BIOL 2401 Week 7 Unit 5.2 & 5.3: Growing microbes in the lab & Measuring bacterial growth

Liquid media

Media used to culture large amounts of bacteria

Solid media

Used to isolate colonies in mixed samples

Agar

Polysaccharide that is used as a solidifying agent

Chemically defined media

Media that is tailored to the specific needs of a microbe, such as an autotroph. Complicated

Complex media

Media that is a mixture of extracts and digests. Not very precise but more commonly used. E.g. tryptic soy broth (TSA)

Selective media

Media that inhibits the growth of unwanted microbes, and encourages growth of wanted microbes

Differential media

Media that uses a chemical to visually distinguish different species from one another

How is mannitol salt agar a selective media?

It is selective since it has a high salt concentration, only allowing growth of halotolerant bacteria

How is mannitol salt agar a differential media?

It is a differential media since fermentation of mannitol changes the color of medium. Helps distinguish different species of staphylococcus (epidermidis vs aureus)

Blood agar

Differential medium which shows if bacterial coloonies are capable of hemolysis

Beta hemolysis

Complete hemolysis with a clear background

Alpha hemolysis

Partial hemolysis with a green background

Gamma hemolysis

No hemolysis with the media unchanged

Hemolysis (in agar)

Destruction of red blood cells by bacteria, such as in blood agar

Methods to allow anaerobes to survive

Reducing media & anaerobe jars → chemically removes oxygen

Anaerobe chamber → excludes oxygen

Refrigeration (preserving bacterial cultures)

Short-term method to preserve bacterial cultures. Slows down bacterial metabolic activity, keeps for 2-4 weeks

Deep freezing (preserving bacterial cultures)

Long-term method to preserve bacterial cultures. Pure cultures are quick frozen at -50C to -95C. Keeps for 6-12 months

Lyophilization

Long-term method to preserve bacterial cultures. Microbes are quickly frozen, water is removed to make a powder. Keeps for a few years

In vivo meaning

In whole organisms. E.g. animals or plants

In vitro

In cells in an artificial environment. E.g. petri dish

How must viruses be grown?

Viruses must be grown inside cells. They cannot survive without a host cell

“Mediums” for growing animal viruses

Embryonated eggs, living animals, cell cultures (e.g. HeLa immortalized cells)

Process of growing bacteriophages

Add phages to lawn of bacterial growth → phages will lyse susceptible bacteria, shows as clearings on plate

Generation time

The time needed for a cell to divide, dependending on species and environment. Typically ranges from 20 minutes to 3 hours

In binary fission, how many cells are produced?

Two cells are produced per one cell (doubles)

Lag phase

First phase of bacterial growth curve. Intense preparation by bacteria for growth, but no growth yet

Log phase

Second phase of bacterial growth curve. Logarithmic or exponential population increase

Stationary phase

Third phase of bacterial growth curve. Period of equilibrium, where death rates = production rates

Death phase

Fourth phase of bacterial growth curve. Population decreases logarithmically

Turbidity

Used to indirectly count bacteria. If turbid, appears opaque. Turbidity is measured using a spectrophotometer

Spectrophotometer

Device used to measure turbidity

Disadvantages of turbidity

Cannot be use for dilute cultures, also counts dead cells, species absorb light differently

Direct microscopic count

Observing a vry small amount of liquid and individually counting the organisms

Advantages and disadvantages of direct microscopic count

Advantages: Results obtained right away

Disadvantages: Cannot be used for very dilute cultures, counts dead cells, not for motile cells

Plate counts

Where diluted sample is spread on medium, and each bacterium results in a visible colony which can be counted. # of colonies = # of bacteria

CFU (Colony Forming Units)

A unit that estimates the number of microbial cells in a diluted sample through counting its individual colonies

Advantages and disadvantages of plate counts

Advantages: Counts live cells, is quite accurate

Disadvantages: Requires incubation (delay), requires dilution

Serial dilution

A process where a known amount of sample is continually diluted. Repeated several times to create a very dilute solution