Lab Final Flashcards
(22 cards)
Oxygen requirement:
- Purpose
- Procedure
- Results
Purpose: To determine the oxygen requirement of bacteria
Procedure:
- label tubes with initials and name of organism
- Sterilize loop
- Gently shake bacteria to disperse
- Flame top of tubes and take one loop from broth culture and inoculate FTM tube by going straight down
- Reflame and recap tube
- Sterilize loop and incubate @37c
Results:
Obligate aerobe, obligate anaerobe, facultative aerobe, microaerophile, aerotolerant anaerobe (even growth everywhere)
Why do we use beads for the antibiotic evaluation experiment?
To uniformly spread bacteria
Antibiotic evaluation:
- Purpose
- Procedure
- Results
Purpose: to determine which drug a microorganism is most sensitive to
Procedure:
- Label bottom of plate with initials and organism
- Use cotton swab to inoculate entire plate with bacteria
- Dry for 5 mins then use sterilized forceps to take out each one antibiotic disc from each cartridge and place on surface of agar of plate (space far apart from each one)
- secure and incubate @37c for 16-18 hours
- use metric ruler to calculate zone of inhibition (diameter)
Results:
Vary for each bacteria
Sa: everything effectiveish
Ecoli: only GM
Alcohol evaluation:
- Purpose
- Procedure
- Results
Purpose: To assess the effectiveness of alcohol on removing microorganisms from the skin
Procedure:
- Divide TSA plate into 4 (A,B,C,D)
- You will swab one thumb with either alcohol wipe or 70% isopropyl later
- Gently press unwashed thumb into A and again into B
- Gently press other unwashed thumb into C
- Disinfect that same thumb using either mentioned above and then press into D
- Incubate @37c for 24-48 hours
Results:
D least amt of bacterial growth
What is the formula for percent reduction?
(# colonies from 1st thumb- # colonies from 2nd thumb)/ # colonies from 1st thumb x 100
Antiseptic (Chemical germicide)
- Purpose
- Procedure
- Results
Purpose: To examine the effectiveness of different germicides
Procedure:
- label 6 broths dilution 1,2,3, sterile water 4,5 and control
- label 5 petri dishes dil 1,2,3 and sw 4,5
- one loop of bacteria to control and set aside
- use forceps and add 4 beads to culture and incubate for 10 mins. Leave one bead alone
- Transfer broths to petri dishes in respective names
- Add one bead each from culture to dil 1,2,3 and sw 4
- take uninfected bead and put in control
Results:
Dil 1,2 or 3 will have no growth depending on the disinfectant (2 for lysol) and sw 5 since no bacteria was added
Gelatinase
- purpose
- procedure
- positive and negative result
Purpose: to see if bacteria has enzyme gelatinase
Procedure:
- sterilize needle and inoculate
- incubate on ice for 5 mins
+ if solid - if liquid
Methyl Red test
- purpose
- procedure
- positive and negative result
Purpose: to determine mixed acid fermentation
Procedure:
Starts yellowish clear
- Inoculate one loop of bacteria
- Incubate at optimal temp (20 deg C for you!!!!)
