MICRO 3060 MIDTERM QUESTIONS WITH VERIFIED
ANSWERS
Describe aseptic technique and understand why it is important. - Answers - -A set of
routine procedures done to prevent sterile solutions and cultures from becoming
contaminated by unwanted microorganisms in the lab.
-It is important because it prevents microbes from unintentionally released into the
environment and/or contaminating lab user in the lab.
1.Inoculating loops is flamed→ this incinerates any possible contaminating organisms.
*Make sure loop has cooled or else it will kill the bacteria.
2.Cap of tube is removed, the mouth of tube (ONLY GLASS) is flamed
3.Loopful of organisms is removed from tube
4.Mouth of tube is flamed again, and the top is put back on
5.Media is inoculated6.Loop is flamed again→ destroys any remaining bacteria
Define and describe culture media. - Answers - -Culture media is a gel or liquid that
contains nutrients and is used to grow bacteria or microorganisms.
-Nutrient broths and agar plates are the most typical growth media for microorganisms.
Compare and contrast how to make a bacterial smear from liquid media and solid
media. - Answers - -when inoculating a solid media. a sterile loopful of water is added to
the slide; also only a small amount of solid media is necessary or else there are too
many bacteria cells on the slide
-When inoculating liquid media water is not needed
Explain the importance of heat fixing and air-drying specimens on slides. - Answers -
heat fixing causes the cells to adhere to the slide so that the application of dye or water
doesnt wash them off, it kills the microorganisms and allows them to be stained better
by causing pores in the cell wall to open. **some bacteria cannot withstand high
temperatures and must be air dried
Compare and contrast simple staining and differential staining. - Answers - -Simple
staining involves one dye
-Differential staining involves multiple dyes to distinguish between different bacteria
Recognize the different bacterial morphologies. - Answers - -Bacillus: rod shaped
bacteria
-Coccobacillus: short rods
-Cocci: spherical bacteria
Compare and contrast gram-positive and gram-negative cells (what colors they stain
using the Gram stain and how cell wall structure determines how they stain). - Answers
- -Gram-positive: stain purple (crystal violet.) The thick peptidoglycan wall traps the
large Crystal violet + Iodine complexes within the cell wall.
, -Gram-negative: stain red (safranin.) The lack of peptidoglycan results in the large CVI
complexes being washed out by alcohol, they get their red color from the counter stain
that is applied at the end of the gram-staining procedure.
Know all of the steps and stains used in a Gram stain. Know the purpose of each stain
and how bacteria look at each step. Know which stain/reagent is the primary stain,
counterstain, mordant, and decolorizing agent. - Answers - 1. The heat fixed cells are
dried on the slide; all cells are colorless.
2. The PRIMARY STAIN, crystal violet, is applied to the sample and all cells appear
purple.
3. The slide is rinsed with water to remove an extra dye, no color change occurs.
4. The MORDANT, Gram's iodine, is applied to the slide. This bonds with the crystal
violent dye and forms large CVI complexes, all cells still appear purple.
5. The sample is rinsed with the DECOLORIZING AGENT, ethyl alcohol. This causes
the gram- cells to lose the large CVI complexes because they lack a thick peptidoglycan
wall to hold onto them. After this step the gram+ cells are still purple while the gram-
cells are now colorless.
6. The sample is rinsed with water to remove any remaining alcohol; no color change
occurs.
7. The final step involves applying the COUNTERSTAIN, safranin; this stains both the
gram+ and gram- cells. After this step the gram+ cells are now a dark purple color and
the gram- cells are a red/pink color.
Understand the purpose of a streak plate and how you would do one. - Answers - Used
to isolate a colony; the quadrant streak method is used in order to have isolated pure
colonies that can be studied
Pure culture - Answers - a culture containing only one species of bacteria
CFU (colony forming unit) - Answers - unit that is used in microbiology to estimate the
number of viable bacteria in a sample
Describe the growth of bacteria in various types of media (cultural characteristics) using
the proper microbiological terms. - Answers -
Know the purpose of a gelatin stab, how you conducted this test, and what a positive
reaction for gelatinase production (gelatin hydrolysis) would look like. - Answers - Some
bacteria produce PROTEASES (enzymes that degrade proteins), an inoculating needle
is stabbed into the gelatin, using the aseptic technique, and the tube is placed in an ice
bath after incubation. If any liquefaction of the gelatin medium occurs, it is a positive
reaction for the production of proteases.
