MB 351 - Lecture 16

Question 1

Two classes of culture media

Answer 1

Chemically defined, and complex

Question 2

Chemically defined media

Answer 2

The exact composition of pure chemicals used to formulate the medium is known. -used extensively for well characterized isolates in lab studies (chemoheterotrophs like E.coli) -media must contain organic growth factors, such as glucose, that serves as a source of carbon and energy

Question 3

Complex media “rich” media

Answer 3

Composed of a mixture of proteins and extracts in which the exact chemical composition is poorly defined, often made of natural source components- red blood cells, yeast extracts, peptone, Casein, etc. Used as a non-specific medium for growing many organisms

Question 4

When are complex media used?

Answer 4

Complex media usually provide the full range of growth factors that may be required by an organism so they may be more handily used to cultivate unknown bacteria or bacteria whose nutritional requirements are complex (organisms that require a lot of growth factors, known or unknown)

Question 5

A nutrient material prepared for the growth of microbes in a lab

Answer 5

culture medium

Question 6

What’s in nutrient broth/agar?

Answer 6

Broth (liquid), agar (solid) -Glucose, peptone (animal, milk, or meat), beef extract, yeast extract, sodium chloride, agar -pH 7.0 to 0.2 -storage 2-8 degC Beef extract added to compliment peptone to add minerals, phosphates, energy sources, things missing from peptone. Yeast provides vitamins an other organic growth factors – rich in vitamin B, also supply organic N and C.

Question 7

Complex media are easier to prepare than ________

Answer 7

chemically defined media

Question 8

Three types of culture media used to detect the presence of specific microbes

Answer 8

selective, enriched, and differential media

Question 9

Selective media

Answer 9

Inhibits growth of unwanted organisms, supports growth of desired organisms, and thereby ‘selects’ for it. -ex. Brilliant green agar isolates gram negative, and prevents growth of gram positive

Question 10

Enrichment Media

Answer 10

-Usually a complex medium base to which additional nutrients such as serum or whole blood are added. -Contains some important growth factors (vitamin, amino acid, carbon source) needed for the growth of fastidious organisms. -These added nutrients better mimic conditions in the host and are required for successful lab culture of some human pathogens like Streptococcus pyogenes. -ex. blood agar, chocolate agar -the medium for an enrichment culture provides nutrients and environmental conditions that favor the growth of a particular microbe, but not others. In that sense it is also a selective media.

Question 11

Differential media

Answer 11

-Allows for the separation of organisms by some observable change in appearance of the media. -ex. blood agar -Media that supports the growth of many different organisms, but differentiates between them. -Media will allow identification of microbes based on their growth, color, and appearance on the medium

Question 12

Sometimes selective and differential media are combined in a single medium ex?

Answer 12

MacConkey agar -bile salts and crystal violet inhibit Gram positive bacteria (selective) and allow growth of gram negative -lactose fermenters produce color change (differential)

Question 13

Mannitol Salt Agar

Answer 13

????

Question 14

puncturing suitable medium such as nutrient agar with an inoculating needle that goes deep into the agar, allows growth within the semi-solid media

Answer 14

Stab (deep) culture - long term storage

Question 15

Inoculating devices

Answer 15

needle or loop

Question 16

Also suitable for longer term maintenance of bacteria. Agar media is poured into test tubes for and allowed to solidify in an angled position, so that media has a ______ to allow for greater surface area and inoculation on this slanted surface.

Answer 16

Slant cultures

Question 17

Microbes grow on the surface of an agar _____ . Helps to show a colony, which is a visible mass of microbial cells arising from one cell or from a group of the same microbes.

Answer 17

Plate culture

Question 18

Use broths for specific assays or for bacteria that will not form colonies on a solid surface. There are two types of broths:

Answer 18

batch cultures and continuous chemostat cultures

Question 19

Batch cultures

Answer 19

microbes are cultured in an enclosed vessel (tube, flask) with a broth medium that supplies its nutritional requirements

Question 20

Continuous Chemostat culture

Answer 20

This culture overcomes the limitations of a batch culture. In a chemostat, the growth chamber is connected to a reservoir of sterile medium. Once growth is initiated, fresh medium is continuously supplied from the reservoir.

Question 21

Chemostat culture helps to prevent this phase…

Answer 21

The stationary phase is often due to a growth-limiting factor such as the depletion of an essential nutrient, and/or the formation of an inhibitory product such as an organic acid. Stationary phase results from a situation in which growth rate and death rate are equal.

Question 22

Advantage of using chemostat culture is that cell population can be maintained in the ________ phase for days or even weeks.

Answer 22

exponential growth phase

Question 23

Shaker is used to? For what types of microbes?

Answer 23

Used to supply oxygen to a aerobe

Question 24

Steps for growing bugs aerobically

Answer 24

1. Make culture media -carbon/energy source -nitrogen source -micronutrients/growth factors -buffer -water (add agar for solid medium) 2. Sterilize medium 3. Inoculate liquid or streak plate 4. Wait until culture grows

Question 25

Anaerobic Culturing: two ways

Answer 25

GasPak jars, and anaerobic chambers (also called a glove box)

Question 26

GasPak jars - small cultivation

Answer 26

inoculated plates or tubes are placed inside the jar, and anaerobic conditions are created by adding water to a gas generator envelope placed in the jar just before sealing.

Question 27

Anaerobic chambers - large cultivation

Answer 27

plastic chamber that contains an atmosphere of H2, CO2, and N2. Culture media are placed within the chamber by means of an airlock which can be evacuated and refilled with N2.

Question 28

inoculated plates or tubes are placed inside the jar, and anaerobic conditions are created by adding water to a gas generator envelope placed in the jar just before sealing.

Answer 28

GasPak jars - small cultivation

Question 29

plastic chamber that contains an atmosphere of H2, CO2, and N2. Culture media are placed within the chamber by means of an airlock which can be evacuated and refilled with N2.

Answer 29

Anaerobic chambers - large cultivation

Question 30

Reducing Media

Answer 30

for anaerobic cultures: during preparating of the culture media, it is boiled for severel minutes to removed dissolved oxygen, and a reducing agent (cysteine) is added to further lower the oxygen content. Oxygen free N2 is bubbled through to keep it anaerobic.