes, many interesting biosynthetic gene clusters

Question 43

Antibiotics are not just for bacteria – some target

Answer 43

eukaryotes, such as fungal pathogens

Question 44

polyene antibiotics from

Answer 44

Streptomyces

Question 45

Polyenes bind to a

Answer 45

cholesterol-like molecule, ergosterol (not in humans), found in fungal
cytoplasmic membranes – permeabilize membrane, cell death.

Question 46

For virtually all medically-important microbial pathogens,

Answer 46

strains resistant to commonly-used antibiotics can be found

Question 47

Antibiotic-resistant bacteria are

Answer 47

altered genetically (mutation or acquiring resistance genes) in a way that reduces sensitivity to that antibiotic.

Question 48

4 main mechanisms of resistance:

Answer 48

1) Modification of drug target (mutation of enzyme targeted by drug)
2) Enzymatic inactivation of the drug (degrade the drug)
3) Removal from cell by efflux pumps (pump drug out of cell)
4) Metabolic bypass (find another way to do what the drug is blocking)

Question 49

For many antibiotics,

Answer 49

derivatives of a parent class have been developed

• improved activity, more broad spectrum, less toxicity in humans

Question 50

derivatives are designed to

overcome known

Answer 50

resistance mechanisms

Question 51

For penicillin, some derivatives are not

sensitive to an enzyme called

Answer 51

β-lactamase

Question 52

β-lactamase is an

Answer 52

antibiotic-resistance gene – an enzyme that degrades penicillin

• β-lactamase-encoding strain can therefore
  be treated with these derivatives

Question 53

Examples of derivatives that are β-lactamase resistant

Answer 53

• methicillin

- oxacillin

Question 54

β-lactamase sensitive derivatives

Answer 54

• ampicillin

- carbenicillin

Question 55

Persistence is when antibiotic-sensitive bacteria includes

Answer 55

rare cells that are transientlytolerant to antibiotics

Question 56

Most antibiotics depend on metabolic activity (& often growth) in
order to be effective – e.g. if you’re not building a cell wall,

Answer 56

inhibiting cell wall synthesis is not effective

Question 57

Dormancy

Answer 57

(slowing or shutting off metabolism) is a common route to persistence

Question 58

Unlike resistant bacteria,

persisters are

Answer 58

genetically unchanged – can be killed by antibiotics once again after they
emerge from their tolerant state.