LAB Week 5 #1

Robert Koch

Father of medical microbiology.

Knew if he wanted to prove a particular bacterium causes a specific disease, it would be necessary to isolate the agent from all other bacteria and characterize the pathogen.

Pure Culture

A culture that contains only a single kind of organism.

Contaminated Culture

A culture that contains a desired organism, but also contains unwanted organisms.

What can we study with a pure culture?

The cultural, morphological, and physiological characteristics of an individual organism.

What are two commonly used methods for obtaining pure cultures?

Streak Plate and Pour Plate.

What do both streak plate and pour plate methods require?

Diluting the bacterial cells in a sample to obtain an isolated pure colony(ies).

Pure Colony

The identical progeny of the original cell that can be picked and used for further study of the bacterium.

Why is it important to master the streak plate?

Future exercises will depend on this technique to obtain isolated cultures.

What is the most popular streak plate procedure?

Quadrant Streak Plate.

Streak pattern that looks like a trellis:

Radiant streak

Materials Needed for Streak Plate Procedure.

Electric Hot Plate

Procedure for pouring an agar plate for streaking.

1. Place nutrient agar pour in beaker of boiling water for 5 minutes.

2. Cool down nutrient agar to 50 degrees Celsius.

3. Remove the cap from tube and flame the open end of the glass.

4. Pour the contents of tube into the bottom of the agar plate and allow it to solidify

What is a consequence of not letting the agar cool before pouring it into a plate?

Condensation / moisture will cover the top and cause the organisms to spread randomly all over the surfaces.

Pour Plate Method.

The method of separating one species of bacteria from another through the dilution of one loopful of organism across three tubes of liquefied nutrient agar resulting in one plate containing an optimum number of organism for successful isolation.

Completing Pour Plate Method.

Once the loop dilutions are completed you will take the inoculated agar pours out of the water bath one by one and pour slowly into the corresponding numbered petri dishes. Make sure you don't produce bubbles and completely fill in the plate by gently tilting the plate around. Let the plates solidify. Once solid, incubate the plates bottom side up at 22 degrees Celsius for 24 to 48 hours.

Why do we incubate plates upside down?

Condensation from the dish lid will be deposited on the agar surface dispersing the organisms, disrupting the desire growth pattern and preventing the formation of isolated colonies.

Subculturing

A new pure culture made by transferring some or all cells from a previous culture to fresh growth medium.

Sub-culturing Techniques

You can transfer an isolated colony from a streak or pour plate to a tube of nutrient broth or a slant of nutrient agar.

Microscopic Examination

The only way to be sure that you have pure cultures.

Best Way to Examine Pure Culture Under Microscope.

Gram-Stained smears allow bacteria to be separated in the case of contamination.

What do higher temperatures do to the S. Marcenscens?

Inhibit production of its red pigment.

What is S. Marcescens described as?

Gram-negative short rod.

What is M. Luteus described as?

Gram-positive coccus.