Describe endospores, their function, and know which genera produce them. - Answers -
Endospores are dormant and highly resistant cells produced by BACILLUS AND
CLOSTRIDIA bacterial cells when they exhaust all essential nutrients in their
environment. The endospores allow the bacteria to survive in these unfavorable
ANSWERS
Describe aseptic technique and understand why it is important. - Answers - -A set of
routine procedures done to prevent sterile solutions and cultures from becoming
contaminated by unwanted microorganisms in the lab.
-It is important because it prevents microbes from unintentionally released into the
environment and/or contaminating lab user in the lab.
1.Inoculating loops is flamed→ this incinerates any possible contaminating organisms.
*Make sure loop has cooled or else it will kill the bacteria.
2.Cap of tube is removed, the mouth of tube (ONLY GLASS) is flamed
3.Loopful of organisms is removed from tube
4.Mouth of tube is flamed again, and the top is put back on
5.Media is inoculated6.Loop is flamed again→ destroys any remaining bacteria
Define and describe culture media. - Answers - -Culture media is a gel or liquid that
contains nutrients and is used to grow bacteria or microorganisms.
-Nutrient broths and agar plates are the most typical growth media for microorganisms.
Compare and contrast how to make a bacterial smear from liquid media and solid
media. - Answers - -when inoculating a solid media. a sterile loopful of water is added to
the slide; also only a small amount of solid media is necessary or else there are too
many bacteria cells on the slide
-When inoculating liquid media water is not needed
Explain the importance of heat fixing and air-drying specimens on slides. - Answers -
heat fixing causes the cells to adhere to the slide so that the application of dye or water
doesnt wash them off, it kills the microorganisms and allows them to be stained better
by causing pores in the cell wall to open. **some bacteria cannot withstand high
temperatures and must be air dried
Compare and contrast simple staining and differential staining. - Answers - -Simple
staining involves one dye
-Differential staining involves multiple dyes to distinguish between different bacteria
Recognize the different bacterial morphologies. - Answers - -Bacillus: rod shaped
bacteria
-Coccobacillus: short rods
-Cocci: spherical bacteria
Compare and contrast gram-positive and gram-negative cells (what colors they stain
using the Gram stain and how cell wall structure determines how they stain). - Answers
- -Gram-positive: stain purple (crystal violet.) The thick peptidoglycan wall traps the
large Crystal violet + Iodine complexes within the cell wall.
, -Gram-negative: stain red (safranin.) The lack of peptidoglycan results in the large CVI
complexes being washed out by alcohol, they get their red color from the counter stain
that is applied at the end of the gram-staining procedure.
Know all of the steps and stains used in a Gram stain. Know the purpose of each stain
and how bacteria look at each step. Know which stain/reagent is the primary stain,
counterstain, mordant, and decolorizing agent. - Answers - 1. The heat fixed cells are
dried on the slide; all cells are colorless.
2. The PRIMARY STAIN, crystal violet, is applied to the sample and all cells appear
purple.
3. The slide is rinsed with water to remove an extra dye, no color change occurs.
4. The MORDANT, Gram's iodine, is applied to the slide. This bonds with the crystal
violent dye and forms large CVI complexes, all cells still appear purple.
5. The sample is rinsed with the DECOLORIZING AGENT, ethyl alcohol. This causes
the gram- cells to lose the large CVI complexes because they lack a thick peptidoglycan
wall to hold onto them. After this step the gram+ cells are still purple while the gram-
cells are now colorless.
6. The sample is rinsed with water to remove any remaining alcohol; no color change
occurs.
7. The final step involves applying the COUNTERSTAIN, safranin; this stains both the
gram+ and gram- cells. After this step the gram+ cells are now a dark purple color and
the gram- cells are a red/pink color.
Understand the purpose of a streak plate and how you would do one. - Answers - Used
to isolate a colony; the quadrant streak method is used in order to have isolated pure
colonies that can be studied
Pure culture - Answers - a culture containing only one species of bacteria
CFU (colony forming unit) - Answers - unit that is used in microbiology to estimate the
number of viable bacteria in a sample
Describe the growth of bacteria in various types of media (cultural characteristics) using
the proper microbiological terms. - Answers -
Know the purpose of a gelatin stab, how you conducted this test, and what a positive
reaction for gelatinase production (gelatin hydrolysis) would look like. - Answers - Some
bacteria produce PROTEASES (enzymes that degrade proteins), an inoculating needle
is stabbed into the gelatin, using the aseptic technique, and the tube is placed in an ice
bath after incubation. If any liquefaction of the gelatin medium occurs, it is a positive
reaction for the production of proteases.
Describe endospores, their function, and know which genera produce them. - Answers -
Endospores are dormant and highly resistant cells produced by BACILLUS AND
CLOSTRIDIA bacterial cells when they exhaust all essential nutrients in their
environment. The endospores allow the bacteria to survive in these unfavorable
