Lab exercises 11-16
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T or F: Polysaccharides, polypeptides, and glycoproteins have all been found in capsules
After using crystal violet on a capsule cell as the primary stain, what is copper sulfate used for? (2 things) ****COPPER SULFATE NOT USED IN LAB****
- It is the decolorizing agent, by removing excess primary stain and color from the capsule.
- It also acts as a counterstain by being absorbed into the capsule, turning it a light blue or pink.
T or F: when capsule staining, you should heat fix the sample
What three chemical substances have been identified in bacterial capsules?
- 1. Polypeptides
- 2. Sugars
- 3. Proteins
What is relationship between presence of capsules and bacterial pathogenicity?
They interfere with phagocytosis, resist dehydration, and to help stick
What certain diseases can the presence of flagella indicate?
Whooping cough, meningoencephalitis, UTI, PNA, tyhpoid fever, cholera
T or F: If a bacteria is unstained, its flagella can only been seen in an electron microscope
In order to observe flagella with a light micrscope, its thickness is increased by coating them with __a__ such as tannic acid and potassium alum, and staining them with basic __b__
How do flagella increase in size to be visible in a light microscope?
A flagella stain employs an alcoholic solution of crystal violet as primary stain, and then a mordant. As alcohol evaporates during staining, the crystal violet forms a precipitate around flagella, increasing the apparent size.
How is a flegalla stain done?
- 1. Add bacterium with loop to slide with 3 drops of distilled water, and spread it around.
- 2. Let it air dry for 15 min then cover with the mordant for 4 mins.
- 3. Rinse throughly with water, add a paper towel on smear and soak paper with stain.
- 4. Place the slide on top of a boiling water bath for 5 mins.
- 5. Rinse with water and let it sit for 1 min, air dry, and use oil immersion to observe.
Why are flagella so difficult to stain?
They are very fragile
Why did we use a young culture for the flagella stain?
Old culture bacteria drops their flagella sooner
Why must the glass slide be free of grease and oil before staining for flagella?
Grease prevents flagella from sticking to the slide
Name these 4 types of flagella arrangements
- a. Minotrichous
- B. Lophotrichous
- C. Amphitrichous
- D. Peritrichous
What does mordant do for flagella?
Helps flagella stick to dye
What happens to size of flagella when they are stained?
Become thicker due to mordant
Which microscopy do you use to look for motility and which one do you use to find flagella?
- Hanging drop slide for motility
- Flagella stain for making the flagella thicker to see
What are the two types of pipettes?
- 1. Blow-out pipette (also called serological pipette)
- 2. To-deliver pipette (Mohr measuring pipette)
T or F: With the "serological pipette", after the proper amount of liquid has been delivered, liquid will remain in the tip of pipette and should not be delievered.
False: the final few drops of liquid must be emptied in order to deliver the correct volume. Only the Mohr measuring pipette should the remaining liquid not be eliminated
What is subculturing?
When microorganisms are transferred from one culture medium to another using specific procedures and aseptic technique.
What is desiccation?
Refers to drying out of a living organism
What 2 ways can microorganisms be stored in a state of dormancy?
- 1. Regrigeration
- 2. Desiccation
What oil can be used to seal cultures to maintain them from drying out?
Sterile mineral oil
What is lyophilization?
Freeze-drying a culture for long periods to eliminate the need for periodic transfers.
What is the purpose of flaming in the aseptic technique?
To steralize the needle/loop
In subculturing, when do you use the inoculating loop?
To transfer bacteria to a media
What are signs of growth in a liquid medium?
Cloudy and it smells
What is the spread-plate technique used for?
An easy, direct way of obtaining a pure culture. This is done by spreading out a dluted bacterial mixture on a plate, and letting some of the disperesed develop into isolated colonies.
This term refers to a large number of bacterial cells on solid medium, which is visible to the naked eye as a discrete entity
What is selective media used for? What are examples?
- Favors the growth of a particular microorganism
- Ex: bile salts or dyes like basic fuchsin favor growth of gram negative bacteria by inhibiting the trowth of gram-positive ones.
What is differential media used for? What are examples?
- Used to distinguish between different groups of bacteria and even permit tentative ID of microorganisms based on their biological characteritics and rxns.
- Blood agar is an example, and distinguishes between hemolytic and nonhemolytic bacteria.
What are two very important differential and selective media that are used to isolate and partially ID bacteria?
- Mannitol salt agar
- Eosin methylene blue agar
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