3.6: Control of Microbial Growth

Question 1

what is the most widely used method of controlling microbial growth? why?

Answer 1

• heat
• high temps denature proteins and other cell structures

Question 2

sterilization

Answer 2

the killing or removal of all viable organisms from an object or a growth medium (if heat or chemical treatment kills most but not all, it’s not considered sterilization)

Question 3

inhibition

Answer 3

• Effectively microbial growth, without killing
• ex: high salt concs inhibit growth of E. coli (stop it from growing), but if it returns to lower conc. it could continue to grow.

Question 4

decontamination

Answer 4

• treatment that makes an object safe to handle
• ex: lab equipment that needs to be destroyed must be decontaminated before its sent off for others to handle.

Question 5

disinfection

Answer 5

A treatment that kills most microbes, but not necessarily all.

Question 6

Decimal reduction time

Answer 6

• amount of time required to reduce viability tenfold (by 1 tenth like 100 to 10)
• ie. time required to kill 90% of microbes present

Question 7

pasteurization

Answer 7

the process of using precisely controlled heat to reduce the microbial load in heat-sensitive liquids
* does not kill all organisms, so it is NOT sterilization

Question 8

how does pasteurization help milk?

Answer 8

• reduced microbial load gives milk longer shelf life

Question 9

what are the two different time and temp combinations that have been used to pasteurize milk? what pathogen do both processes do?

Answer 9

• LTLT (low temp/long time)
  -63 C for 30 mins
• HTST (high-temp/short time)
  -72 C for 15 seconds
• both processes kill the most resistant pathogen found in milk.

Question 10

what’s an autoclave, what temp and pressure and time does it use?

Answer 10

• a sealed device that uses steam under pressure to sterilize materials
• typically set to 121 C and 15 pounds per square inch of pressure for 15 to 20 min

Question 11

what internal temp must all materials in a autoclave reach? why this temp?

Answer 11

121 C, this temp is sufficient to destroy the endospores of Geobacillus stearothermophilus (previously most resistant endospores known)

Question 12

what does a typical cycle in an autoclave require?

Answer 12

• requires that material be kept at 121 C for 15 mins to be sterilized

Question 13

what does time it takes for an autoclave to reach 121 C depend on?

Answer 13

depends on the shape and size of the vessel
-larger materials take much longer to sterilize

Question 14

how does radiation control microbial growth?

Answer 14

• short wave length radition such as Gamma, X-ray, and UV can damage DNA and destroy microorganisms

Question 15

how can Gamma and X-ray radiation control microbial growth?

Answer 15

• Gamma and X-rays penetrate directly through materials
  -can sterilze lab equipment inside plastic packaging
  -used to prolong the shelf-life of food (meats, fruits, vegetables, spices, etc)

Question 16

how can UV radiation control microbial growth?

Answer 16

• UV does not penetrate well (even through glass or plastic)
  -Useful for decontaminating lab surfaces (only makes safe to handle)
  -Used along with chlorine for drinking water treatment

Question 17

what are membrane filters, how do they work?

Answer 17

• fine filter with 0.45 um or 0.22 um pore size
• Allow liquid to pass through, but restrict passage of bacteria bigger than pore size
• used to remove microbes from temp sensitive liquids
• ex. some media ingredients, antibiotics, enzymes, vaccines.

Question 18

what’s something to consider when choosing a membrane filter?

Answer 18

• size of microbes that could be present, the pore size must be smaller than the size of the microbe, so it prevents most bacteria from going through.

Question 19

what are the 4 chemical antimicrobial agents? order from most to least powerful?

Answer 19

1. Sterilant (most powerful)
2. Disinfectant
3. Sanitizer
4. Antiseptic (least powerful)

Question 20

what does sterilant do and example

Answer 20

• destroys all microorganisms including endospores
• Ex. Ethylene oxide -gas used to sterilize some temp sensitive lab equipment (paper on swabs allows ethylene oxide through)

Question 21

what does disinfectant do and example

Answer 21

• kills most microorganisms but not all endospores
• ex. Chlorine bleach -used to disinfect lab surfaces

Question 22

what does Sanitizer do and example

Answer 22

• less harsh than disinfectants, just used to reduce microbe **numbers **
• ex. 70% Ethanol hand-santizers

Question 23

what does an Antiseptic do and example

Answer 23

• kills or inhibits the growth of microorganisms, but sufficiently non-toxic to be applied on living tissue (won’t kill living tissue)
• ex. iodine used to treat small cuts

Question 24

what are the 3 classes of antimicrobial agents, describe them

Answer 24

1. Bacteriostatic: Prevents cell growth as long as the antimicrobial agent is present (think static like not moving =present)
2. Bacteriocidal: kills the cells, but does not lyse them (does not change their shape) (think cidal like homocidal =kills)
3. Bacteriolytic: kills the cells and lyses them (think lytic like lyse)

Question 25

what is the only bacteriolytic agent

Answer 25

lysozyme, cleaves beta bonds

Question 26

what's minimum inhibitory concentration (MIC), what does it vary with? ex?

Answer 26

* the smallest amount of an agent needed to inhibit growth of a microorganism * varies with the organism used, inoculum size, temperature, pH, etc * ex. Test tubes containing media and increasing concentration of an antibiotic from left to right, tube with MIC has the lowest conc of the antibiotic it can have and still be completely clear of growth

Question 27

what is Minimum lethal concentration (MLC)? how does it work?

Answer 27

* The smallest amount of an agent needed to actually kill the microorganism * starts with the MIC test * plate aliquots of broth from the MIC (that had no growth) and from tubes with higher conc. of antibiotic, on nutrient agar plates (without antibiotic) * If colonies form: conc. was bacteriostatic, not bacteriocidal * lowest conc. that results in no colonies on a plate = MLC

Question 28

how does a disc diffusion assay work?

Answer 28

* a filter paper disk is soaked in an antimicrobial agent * placed on a petri plate that has already been inoculated with a thin layer of bacteria * the chemical diffuses out of the disk as the plate is incubated * creates a zone of clearing, in lawn of bacteria (clearing shows that chemical is antimicrobial, size of clearing indicates how well) * corresponds to MIC (through a standard curve)

Question 29

what could a small zone of clearing also indicate in a disc diffusion assay if not that it's just a bad antimicrobial agent

Answer 29

could mean chemical just doesn't diffuse well through plate