Day 2
- Add 4-5 drops of methyl red
+ red - no change
VP test
- purpose
- Indicators
- procedure
- positive and negative result
Purpose: To see if acetoin is produced as a byproduct
Indicators: Barrit’s reagent A and B
Procedure:
Start with yellow/clear media
- Inoculate one loop of bacteria
- Incubate in optimal temperature
Day 2
- Add 20 drops of VP A and 20 drops of VP B (KOH: potassium hydroxide)
- Tap and mix every five minutes for 2.5- 3 hours
Results:
+ top turns red - entire tube remains the same
Mannitol Salt Agar plate
- purpose
- indicator
- procedure
- positive and negative result
Purpose: selective for staphylococci and differentiates for mannitol fermentation
Indicator: phenol red (gives agar red colour)
Procedure:
- draw line on plate
+ if turns yellow (only S. aureus)
Eosin Methylene Blue plate
- purpose
- indicators
- procedure
- positive and negative result
Purpose: to see if bacteria ferments lactose
BOTH SELECTIVE AND DIFFERENTIAL PLATE
Indicators:
Eosin Y and methylene blue combine to form dark purple
Procedure:
- draw line on plate
+ purple normal levels or metallic green/black aggressive levels - clear colour
coliforms: bacteria that grows in intestine and able to use lactose for fermentation purposes
MacConkey Plate
- purpose
- procedure
- positive and negative result
Purpose: to selectively identify enterobacteriaceae
Procedure:
Start light pink colour
- draw line on plate
+ purpleish red normal growth - clear yellowish
Durham tube sugar fermentation
- purpose
- Indicators
- procedure
- positive and negative result
Purpose: to determine if bacteria can use specific sugar for fermentation
Indicator: phenol red
Procedure:
tiny tube in tube
+ yellow (fermentation but no gas)
+ yellow but some empty (ferm + gas)
- still red
Nitrate reduction test
- purpose
- procedure
- positive and negative result
Purpose: to see if bacteria can change nitrate to nitrites
Procedure:
- Inoculate 1 loop of bacteria
- Incubate at optimal temperature
Day 2
- Add 3 drops of nitrate A and add 3 drops of nitrate B
- Mix every 5 minutes for 30 mins
+ red/ pink - no colour change
Catalase production test
- purpose
- procedure
- positive and negative result
Purpose: to determine if bacteria is positive for catalase which breaks down H2O2 into H2O and O2
Procedure:
- Fishtail one loop of bacteria
- Incubate at optimal temp
Day 2
- Add five drops of H2O2
+ bubbles - no bubbles
Oxidase production test
- purpose
- procedure
- positive and negative result
Purpose: to determine if oxygen is the final electron acceptor
Procedure:
- One loop of bacteria on the plate
- Incubate at optimal temp
Day 2
- Grab two long filter papers and place it on lid of the plate
- Add one drop of oxidase solution to each filter paper
- Grab loop of bacteria and place on top of oxidase/ filter paper.
+ turns dark blue/ purple within 30 secs - remains clear/yellow
Starch hydrolysis
- purpose
- procedure
- positive and negative result
Purpose: to determine if bacteria produces exoenzyme amylase which breaks down starch
Procedure:
Start with clear colour plate containing starch
- Loop of bacteria on plate
- Incubate at optimal temp
Day 2
- Add iodine to plate
+ Clear area around bacterial growth means amylase produced - if area around bacteria turns black/dark blue
Casein hydrolysis
- purpose
- procedure
- positive and negative result
Purpose: to see if bacteria has casease which breaks down casein
Procedure:
start with white colour milk plate
- draw line
+ clear area around bacterial growth - no clear area
Urea hydrolysis
- purpose
- procedure
- positive and negative result
Purpose: to see if bacteria has enzyme urease
Procedure:
Start with clear/yellow colour media
- Inoculate one loop of bacteria
- Incubate at optimum temperature
Day 2
- Observe results
+ bright pink colour - yellow/ clear
Tryptophan hydrolysis (indole)
- purpose
- procedure
- positive and negative result
Purpose: to see if bacteria contains enzyme tryptophanase
Procedure:
Start clear/ yellow media
- Inoculate one loop of bacteria
- Incubate at optimal temperature
Day 2
- Add 12 drops of Kovack’s solution
+ red layer on top - all yellow and oily on top
Hydrogen sulfide production test
- purpose
- procedure
- positive and negative result
Purpose: To determine if bacteria can produce hydrogen sulfide
Procedure:
- Inoculate tube with bacteria
- Incubate
Day 2
- Observe
+ black/ red with black - red
Citrate utilization
- purpose
- procedure
- positive and negative result
Purpose: to determine if bacteria is able to use citrate as a sole source of carbon
Procedure:
Start with green coloured slant
- Take one loop of bacteria, then fishtail on slant
- Grab bacteria with needle, then stab slant
- Incubate at optimum temperature
Day 2
- Observe colour change
+ Dark bluish colour - Green colour
