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138 Cards in this Set
- Front
- Back
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In aseptic transfer, why do you flame the neck of the test tube?
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Hot air is pushed out, prevent contamination of the culture.
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What's the main difference in procedure when preparing a smear from a solid medium versus a broth culture?
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For a solid medium, you must add one or two loopfuls of water to the slide before transfer the organism.
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Why do you pass a bacterial smear through a flame?
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Heat fixing so the sample doesn't move on the slide.
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What order do you press the valves of a graduated pipette?
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A creates a vacuum, S draws in the liquid, E release the liquid.
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How do you sterilize the rod used in the spread plate technique?
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You don't hold it in the flame: dip the rod in alcohol, pass it through the flame and let the alcohol burn off.
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In a lab report, where do table numbers and descriptive titles go? Figures?
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Tables: above the table, Figures: below the figure.
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What terms can you use to describe the consistency of a colony?
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smooth, dry, fibrous, mucoid, wrinkled
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What terms can you use to describe the form of a colony?
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circular, punctiform, irregular, filamentous, rhizoid
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What terms can you use to describe the elevation of a colony?
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flat, raised, convex, pulvinate (bubblish), umbonate (irregular lumps)
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What terms can you use to describe the margin of a colony?
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entire, undulate, lobulate, filamentous
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What equipment did you use to perform the direct cell count?
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Quebec colony counter
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What equipment did you use to measure turpidity?
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spectrophotometer
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What dyes are commonly used for simple staining?
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methylene blue, basic fuchsin, crystal violet
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What can you see in a simple stain?
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All cells & structures stain the same color; can see morphological types like cocci, rods, spirals & vibrios (curved rods)
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In positive staining procedures, the stain has a ____ly charged chromophore.
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positively charged - the stain is basic - sticks to slightly negative outer surface
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Methylene blue's chromophore is ___ charged.
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positively
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What are the lenses in a microscope?
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ocular and objective
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The 100X lens allows vhat kind of viewing?
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viewing under oil immersion: oil improves the resolution
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What is the usual magnifying power of the ocular lens?
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10X
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What is resolving power?
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The resolution is the ability to distinguish two images as separate.
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If the image in the microscope is dim or there is no light, what do you do?
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adjust the iris diaphragm and/or intensity of the light
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If the microscope is rocking, what do you do?
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right front rubber foot is adjustable
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If the image in the microscope is foggy, you should ...
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Remove traces of oil from the lens - use lens cleaner & lens tissue (not alcohol).
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A spirochete or spirillum is ...
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spiral-shaped
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Caulobacter has what form?
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stalked
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A comma-shaped organism is called
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vibrio
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Why might you see morphological variation?
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age, composition of media
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What types of cells (based on morphology) are mostly motile? Mostly nonmotile?
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Motile: Bacillus & spirillum; nonmotile: coccus
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What does strepto- mean?
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chain
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What shape of cell tends to produce spores?
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Bacillus
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Corynebacterium has what form?
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club-shaped
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What shape of bacteria typically exists singly?
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sprillum,
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How do you calculate the magnification of a specimen in a drawing?
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size of specimen in microm/estimated or actual size of image
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Gram stain is what kind of staining technique?
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differential
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Gram-positive organisms are ___ and gram-negative organisms appear ____ because ...?
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purple, red, because Gram +ve has thick peptidoglycan wall that retains the stain, Gram -ve shows the counterstain after decolorization by 95% ethyl alcohol
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What does the mordant do?
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fix the primary stain in cells
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Why might a Gram +ve organism appear mixed or Gram negative?
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old culture, acid medium
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What is the Kinyoun method for?
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acid-fast stain, a differential stain Mycobacterium - cold stain
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Why don't Mycobacterium stain readily by simple or Gram stain?
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waxy cell walls rich in mycolic acids (lipoidal material) - their walls are impermeable to most stains
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What's the difference between Ziehl-Neelsen & Kinyoun methods for the acid-fast stain?
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Z-N uses heat to drive the stain into walls, Kinyoun has higher concentrations of phenol and carbolfuchsin (primary stain)
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acid-fast organisms
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Are not decolorized by acid alcohol - all are Gram positive
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acid fast organisms appear ___ and non-acid-fast organisms appear __ because of?
