Growth L6 P2

What is bacterial growth in terms of cell division

An increase in cell size followed by an increase in the number of cells

How do most prokaryotes divide

By binary fission

What is generation time

The time required for one cell to divide into two cells

What happens to the number of cells with each generation

The number of cells doubles

What factors can affect generation time

The species of bacterium and the environmental conditions

What is the usual range of bacterial generation times

About 20 minutes to 3 hours

What is exponential growth

Growth where the number of cells increases exponentially because cells repeatedly double

What is a growth curve

A graph showing how a microbial population changes over time

Why are growth curves usually plotted using the logarithm of cell number

Because bacterial populations become very large, making them difficult to plot on a regular linear scale

What is mean growth rate

The number of generations per unit of time; it is the inverse of generation time, r = 1/g

What are the four phases of a typical bacterial growth curve

Lag phase, exponential phase, stationary phase, and decline phase

What is a batch culture

A culture grown in a closed vessel containing a single batch of growth medium

What happens during the lag phase

Cells adapt to the growth conditions before rapidly dividing

What happens during the exponential phase

Cells divide at a regular rate and the population increases exponentially

Why is the growth rate highest during the exponential phase

The generation time is lowest and cells are dividing regularly

What does the note “more growth than death” mean during exponential growth

The number of new cells produced is greater than the number of cells dying

What happens during the stationary phase

The number of cells stops increasing

What causes the stationary phase

Essential nutrients become depleted and/or waste products accumulate

Why are many microbes in the real world usually in the stationary phase

Their natural environments often limit nutrients or cause waste to accumulate

What happens during the decline phase

Cells begin dying, causing the population to decrease

What are the four main methods used to measure bacterial growth

Microscope counts, plate counts, most probable number, and turbidity

Which bacterial growth measurement methods are direct methods

Microscope counts, plate counts, and most probable number (MPN)

Which bacterial growth measurement method is indirect

Turbidity, measured using a spectrophotometer

What is a microscope count

Directly counting bacterial cells using a microscope and a counting chamber with a known volume

What is an advantage of microscope counts

They are very fast because no incubation is required

What is a disadvantage of microscope counts

They count both living and dead cells

The bacteria must be still, or move very slowly to count.

How is the number of bacteria per mL calculated from a microscope count

Number of cells counted ÷ volume of the area counted

What is a viable count

A direct method that estimates living bacteria by counting the colonies they produce

What is the basic idea behind a plate count

Each viable bacterial cell produces a visible colony, so colonies are counted to estimate viable cells

What is the goal of a plate count

To spread bacteria out enough that individual cells form separate visible colonies

# of colonies = # of bacteria

What are colony-forming units (CFU)

Units representing viable cells that can grow into colonies

How are viable cells reported from a plate count

As colony-forming units per millilitre (CFU/mL)

What is a spread plate

A method where bacteria are spread across the surface of a solid medium

What is a pour plate

A method where bacteria are mixed throughout molten solid medium

What is an advantage of viable plate counts

They count only living cells that can grow into colonies

What is a disadvantage of viable plate counts

They require incubation, so they are slower

Why are serial dilutions needed before plating many bacterial samples

An undiluted sample may produce too many colonies to distinguish individually

What is a bacterial lawn

A plate completely covered with bacterial growth, making individual colonies indistinguishable

What is the recommended countable range for a plate

About 20–200 individual colonies

What is the purpose of serial dilutions

To reduce the number of bacteria to a countable level

What is added to the diluent during the first step of a serial dilution

A known amount of the original sample

What happens after making the first dilution

A known amount of the first dilution is transferred into more diluent

Why is the serial dilution process repeated

To progressively reduce the bacterial concentration

What is a dilution factor

The reciprocal of the dilution made

What is the dilution factor for a 1/10 dilution

10

What does CFU stand for

Colony-forming unit

How is the original bacterial concentration calculated

CFU/mL = (number of colonies × dilution factor) ÷ plating factor

What does the number of colonies represent in a CFU/mL calculation

The number of colonies counted on the plate, usually from a countable plate

What is the plating factor

The volume of the bacterial suspension plated, expressed per mL or g

What is the formula for an individual dilution

Amount transferred from the previous dilution ÷ total final volume

If 1 mL of sample is added to 9 mL of diluent, what is the dilution

1/10

What is the dilution factor for a 1/10 dilution

10

If 1 mL of a 1/10 dilution is added to 9 mL of diluent, what is the new dilution

1/100

What is the dilution factor for a 1/100 dilution

100

Why is the dilution factor the reciprocal of the dilution

It converts the diluted concentration back to the original concentration

What colony range should be used for calculating CFU/mL

About 20–200 colonies

What is the formula for original bacterial concentration

Original concentration = (colonies × dilution factor) ÷ plating factor

What happens if the undiluted bacterial sample is plated

It usually produces a bacterial lawn with indistinguishable colonies

Why are very small samples difficult to use for plate counts

They may contain too few bacteria to produce enough colonies for accurate counting

What is the Most Probable Number (MPN) method

A statistical method used to estimate the number of microorganisms in a sample

When is MPN especially useful

For samples with low microbial counts, spreaders, or interfering particles

What is done during an MPN test

Serial dilutions are prepared and tubes are observed for microbial growth

What must be true about the microorganism for MPN to work

The microorganism must be capable of growing in the laboratory

How is a positive MPN tube identified in your lab

By a change in colour and production of gas

How is an MPN result determined

The number of positive tubes is matched to a specific MPN table

What do the confidence limits of an MPN result tell you

The range in which the actual number of bacteria is expected to fall

In the lecture's MPN example, what was the positive-tube pattern

5–3–1

What was the most probable number in the lecture's example

110 bacteria per 100 mL, with 95% confidence

What is one advantage and one disadvantage of MPN

Advantage: high statistical confidence; disadvantage: requires incubation and only works for some bacteria

What is spectrophotometry used for in measuring bacterial growth

It indirectly estimates cell numbers by measuring light scattering

What is turbidity

The cloudiness of a bacterial culture caused by cells scattering light

What happens to turbidity as the number of bacteria increases

The solution becomes more turbid

How is turbidity measured

Using a spectrophotometer

Why is light scattering used instead of light absorption to measure bacterial growth

Light absorption can vary with culture conditions and bacterial species

What is one advantage of using turbidity to measure bacterial growth

It is very quick and useful for routine analysis

What are two disadvantages of turbidity measurements

They require a standard curve and cannot measure very low cell numbers

Does turbidity distinguish between living and dead cells

No. Both living and dead cells contribute to the measurement

Why can turbidity measurements usually only be used with a pure culture

Different bacterial species scatter light differently

What is a standard curve used for

To determine an unknown concentration by comparing it with measurements from known samples

How can OD measurements be used to determine generation time

OD can be converted to CFU/mL using a standard curve, then used to create a growth curve and determine generation time