Q1. Acid fast bacteria are have some significant differences as compared to non-acid fast bacteria. What are
the major chemical (structural) differences among these different bacteria that might explain the acid-fast
stain? Hint: This structural difference also affects the way the bacteria grow on media (both broth and media
containing agar) and the length of time they take to grow.
Q2. Why do you suppose the acid fast stain in not as widely used as the Gram stain? When is it more useful
than the Gram stain? Can you name the bacterial genera that are considered to be acid fast?
Q3. (Use the lab manual to answer this) How does heating the bacterial smear during a Ziehl-Neelson stain
promote entry of carbol fuchsin into the acid fast cell wall? Why is the Kinyoun procedure referred to as the
“cold” acid fast stain procedure? Explain.
Q4. Are acid fast negative bacteria stained by carbol fuchsin? If so, explain why the acid fast stain is
considered to be a differential stain?
Q1. What is the function of a bacterial endospore? Hint: How easy is a spore to destroy? Use your
textbook/lecture notes to help you.
Q2. Why does this exercise call for an older culture of Bacillus? What does a positive result for a spore stain
indicate about the organism? What does a negative result for the spore stain indicate about the
microorganism?
Q3. List three specific functions of capsules. Use your textbook/lecture notes to help you. (Hint: two of these
functions are often listed as functions of various virulence factors). Note: It is not enough to say that it protects
bacteria from the immune system or that it is a virulence factor.
Q4. In the capsule stain process used in class (using nigrosin as described in Ex. 3-3), are the bacterial cells
heat-fixed to the slide? Why or why not?
Q5. Why doesn’t a negative stain, which is also an acidic stain, such as the nigrosin colorize the cells in the
smear?
Q6. Eosin is an acidic red stain and methylene blue is a basic blue dye. What should be the result of staining a
smear of encapsulated bacteria with a mixture of eosin and methylene blue?
Please note if the capsule is stained or not stained at all.
Q1. How does complex medium compare to synthetic (chemically defined) media? What kind of medium were
you using in exercise 1-4 and exercises 2-2, etc.?
Q2. What is the meaning of the term colony forming unit? Why is it applicable when talking about the growth on
a streak plate?
Q3. Define or explain:
a) pure culture
b) obligate anaerobe
c) facultative anaerobe
d) aseptic technique (think about this in laboratory terms)
Q4. What is the purpose of inverting inoculated plates during incubation?
Q5. Match the following terms from exercise 2-3
_ filiform 1. Produces colored growth spreading edge 2. Smooth texture with solid edge _ transparent 3. Solid growth seeming to radiate outward
______friable 4. Almost invisible or easy to see light through
______pigmented 5. Rough texture with a crusty appearance
Q6. Match the following terms from exercise 2-4
_____Flocculent a. Evenly cloudy throughout
_____Sediment b. Growth at top around the edge
_____Ring c. Growth at the bottom
_____Pellicle d. Membrane at the top
_____Uniform fine turbidity e. Suspended chunks or pieces
Q1. What is the role of extracellular enzymes in bacterial metabolism?
Q2. How might the results obtained in these exercises be used for the identification and/or classification of
bacteria?
Q3.Suppose you had poured iodine on your plate and noticed clearings in the uninoculated area, as well as
around both of your transferred cultures? What are some possible explanations for this occurrence? Was the
integrity of the exercise compromised? What kinds of things might be done to avoid this problem in the future?
Q4.Is it acceptable to read a positive test before the incubation time is completed? Why is this not the case in a
starch agar assay? How about an early negative result?
Q5.Why is it advisable to use a positive control along with organisms that you are testing?
Q6.With respect to the Starch Agar assay, how would you expect the results to change if you were to add
glucose to the medium (in addition to the starch already present)?
Q1. All enterics are facultative anaerobes. What color results would you expect for organisms in O-F glucose
media inoculated with an enteric. Remember to describe both sealed and unsealed tubes. How would an
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obligate aerobe appear in both the sealed and unsealed tubes? What would happen if lactose were used
instead of glucose? Would you expect the results for E. coli and P. vulgaris to be identical? Explain based on
your knowledge from how these two bacteria grow on MacConkey media.
Q2. With respect to Exercise 5-2, what is the Durham tube and what is its purpose?
Q3. List two advantages of a multi-test system such as triple sugar iron agar or SIM.
Sample Solution