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acid-fast is red (carbolfuchsin), nonacidfast blue (methylene blue counterstain)
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primary stain in the spore stain
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malachite green - driven into the cell with heat
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spore stain is what kind of stain
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differential
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spores appear __ because of? vegetative cells appear __ because of?
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green (malachite green); vegetative red (safranin)
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possible locations of endospores
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terminal, central, free
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stain used for negative stain
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acid dye nigrosin (india ink) - is repelled by the cell so shows it on a black background
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advantages of negative staining
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looking at cells that are hard to stain e.g. Mycobacterium, looking at capsules, no heating so doesn't distort cell shape or size
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capsule is made of ___. also called?
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polysaccharides, polypeptides &/or carbohydrates; also called slime layer or glycocalyx
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Capsules can be stained with __ because?
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Alcian blue because it's basic & forms linkages with acid groups in slime layer
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Why can't you use a simple stain to look at capsules?
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won't adhere to polysaccharides because they're water soluble and uncharged
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What does a capsule look like in a negative stain?
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unstained halo around cell
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primary constituent of agar
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galactan, complex carbohydrate from marine alga - solidifying agent
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when does agar melt? solidify?
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100 degrees C, stays liquid until cooled to 43 degres
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what's in nutrient broth
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beef extract & peptone in distlled water
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sterile
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free of all life, including viruses
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nutrient broth or agar is a ___ medium
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general purpose or all purpose medium
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the principal lethal agent in steam sterilization is
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heat
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instrument used to sterilize media & temperature thereof?
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autoclave, 121 degrees - acts like a pressure cooker - builds up steam pressure without boiling
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examples of selective agents
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antibiotics, bile salts, high [NaCl], dyes like crystal violet, specific sugars
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example of differential and selective media
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EMB agar
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Why is EMB agar selective?
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eosin Y and methylene blue dyes suppress gram-positive organisms
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Why is EMB agar differential?
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lactose as energy source, pH indicator
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What is EMB used to isolatE?
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enteric bacteria like E. coli, Enterobacter will grow on it, organisms that produce beta-galactosidase
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What does beta-galactosidase do
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break lactose down to glucose and galactose
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Why is KF Streptococcal agar selective?
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Sodium Azide is the selective agent, suppressing Gram-negative organisms.
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What does E cooli look like on EMB agar?
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blue/green, shiny metallic
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What does Enterobacter aerogenes look like on EMB agar
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large pinkish mucoid colonies with dark centers
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What's an organism that grows on KF streptococcal agar?
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streptococcus faecalis
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streak plate method is used for what
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isolation of pure cultures
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3 dilution methods commonly ised for isolation of bacteria
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streak plate, spread plate, pour plate
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standard plate count is used where in real world
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to determine number of organisms in water, milk & food
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direct count tells you what
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total number of cells dead & living
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Calculating CFU/mL culture
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# colonies & dilution factor divided by size of aliquot
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what does Brownian motion look like? True motility?
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organisms stay in one place but shake, true motility moves across slide usually zig-zagging
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Motility of lactobacillus plantarum
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not motile
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motility of pseudomonas fluorescens?
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motile
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West method
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increase thickness of flagella by coating with mordants like tannic acid then staining with silver nitrate - mordant incorporates stain into precipitate and increases contrast but mordant will precipitate on anything present
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Proteus vulgarous has what kind of flagella?
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peritrichous
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most microorganisms use what as a nitrogen source
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protein & amino acids, some use inorganic nitrogen compounds
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Lactobacillus plantarum can't grow without
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nitrogen source
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most microbes use what as energy sources
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carbs and amino acids
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urease activity is typical of __ & detected with ---?
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Proteus; phenol red pH indicator - dark pink for alkaline
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indole is a by-product of what? detected with what?
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tryptophan break-down, Kovac's agent
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what's in SIM medium? What's it for?
