Midterm Flashcards
Why is the body of the microscope never lowered while looking through the ocular lens?
This is to ensure that the objective lens and slide are not damaged by the forceful contact between the two \
What does the iris diaphragm do?
It adjusts the amount of light coming through the specimen
What does the coarse adjustment do?
It is used to bring the specimen into view
What does the fine adjustment do?
It is used to bring the specimen into sharp focus
What does the condenser do?
It directs the light from the light source into the lens system
What does the mechanical stage do?
It controls the position of the specimen over the central opening in the stage
What is the reason for the inability to bring the specimen into sharp focus?
- insufficient or an excess amount of oil on the slide
- failure to position the fine adjustment at the midpoint of its range prior to focusing with coarse adjustment
What is the fix for insufficient light going into the lens system?
Raising the Abbe condenser completely and adjusting the iris diaphragm
Why are thick and dense smears bad in staining?
- thick smears do not allow for sufficient light to pass through the preparation for good visualization of the organisms
- dense smears contain tightly packed and superimposed cells that do not lend themselves to accurate determination of cell shape and arrangement
Why do we let bacterial smears air dry?
Air drying prevents the cells from shrinkage and distortion and allows for the visualization of the natural cellular morphology
What does excessive heat do to bacterial smears?
It can distort the morphology, causing plasmolysis of the cell wall
If done improperly the smear can wash off of the slide
Why is basic dye preferred when staining?
Because the chromogen in the dye is cationic and has an affinity to negatively charged DNA. Most bacteria has a negatively charged cell surface
Why can Methylene blue not be used in negative staining?
Because it is a basic cationic dye. Only an acidic stain can be used
What is the point of negative staining?
It allows the visualization of living microbial cells that have not undergone distortion by heat fixation
How does differential staining work?
It utilizes two stains of contrasting colors that allow for the separation of bacteria into groups or for the visualization of cellular structures
What is the purpose of the primary stain?
To impart color to all cells
What is the mordant used for?
It is a chemical that acts as an intensifier in the Gram staining procedure
it forms a complex with the crystal violet which cannot easily be removed from gram positive cells
What is the counterstain used for?
It is the second stain applied at is absorbed by decolorized cells
Why is heat used in acid-fast staining?
to soften the waxy cell wall components to facilitate the penetration of the primary stain into the cells
Why is acid alcohol used in acid-fast staining vs. 95% ethyl alcohol?
Acid-alcohol is used preferentially over 95%
ethyl alcohol to ensure that the primary stain is
removed from the non–acid-fast organisms.
What is acid-fast staining used to diagnose?
The acid-fast staining procedure is used for the
diagnosis of leprosy and tuberculosis, both of
which are caused by members of the genus
Mycobacterium.
Why is heat not needed after counterstaining acid-fast stains?
Application of heat or a surface-active agent is
not required during the application of the counterstain.
The acid-fast organisms, because of the
waxy nature of their cell walls, are not decolorized,
and the red stain remains trapped inside
the cells. The non–acid-fast organisms lack the
lipoidal cell wall components. Therefore, the
primary stain is easily removed during decolorization,
and the colorless cells are readily stained
by the counterstain.
Why is the stain heated when dealing with spores?
to ensure penetration of the stain into the spore.
Why is water used to rinse in spore staining?
The function of water is to remove excess primary
stain from the spore. The vegetative cells
lack an affinity for this stain; thus it is removed
by water, rendering the vegetative cells colorless.
What color are gram positive bacteria?
blue
What color are gram negative bacteria?
red
Why do we not use acid-alcohol for staining in spores?
Because it has the same effect that water would have
What happens when you fail to apply heat with the primary stain?
The stain is unable to penetrate into the endospore
You used safranin as the primary stain and malachite green as the counterstain
If safranin is applied with heat, both the
endospore and the vegetative cell will accept
the stain and appear red in color. Tap water will
not remove the stain, and, therefore, malachite
green would not be accepted. Both the endospore
and the vegetative cell will be red.
Explain the medical significance of a capsule
presence
renders the cell resistant to the phagocytic
activities of WBCs, thereby increasing the virulence
of the organism.
Explain the function of copper sulfate in this procedure
The capsule is nonionic and as such will not bind
with the cationic primary stain, crystal violet. In
this method, copper sulfate is used rather than
water to wash out excess stain from the cell. During
this process, the copper sulfate is absorbed
into the capsule, giving it a light blue color in
contrast to the deep purple color of the cell.
What happens when enzymes are not present?
Without the action
of enzymes many of these reactions would
not take place at perceptible rates. All aspects of
cellular metabolism are catalyzed by enzymes.
This includes the digestion of large nutrient
molecules such as proteins, carbohydrates, and
lipids that are broken down into smaller molecules;
the production and transformation of energy;
and the synthesis of larger macromolecules
essential for the viability of the cell.
Why is milk agar a good growth environment?
Although milk is a sterile body fluid, microorganisms
gain entry during the milking process.
Many of these microorganisms contain enzymes
that degrade milk carbohydrates, proteins, and
lipids with the production of acid end products.
