Laboratory Culture of Microorganisms

Question 1

What are the essential elements for all microorganisms?

Answer 1

C, O, N, H, P, S, Se

Question 2

What is the culture media?

Answer 2

• nutrient preparations devised to support the growth (reproduction) of microorganisms
• contains macronutrients and micronutrients (trace metals, growth factors)
• can be liquid or solid (agar)

Question 3

What are the bases of classification for culture media?

Answer 3

• chemical composition: defined and complex
• physical nature: liquid, semisolid, solid
• function: supportive, enriched, selective, differential

Question 4

What is defined media?

Answer 4

• exact composition is qualitative and known
• very specific
• usually used for one particular organism

Question 5

What is complex media?

Answer 5

• contain ingredients of unknown composition and/or concentration
• one complex ingredient automatically makes the media complex
• complex ingredient examples: peptones, extracts, agar

Question 6

What is general purpose media?

Answer 6

• supports the growth of many microorganisms
• typically a complex media
• ex: tryptic soy agar (TSA)

Question 7

What is enriched media?

Answer 7

• general purpose media supplemented with highly nutritious substances such as blood
• also typically complex media
• ex: chocolate agar (boiled blood agar)

Question 8

What is minimal media?

Answer 8

• contains the minimal necessities for growth of the wild type
• only contains inorganic salts, a simple carbon source, and water

Question 9

What is selective media?

Answer 9

• favors the growth of some microorganisms and inhibits the growth of others
• ex: EMB agar selects for gram-negative bacteria

Question 10

What is differential media?

Answer 10

• allows you to distinguish between groups of microorganisms based on their biological characteristics
• ex: blood agar (hemolytic v. non hemolytic bacteria)
• ex: MacConkey agar (lactose fermenters v. non-fermenters)

Question 11

What are alpha hemolytic bacteria?

Answer 11

• bacteria that partially break down blood

- leave a yellow/green ring around them in blood agar

Question 12

What are beta hemolytic bacteria?

Answer 12

• bacteria that fully break down blood

- leave a clear ring around them in blood agar

Question 13

What are gamma hemolytic bacteria?

Answer 13

• bacteria that do not break down blood

- no ring around them in blood agar

Question 14

What is a pure culture?

Answer 14

• a population of cells arising from a single cell

- allows for the study of a single type of microorganism

Question 15

How is a pure culture obtained?

Answer 15

• a mixture of cells is applied to an agar surface so that individual cells are well separated from each other
• streak plate, pour plate, and spread plate techniques

Question 16

How does streak plating work?

Answer 16

1. Loop is sterilized and a loopful of inoculum is removed from the tube
2. Initial streak is worked in well in one corner of the agar plate
3. Subsequent streaks are at angles to the first streak
4. After incubation, you should obtain isolated colonies at the end of your streak

Question 17

How does spread plating work?

Answer 17

• small volume of diluted culture is transferred to an agar surface
• culture is spread evenly over surface with a sterile bent glass rod (hockey stick)

Question 18

How does pour plating work?

Answer 18

• diluted samples are mixed with liquid agar
• mixture of cells and agar are poured into sterile culture dishes
• some colonies may be embedded into the agar (still counted in your plate count)

Question 19

What is a countable plate for either pour or spread plating?

Answer 19

30-300 colonies

Question 20

How is cell concentration determined from a plate count?

Answer 20

• find the dilution plate that is countable (30-300 colonies)
• divide the number of colonies by the total dilution
• if you added 1 mL to your plate, then you are done, but if you added 0.1 mL, then you need to multiply by 10

Question 21

How do microbes grow on solid surfaces?

Answer 21

• species form characteristic colonies (color, texture, arrangement)
• differences in growth rate from edges to center us due to oxygen, nutrients, toxic products, and cell death
• oxygen is available for cells on top
• nutrients are available for cells on bottom
• toxic products are produced from fermentation by cells who don’t have oxygen access

Question 22

What are the different ways to characterize bacterial colonies?

Answer 22

• form
• elevation
• margin
• color
• consistency