Module 3: Bacterial Reproduction and Growth Curves

Question 1

Define generation time

Answer 1

Time required for binary fission to take place and bacterial population to double

Question 2

What factors affect generation time?

Answer 2

Genetic control (DNA)
Available nutrients (some metabolized faster than others [glucose=fast, proteins=slow])
Environmental conditions (temperature, oxygen, carbon dioxide)

Question 3

Is generation time faster or slower in vivo?

Answer 3

Slower due to a number of factors that slow bacterial growth

Question 4

What are the 2 types of bacterial counts?

Answer 4

Viable count - live bacteria

Total count - live and dead bacteria

Question 5

How is the viable bacteria count determined?

Answer 5

By demonstrating their ability to reproduce

Colony count by adding bacteria to molten agar and letting harden, each colony = 1 viable bacteria

Question 6

How is a total bacteria count determined?

Answer 6

Under microscope - counting all bacteria in a ruled chamber

Visual turbidity - cloudiness of broth = number of bacteria (compared against McFarland standards)

Question 7

How are McFarland standards prepared?

Answer 7

Varying amounts of 1% sulfuric acid to varying amounts of 1.175% aqueous barium chloride creating a white precipitate

Question 8

What is the amount of bacteria compared to a 0.5 McFarland standard?

Answer 8

1.5 x 10^8 bacteria/mL

Question 9

What instruments can measure bacterial counts in solution?

Answer 9

Spectrophotometer and nephelometer

Question 10

How does spectrophotometry measure bacterial counts in solution?

Answer 10

Turbidity is measured at either %T (transmittance) or optical density
Calibration curves convert value to number of bacteria/mL

Question 11

How does nephelometry measure bacterial counts in solution?

Answer 11

Measures the amount of light scattered

Question 12

What are the 4 phases of the bacterial growth curve?

Answer 12

1. Lag phase
2. Logarithmic or Exponential phase
3. Stationary phase
4. Death phase

Question 13

What happens in the lag phase of the growth curve?

Answer 13

Little to no cell number increase, cells are metabolizing enzymes and getting larger

Question 14

How can the lag phase be shortened?

Answer 14

Take inoculum from bacteria already in the log phase (actively growing)
Use a medium most suitable for the bacteria to allow it to adjust more quickly

Question 15

What happens in the log/exponential phase of the growth curve?

Answer 15

All cells are dividing at a constant rate

Liquid medium becomes cloudy, colonies start appearing on plates

Question 16

Why is the log phase most desirable for working with bacteria?

Answer 16

Best cellular morphology
Most likely time to show motility
Most susceptible to antibiotics

Question 17

Why might the log phase of growth end?

Answer 17

Exhaustion of nutrients

Build up of toxins

Question 18

What happens in the stationary phase of the growth curve?

Answer 18

Number of viable cells remains constant, cells stop dividing, growth rate equals death rate

Question 19

Why might the stationary phase be of use in the lab?

Answer 19

Easier to find spores as they start growing at the end of the log phase

Question 20

What happens in the death/decline phase of the growth curve?

Answer 20

Viable cells decline and dye exponentially

Broth may clear due to autolysis

Question 21

Why is the death phase important in lab?

Answer 21

Gram stains are poor
Involution morphology is most likely
May not be motile

Best time to find spores

Question 22

Bacteria that are able to form colonies on solid medium after overnight incubation have a generation time of about

a. one hour
b. twenty minutes
c. four hours
d. ten hours

Answer 22

b. twenty minues

Question 23

Which of the following methods gives an accurate viable count?

a. nephelometry
b. electronic particle counter
c. colony count
d. turbidity measurements

Answer 23

c. colony count

Question 24

When the growth rate equals the death rate, the culture is in the

a. lag phase
b. log phase
c. stationary phase
d. death phase

Answer 24

c. stationary phase

Question 25

Spores are not usually found in the

a. lag phase
b. log phase
c. stationary phase
d. death phase

Answer 25

a. lag phase

Question 26

Bacteria are most susceptible to antibiotics in the

a. lag phase
b. log phase
c. stationary phase
d. death phase

Answer 26

b. log phase

Question 27

Cultures for microscopic examination for motility should be in the

a. lag phase
b. log phase
c. stationary phase
d. death phase

Answer 27

b. log phase

Question 28

0.1 mL of urine is added to approximately 20mL of molten agar, mixed well, and poured into a petri dish. Inspection of the plate the following day showed 86 small white bacterial colonies and 24 large yellow colonies.

a. What is the total colony count in CFU/L (SI units)?
b. What is the colony count of the large yellow colonies in SI units?
c. Is this a viable or total colony count?

Answer 28

a. 1.1 x 10^6 CFU/L
110 colonies/0.1mL x 1000mL/L = 1100000

b. 2.4x10^5 CFU/L
24/0.1mL x 1000mL/L = 240000

c. viable