Organisms such as Lactobacillus and Streptococcus
spp. ferment lactose to lactic acid and
acetic acid, turning milk sour. They may produce
enough acid to curdle the milk protein,
forming a curd.
What is cellular respiration?
Cellular respiration is a biooxidative process
that occurs aerobically, with molecular oxygen
serving as the final electron acceptor, or anaerobically,
with an inorganic ion acting as a final
electron acceptor.
What is fermentation?
Fermentation is a biooxidation
that utilizes an organic compound as the
final electron acceptor.
How do organisms use pyruvic acid?
Microorganisms differ in their use of pyruvic
acid. Some organisms use the pyruvic acid as a
final electron acceptor, resulting in the formation
of acids, alcohols, and solvents. Other organisms
use this compound as a stepping stone
into the Krebs cycle for further ATP production.
How do strict anaerobes get energy?
The strict anaerobes utilize the Embden-
Meyerhof glycolytic pathway to pyruvic acid
with limited ATP production. The pyruvic acid
is then further metabolized through fermentative
pathways.
What cycle to Pseudomonas use for energy?
Pseudomonas species hydrolyze proteins to
amino acids that then enter the Krebs cycle for
generation of ATP.
What is the purpose of the TSI test?
The TSI test is designed for the rapid separation
and presumptive identification of enteric organisms.
What does the low concentration in TSI agar allow?
The lower concentration of glucose in the medium
allows for detection of the utilization of
this substrate only
What does TSI stand for?
Triple Sugar-Iron Agar
What is the purpose of phenol red PH indicator in TSI agar?
The purpose of the phenol red pH indicator is to
detect carbohydrate fermentation that is indicated
by a color change in the medium from orange-
red to yellow, which is caused by the
presence of acidic end products.
What is the purpose of thiosulfate?
substrate for hydrogen sulfide production
Why is the media incubated for a specific amount of time?
The limited length of the incubation period is
important to prevent the breakdown of proteins
in the medium, which would result in the formation
of end products that would obscure the
observation of the results.
What is the IMViC test used for?
The IMViC test is used for the identification of
enteric organisms, which include both pathogens
and nonpathogens.
What is the purpose of Kovac’s reagent?
Kovac’s reagent acts to extract indole from the
medium into the reagent layer. The indole then
forms a cherry-red complex with pdimethylaminobenzaldehyde.
Why does the media become alkaline when citrate utilized?
Citrate utilization produces oxaloacetic acid and
acetate, which are enzymatically converted to
CO2 and pyruvic acid. The CO2 combines with
sodium and water to form Na2CO3, an alkaline
product.
Why is E.coli positive for the methyl red test and E aerogenes negative?
Both E. coli and E. aerogenes produce acidic
end products during early incubation. The low
pH is maintained by E. coli, thereby producing
a positive methyl red test. On the other hand, E.
aerogenes converts the acids to acetylmethylcarbinol,
a nonacidic end product that elevates
the pH of the culture later in the incubation period.
Why is pyruvate not detected in the medium but indole is?
Pyruvic acid is a utilizable intracellular metabolite
and therefore is not excreted into the medium.
Indole is a waste product and can be detected
in the medium.
Why is Simmons citrate used to identify organisms?
The rationale for the use of Simmons citrate is
to identify organisms that are enzymatically capable
of metabolizing citrate as the sole carbon
source for energy production.
What are the substrates for hydrogen sulfide production?
The substrates for hydrogen sulfide production
include the amino acid cysteine and inorganic
sulfur-containing compounds such as thiosulfates,
sulfates, and sulfites.
Why is the ferrous ammonium sulfate important in the hydrogen sulfide test?
The ferrous ammonium sulfate serves as an
indicator by combining with the hydrogen sulfide
gas to form a detectable, insoluble, black
ferrous sulfide precipitate within the medium.
What is urease?
Urease is a hydrolytic enzyme that degrades
amide compounds such as urea with the formation
of ammonia, which is alkaline.
Why is phenol red incorporated in urea broth?
Phenol red is incorporated into the urea broth
for the detection of the alkaline end products
with the resultant development of a deep pink
color.
What is the difference between an acid curd and a rennet curd?
An acid curd is a nonretractable, hard clot,
whereas the rennet curd is a soft clot that retracts
from the walls of the tube
What happens during proteolysis?
In proteolysis, the proteins are degraded
to amino acids with the production of
ammonia, an alkaline end product. This is evidenced
by the appearance of a deep-purple band
at the surface of the culture. The medium below
shows a translucent brown color.
What happens in an alkaline reaction in litmus milk?
In an alkaline
reaction, the casein is partially degraded to
shorter polypeptides with the release of some
alkaline end products. As a result, the medium
assumes a deeper blue color.
What happens to litmus when it is reduced?
It becomes
reduced when it gains hydrogen ions and the
medium turns white starting at the bottom of the
test tube.
What is the purpose of the agar in nitrate reactions?
The agar in the nitrate medium serves to lower
the redox potential to favor the anaerobic requirement
for nitrate reduction.
