Micro Lab Practical 1
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Micro Lab Practical 1
How long is the sterilization in an autoclave?
What is the temp and pressure for autoclave sterilization?
121 degrees; 15lbs pressure
Why so much pressure in the autoclave?
To penetrate the thick coat of the endospore and destroy the genome of the bacteria.
Define Aseptic technique.
A procedure used to inoculate a pure culture without (or limited) contamination to media for growth.
What is the purpose of a streak plate?
To get isolated colonies
What kind of colony does Serratia Marcescensa produce?
Small & red
One flagella at the end of a bacterium.
One flagella on either end of a bacterium.
More than one flagella at one end of a bacterium.
Flagella all around the bacteria.
When should you use safety goggles instead of safety glasses?
If you wear contacts you should wear goggles when dealing with endospores and acid fast bacteria due to the steam heating step
What can be disposed of in the biohazard BAG?
paper towels, Petri dishes, gloves
What can be disposed of in the BASKET?
tubes (at an angle)
What can be disposed of in the BOX?
broken glass, slides (contaminated slides in the biohazard box)
What is the indicator on the tubes that are motile?
What is added to the media to stop the non-motile bacteria from moving out of the slab?
How do you innoculate a motility tube (an agar deep)?
With a needle
What is an agar plate?
solidifying agent - polysaccharide seaweed nutrients for growing bacteria
Define agar deep.
Used for motility; allowed to cool & solidify in an upright position
This type of agar is used to keep stock cultures longer.
Why agar for growth media?
Because it cannot be digested; it is more solidifying & remains solid
What is agar?
What is the purpose of agar plates, deeps, and slants?
To produce a pure culture; its not digestible
What is nutrient broth?
a liquid that has nutrients which support the growth of microbes (does not contain agar)
What is a Brewer jar used for?
To create an anaerobic environment for bacteria growth
What genus of bacteria grow in the Brewer jar?
What are 4 types of Clostridium?
What two gases are generated when water is added to the Brewer jar?
hydrogen & carbon dioxide
What is the inorganic catalyst in a Brewer jar?
What is the mb strip in the Brewer jar?
methylene blue, which confirms absence of oxygen
-blue = oxydized
-red = reduced
What happens to the color of the mb strip in the Brewer jar?
It should turn white, meaning there is no oxygen. If it stays blue, it didn't work.
What is the purpose of the mb strip in the Brewer jar?
To make sure it works (white if it does, blue if it doesn't)
What kind of special gas environment is produced in the candle jar?
High concentration of CO2
Low concentration of O2
Which pathogen grows best in the candle jar?
S. pyogenes (can cause pharyngitis)
Name four pathogens that grow in the Brewer jar.
What group of microbes is this medium (fluid thioglycollate broth) used to cultivate?
Aerobic, Anaerobic, and/or microaerophilic
-based on growth at various levels in the media
What is the indicator in our tubes?
How does methylene blue indicate oxygen levels?
If O2 is present, it turns greenish.
Where would aerobes grow in this medium?
at the top
Where would anaerobes grow in this media?
at the bottom
Where would microaerophiles grow in this media?
right below the top
Where would facultative bacteria grow in this media?
What is the primary stain in a gram stain?
What is the mordant in a gram stain?
What is the decolorizer in a gram stain?
What kind of solvent is the decolorizer in a gram stain?
What is the counterstain in a gram stain?
What is the purpose of mordant (in a gram stain)?
it tightly binds the dye to the cell wall
What is the primary dye in an acid fast stain?
What is the decolorizer in an acid fast stain?
What is the counterstain in an acid fast stain?
Which genus is "acid fast"?
What are the two pathogens in mycobacterium (acid fast)?
What is the basis of an acid fast stain?
presence of mycolic acids in the cell walls of acid fast organisms
Why is the presence of mycolic acids in cell walls an issue?
Acid fast bacteria are resistant to chemical compounds and may be resistant to medications.
What procedure is used to drive the primary dye into the cell wall of an acid fast organism?
The waxy material in the cell walls of acid fast bacteria.
The decolorizer in an acid fast stain is called a "harsh" decolorizer because it destains all bacteria except the what genus?
What is the term for cloudy?
Term for "ring around the top".
What is the term for a material that settles to the bottom of the tube?
What is the term for "flaky" clumps in the medium?
What is the function of the endospore?
survival (NOT reproduction)
What are two genera that produce spores?
Name a pathogenic species for each genus that produce spores.
Bacillus = B. antracis, B. subtilis
Clostridium = C. tetani
Whats the difference between colony pigments and soluble pigments?
Colony - stain the colony
Soluble - Stain the whole plate; they sink into the agar
What kind of stain is this?
Capsule stain - a modified
A capsule stain uses acidic dye that stains the _______.
Slide, NOT the cell.
The purple aniline dye in a capsule stain stains the _______.
What is the capsule made of?
Why is the capsule a virulence factor?
It helps resist phagocytosis by WBCs.
Two pathogens of a capsule.
Serratia - what kind of pigment?
M. luteus plate: What is the “key” morphological characteristics of the colonies seen on this plate?
lemon yellow color
What procedure is this?
Hanging drop procedure
What is the purpose of a hanging drop procedure?
It allows longer observation because it does not dry out as fast; also allows you to see live motility and natural size
Why are stains used?
To increase contrast and make the bacteria easier to see
What kind of dye is this? What kind of stain?
Crystal Violet Dye
Simple stain (one kind of dye)
How much is the morphological unknown worth?
What are the first 2 things done in lab when given an unknown to ID?
1. Streak plate
2. Gram stain
A gram positive, rod shaped, spore-forming anaerobe that causes gangrene.
A species of gram-positive Clostridium that causes diarrhea.
What is the purpose of an acid fast stain?
To identify the genus mycobacteria (used for TB)
A type of negative stain where the background is stained and not the organism.
What does the gas pack generate in the Brewer jar?
sodium borohydride (NABH4) & CO2
What does the lever under the stage open and close to reduce or increase the amount of light?
iris diaphragm - regulates light intensity
Why is an agar deep used?
it promotes growth of a broad range of organisms;
it has an oxygen gradient from aerobic at top to anaerobic at bottom;
The depth ensures that the bottom portion is anaerobic