Lab Practical 1 Flashcards
Rods. Can have rounded, flat, or tapered ends. Can be motile or non-motile.
Bacilli
Bacteria may occur singly, in chains, in a tetrad, or irregular masses. Most are nonmotile because they lack flagella
Cocci (spherical)
Bacteria are motile, using axial filaments, which are a type of flagella that originate form both ends of the cell and wrap around the cell body. Rotation of these causes the cell to move in a cork screwlike motion.
Spiral or curved
Agar plates
Incubated upside down to prevent moisture from collecting on the cultures and ruining them, spreading the inoculated organisms around. Agar-agar, the solidifying agent, becomes liquid when boiled and solid at about 42C.
Transfer from broth to broth
TSB of E. coli. Use a loop, swirl in the culture, swirl in the in sterile broth, flame the loop. 37C.
Slant to slant
TSA slant to sterile agar slant. E. coli. Use loop to draw culture, streak surface of sterile slant in a serpentine manner, flame loop
Plate to slant
TSA plate, slant, bacterial colonies. Use a needle. Raise the lid of the plate only a little to prevent contamination, touching the center of the colony with the needle. Streak the slant with the needle, not gouging it. Flame
Endospores require what to die?
Sterilization in the autoclave at 121C for 15-20 minutes, at 15 pounds per square inch.
What is used to clean living tissue? Surfaces?
Antiseptic is for living tissues. Disinfectant is for surfaces (leave covered for 20 minutes on a spill)
Hospital acquired infections are?
Nosocomial infections
Part of the microscope that varies intensity of light. Part of the microscope that reduces the intensity of light below the lower limit allowed by voltage control.
Light intensity control
Neutral density filter
Explain the objective lenses
10x is yellow, and 20x is green (coarse adjustment). 40x is blue, and 100x is white (fine adjustment). White is the oil immersion lens. Low power, high dry, oil immersion.
The resolving power of the microscope is a function of
The numerical aperture of the lenses and the wavelength of the light.
The ability of a lens to completely separate two objects in a microscopic field.
Resolving power, on most microscopes 1000x. It’s a function of the wavelength of light and the numerical aperture. Limit of resolution is typically 0.2um. Objects closer than that would not be seen as tow distinct objects.
A mathematical expression that describes how the condenser lens concentrates and focuses the light rays from the light source.
Numerical aperture. Its value is maximized when the light rays are focused into a cone of light that then passes through the specimen into the objective lens. The greater the loss of refracted light, the lower the numerical aperture. 0.2um
Oil immersion
This oil has the same refractive index as glass. When used it forms a continuous system that limits the loss of light due to refraction. Malaria caused by the Plasmodium species.
What microscopy is used for transparent delicate living organisms?
Which pathogenic microbe?
Darkfield. The darkfield stop is placed below the condenser, so that only oblique rays strike the objects being viewed.
Used to identify a spirochaetes in exudates from syphilitic lesions.
Goals of making a smear
Cause the cells to adhere to the slide so that they aren’t washed off during the staining and washing. Also makes sure that shrinkage of cells doesn’t occur during staining, so that distortion doesn’t result. Prepare thin smears, because thick smears obscure details and entrap the stain
Heat fixation
Allow the slide to air dry first. Then pass through the flame a few times, not leaving it too long or else the slide will shatter. It fixes organisms to the slide and kills anything that’s still living. Used in simple staining, gram staining, and spore staining.
Simple staining
The use of a single stain to color a bacterial cell. Uses S. aureus and E. coli. Bacterial cells are negative. Basic dyes are cationic (+) because of the chromophores, color-bearing ions. Methylene blue (!), basic fuchsin, crystal violet. Can determine morphology, size, and arrangement.
Negative stains
Uses B. megaterium. They are acidic (-) anionic with a chromophore that does not penetrate the cell but is rather repelled by the negative bacterial cell. Negative/indirect staining because the background is stained. India ink and nigrosin. Can be used in studying morphology, capsules, no shrinkage of cells because no heat fixing, so size determinations are more accurate. Good for observing spirochaetes.
Reactions that take advantage of the that cells or structures within cells display dissimilar staining reactions that can be distinguished by the use of different dyes
Differential staining, eg gram staining
Gram stain procedure and microbes
E. coli (-) bacillus with S. aureus (+) cocci
M. catarrhalis (-) cocci with B. megaterium (+) bacillus
Primary: crystal violet, 1 minute
Mordant: gram’s iodine, 1 minute
Decolorizer: ethyl alcohol, 20-30 seconds
Counterstain: safranin, 1 minute
Spore stain overall
The bacteria of the genera Bacillus and Clostridia become endospores when they exhaust essential nutrients. They have a protein coat, or exosporium, that forms a protective barrier around the spore. Resistant to heat, radiation, acids, and many chemicals like disinfectants.