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IRON - detection of metal sulfides (turns dark) - shows organisms is metabolizing sulfur-containing aas
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how to detect carb metabolism
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Durham tube for gas, bromocresol purple for pH (- dark blue for alkaline, yellow for acid
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P aeruginosa does what in the carb fermentation test
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consumes proteins and makes alkaline - doesn't use the sugar
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Proteus vulgaris and B subtilis ferment what sugars
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glucose and sucrose not lactose
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E coli ferments what sugars
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glucose and lactose
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E coli does what in indole test
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positive -red ring forms
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Proteus vulgaris does what in the indole test
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is positive - red ring forms
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what organism is positive for hydrogen sulfide production
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proteus vulgaris
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examples of endoenzymes
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catalase & oxidase
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catalase does what reaction
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2 peroxides to 2 water and molecular oxygen
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oxidase does what
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activate oxidation of reduced cytochrome c by molecular oxygen during electron transport in aerobes
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detecting oxidase
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oxidized cytochrome c will tranfer oxygen to tetramethyl-p-phenylenediamine - makes purple/black color
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evidence of protease activity
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clear zone in milk as casein breaks down
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evidence of lipase activity
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cloudy or turbid precipitate from hydrolysis of Tween 80
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evidence of amylase activity
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iodine + starch is black, amylase makes a clear zone
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organisms that hydrolyzyes starch
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B subtilis
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What organism has lipolase
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P aeruginosa
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Temperatures of psychrophiles, mesophiles, thermophiles
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P: optimal <<15, max <20; M: 20-45, T: 45-80
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organisms with temperature sensitive pigment
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Serratia marcescens
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a thermophile
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Bacillus stearothermophilus
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Saccharomyces cerevisiae is a __ that likes ___pH.
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yeast, neutral to acidic
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organisms that can grow at 3 degrees C
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Lactobacillus plantarum
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microaerophiles
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require oxygen in limited amounts
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obligate anaerobes
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can't grow with oxygen present
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indifferents
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grow equally well with or without oxygen
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redox indicator
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resazurin - turns pink with oxygen -> in thioglycolate midium
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what kind of media favor anaerobes
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contains reducing compounds like cysteine and sodium thioglyclate
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an indifferent bacterium
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lactobacillus plantarum
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a strict anaerobe
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Clostridium species
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facultative anaerobe example
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E coli, Staphylococcus aureus
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thymine dimers form when
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cell exposed to UV
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thermal death time
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time required to kill cells or spores at given temperature
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thermal death point
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temperature at which organisms dies in 10 minutes
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a true halophile
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Halobacterium salinarium
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hypertonic solution
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high solute concentration - dehydrates cell membrane
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hypotonic
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low solute concentration - causes swelling but usually not harmful
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lethal mode of action for heavy metals
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inactivation of key enzymes
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oligodynamic action
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ability of small amounts of metal to kill bacteria
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antiseptic example
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Dettol
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disinfectant + example
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disinfectant kills microorganisms that cause disease -> Lysol, Iodine
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static activity
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activity that arrests growth but doesn't kill
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stuff used to kill things on inanimate surfaces
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disinfectants - lysol, dettol
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stuff used to kill things on animate surfaces
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antiseptics - iodine, scope
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agent causes disulfide bridges
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iodine
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agent inhibits enzymes
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dettol, lysol, iodine
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contains methylated phenols (cresols)
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lysol, dettol
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antibiotic
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chemical agent produced by organisms that control other organisms
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broad spectrum antibiotic examples
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tetracycline (prevents binding of tRNA to 70s ribosome), erythromycin (inhibits protein synth, targets 50s subunit of ribosome), some derivatives of penicillin
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agent effective against gram negative bacteria
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Streptomycin (attaches to ribosome, messes up translation), Polymyxin B
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antibiotic agent effective against gram positive bacteria
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penicillin (inhibits synth. of peptidoglycan)
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polymyxin B works by
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altering membrane permeability, bonds to phospholipids - will kill gram negative
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disc-plate procedure
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look for zones of inhibition around disc containing antibiotic or whatever agent
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examples of mutagens
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UV, X-ray, nitrous acid, alkylating agents, base analogues
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Medium suitable for fungus and yeasts
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Sabouraud's agar
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