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Lab Quiz 2 Review

Lab Ex.'s 3,4,5,7

What is a smear? To stain bacteria, a thin film of bacterial cells must be placed on a slide to be viewed through a microscope.
Fixing Denatures bacterial enzymes, preventing them from digesting cell parts, which causes the cell to break(autolysis).
Heat fixing A slide is run through a Bunsen burner flame several times to dry the smear, does not kill the bacteria.
Chemically fix Cover the smear (with bacteria on it) with 95% methanol for 1 minute.
What is a chromophore? The ion that is colored. If chromophone is a (+) ion like methylene blue , the stain is considered a basic stain; if it is a (-) ion it is an acidic stain.
Simple stain Staining procedures that use only one stain. Can be used to determine cell morphology, size, and arrangement.
Direct stain A simple stain that stains the bacteria.
Negative stain A simple stain that stains the background but leaves the bacteria unstained.
Of what value is a simple stain? quick observation of the morphology and arrangement of bacteria.
What is the purpose of heat fixing the smear? To makes the bacteria sticky so they stick to the slide.
In heat fixing, what would happen if too much heat were applied? It would burn and/or kill the bacteria. Cells would be completely destroyed "plasmylosis" and burst and bacteria would not hold stain.
What bacterium is a rod? Baccillus
Which bacterium is larger? B. megaterium
Gram stain A differential stain that allows you to identify and classify bacteria as either gram- positive (+) or gram-negative (-).
Primary stain (1st step in gram staining) Crystal Violet (basic dye stains bacteria purple).
Mordant (2nd step in gram staining) Gram's Iodine (combines with crystal violet in cell to from a crystal violet-iodine complex (CV-I)).
Decolorizing agent Ethanol of ethanol-acetone (primary stain is washed out (decolorized) of some bacteria, whereas others are unaffected.
Secondary Stain or Counter Stain Safranin (a basic dye which stains the decolorized bacteria red).
Gram-negative(-) Those that decolorized easily
Gram-positive(+) Those that decolorize slowly and retain the primary stain.
Peptidoglycans Bacteria cell walls are composed of this complex lattice structure along with other compounds. Gram(+) cell walls contain multiple layers of peptidoglycans, whereas gram(-) bacteria contain a thin layer of peptidoglycan.
Why will gram (+) cells more than 24 hours old stain gram (-)? They may be old cells that have a deteriorated cell wall, as cell age there cell walls degrade which may nor allow bacteria to retain stain and give inaccurate results.
Can iodine be added before the primary stain (crystal violet) in a Gram stain? No
Suppose you performed a Gram stain on a sample from a pure culture of bacteria & observed a field of red and purple cocci. Adjacent cells were not always the same color. What do you conclude? That some were a successful stain & the others may have been dead or just didn't retain the stain.
Suppose you are viewing a Gram-stained field of red rods and purple cocci through the microscope. What do you conclude? A mixed culture OR contamination has occurred.
Considering you can't identify bacteria from a Gram stain, why might a physician perform a Gram stain on a sample before prescribing an antibiotic? Antibiotic sensitivity correlates with the cell wall type of the bacteria.
If you performed a Gram stain on human cells, what would happen? NO cell walls, the cells would not hold the stain, ETOH (alcohol) would destroy the membranes (phospholipid bilayer).
Chemically defined medium A medium whose exact chemical composition is known.
Complex media Media for which the exact chemical composition varies from batch to batch.
Nutrient broth Is a commonly used liquid complex medium.
Nutrient agar When agar is added to the nutrient broth it becomes a solid medium.
Agar An extract from marine red algae, has some unique properties that make it useful in culture media. Few microbes can degrade agar, so it remains solid during microbial growth.
Steam sterilization OR autoclaving Which uses steam under pressure( during this process, material to be sterilized is placed in auto-clave and heated to 120 C at 15 lbs. of pressure for 15 minutes. It's the most common method of sterilizing culture media that are heat stable.
Petri plates Contain solid media which provide a large surface area for examination of colonies.
Inoculated Intentionally introduced, into various forms of culture media to keep them alive and to study their growth-without introducing unwanted microbes.
Incubation period Bacteria that are inoculated into culture media increase in number during this period. A period of usually 24-48 hours were bacteria are heated a 98.6 degrees Fahrenheit to allow microbial growth.
Turbid After a suitable incubation, liquid media become cloudy as a result of bacterial growth.
Colony A population of cells that arise from a single bacterial cell.
Colony-forming unit When a colony arises from a group of the same microbes attached to one another.
What is the minimum number of different bacteria present on one of your plates? How do you know? > or equal to 1. > or equal to 1 colony type seen.
What is the value of Petri plates in microbiology? Have a larger surface area, easier to subculture so that you can see the isolated colonies of bacteria.
What are bacteria using for nutrients in the nutrient agar? Peptones and beef extract
What is the purpose of the agar? A solidifying agent ( solid/semi-solid surface to grow bacteria).
Why is agar preferable to gelatin as a solidifying agent in culture media? It stays solid at incubator temp of 98.6 degrees Fahrenheit and most bacteria can't eat or digest it.
Did all the organisms living in or on the environments sampled grow on your nutrient agar? No Way! We only provided one food type and one growing temperature.
Contaminants Unwanted microbes
Aseptic technique Used in microbiology to exclude contaminants and is performed under sterile conditions.
Sterilized Rendered free of all life, prior to use, usually accomplished during autoclave.
Broth cultures Provide large numbers of bacteria in a small space and are easily transported.
Agar slants Test tubes containing solid culture media that were left at an angle while the agar solidified. Provide solid growth surface but are easier to store and transport than Petri plates.
Inoculating loop and Inoculating needle When end of wire is bent into a loop AND when it's straight.(Aseptic transfer and inoculation are usually performed with this sterile, heat-resistant, noncorroding Nichrome wire attached to an insulated handle).
Agar deep Solidifies at the bottom of the test tube, often used to grow bacteria that require less oxygen than is present on the surface of the medium.
Were any of the bacteria growing in the semisolid agar deeps motile? Yes, resembled an upside down Christmas/pine tree. (Motile bacteria will move away from the point of inoculation, giving the appearance of an inverted pine tree)
What is the primary use of slants? Solid surface,easier to store than plates and not easily contaminated.
What is the primary use of deeps? Motility and oxygen requirements.
What is the primary use of broths? To maximize growth (the rate/amount)
What is the purpose of flaming the inoculating loop before use? After use? Before for sterilization to minimize contaminants. After for the safety of people within the lab.
Why must the loop be cool before you touch it to a culture? Should you set it down to let it cool? So you don;t burn and or kill the bacteria your trying to obtain. No because you risk contaminating it again and will have to re-heat the loop, always keep in your hands when in use.
Why is aseptic technique important? It minimizes the transfer of bacteria and contaminants from one thing to another.
Created by: KJones040607