Lab Exam I Flashcards
Definition of Blood Agar Plate
Differential medium (RBC from sheep) is used to observe hemolytic activity of bacteria. Most commonly used to differentiate Streptococcus pneumoniae (alpha hemolysis) and Streptococcus pyogenes (beta hemolysis).
alpha, beta, and gamma hemolysis
alpha hemolysis - partial lysis, brown or green
beta hemolysis - complete lysis, clearing
gamma hemolysis - no hemolysis, no change
Procedure for smear preparation
- Place small drop of water on slide.
- Spread small amount of culture.
- Air dry.
- Heat fix by passing slide over flame 3 times.
- Circle smear with wax pencil.
Purposes of Quadrant Streak Plate Technique
- obtain pure culture
- determine if culture was pure or mixed
- determine colony morphology
What is the purpose of streaking a plate in 4 quadrants?
Streaking the plate in 4 quadrants dilutes the culture, leaving bacteria further and further apart. Isolated colonies arise from a single cell.
Label of each plate
- Name
- Specimen ID
- Medium
- Incubation conditions
- Date
Terms used to describe colony morphology
- Shape (circular, irregular)
- Surface (dry, moist, rough, wrinkled)
- Margin (smooth, wavy, lobed, branched)
- Elevation (flat, raised, convex, raised in center)
- Opacity (transparent, opaque, transluscent, iridescent)
- Pigmentation (white, tan, cream red, yellow)
- Other
- agar color change
- isolated colonies, swarmer
Purpose of TSA Slant
Used to grow and maintain culture. Not for isolating colonies. Advantages of Slants include:
- easier to maintain sterility
- less desiccation for long-term storage
- less media used
Definition of Gram stain
Used to determine if bacteria is Gram positive (purple) or Gram negative (pink)
Procedure for Gram staining
- Prepare smear
- Soak in Crystal Violet for 1 minute; rinse with water
- Soak in Gram Iodine for 1 minute; rinse with water
- Decolorize with EtOH until runs clear; rinse with water
- Soak in Safranin for 1-3 minutes; rinse with water
Define Kirby Bauer/Antibiotic Sensitivity Test
Purpose is to demonstrate the difference between broad and narrow spectrum antibiotics, using Gram-negative bacterium (E. coli) and Gram-positive bacterium (Staphylococcus epidermidis).
Describe how Kirby Bauer Test is interpreted
- Zone of inhibition around each antibiotic disk measured in mm and compared against chart of interpretive standards.
- Broad spectrum antibiotics will be effective against both Gram-positive and Gram-negative bacteria.
- Narrow spectrum antibiotics will be effective only against a subset of bacteria.
Define MacConkey Agar (MAC)
A Selective and Differential medium used to detect and differentiate Gram-negative enteric bacteria based on ability to ferment lactose. E. coli is lactose positive. Salmonella, Shigella are lactose negative.
Selective component of MAC
- Crystal violet selects for Gram-positive
- Bile salts select for enteric
Differential component of MAC and indicator
- Lactose
- Neutral red pH indicator turns red when pH <6.8, remains colorless when pH >6.8
Define Mannitol Salt Agar (MSA)
A Selective and Differential medium used for isolation and differentiation of pathogenic Staphylococci, principally Staph. epidermidis and Staph. aureus.
Selective component of MSA
- 7.5% NaCl selects for Staphylococci capable of growing in high salt concentration
Differential component of MSA
- Mannitol
- Phenol red
- pH < 6.8 yellow
- pH 7.4-8.4 red
- pH > 8.4 pink
- Staph. aureus (pathogenic) capable of fermenting mannitol, producing acid, which turns the pH indicator yellow.
Define Catalase Test
- Detects the presence of catalase, and enzyme found in aerobic and facultative anaerobic bacteria.
- Superoxide dismutase converts superoxide radicals into H2O2 and O2. Catalase converts H202 into H2O and O2.
- Differentiate between Staphylococcus (catalase positive) and Streptococcus (catalase negative).
Define KOH Test
A nonstain alternative to the Gram stain test. If Gram-negative bacteria is mixed with KOH solution, the KOH crosses the thin cell wall and lyses the cells. Chromosomal material is released into the emulsion. The DNA unwinds into a viscous, stringy material. Gram positive bacteria remain unchanged.
Define Snyder Agar
Selective and differential agar typically used to determine the relative risk of an individual developing dental caries (due to Lactobacilli). Agar is green.
Selective component of Snyder Agar
pH 4.8
Differential component of Snyder Agar
- Glucose
- Lactobacillus ferments glucose to produce lactic acid.
- The green agar turns the agar yellow faster in the presence of lactobacilli.
Procedure for conducting Snyder Agar Test
- Collect saliva in small cup.
- Obtain molten Snyder agar from hot water bath. Do not allow to let cool.
- Pipette o.2mL of saliva into tube.
- Vortex tube.
- Incubate at 37C
yellow in 24 hours –> high susceptibility to dental caries
yellow in 48 hours –> moderate susceptibility
yellow in 72 hours –> slight susceptibility
Define Thioglycolate Broth
Used to determine oxygen requirement for bacteria. An indicator is added to detect presence of free oxygen. Observing location of growth in the column will indicate oxygen requirement.
