Microbiology Lab Exam Flashcards Table View

     

Aseptic Technique This is the use of specific methods and sterile materials to exclude contaminating microorganisms from an environment.
Autoclave This is a device employing steam under pressure used for sterilizing materials that are stable to heat and moisture. (15 psi, 121 C for 15 minutes.)
Broth Liquid media used to grow bacteria.
Colony A visible mass of bacterial cells arising from a single cell.
Sterilization
The process of removing or destroying all microorganisms and viruses on or in a product.
Transfer Loop
Thin wire with a loop on the end used in transferring bacteria to a culture medium.
Transfer Needle
Thin straight wire used for stabbing agar (to get culture away from oxygen), or for fine manipulations
Mixed Colony
Population of organisms descending from more than one kind of cell
Isolated Colony
These are colonies separated from other colonies on a petri dish.
Pure Culture This is a population of organisms descending from a single cell.
Streak Plate
The simplest and most commonly used technique for isolating bacteria, a series of successive streak patterns is used to sequentially dilute an inoculum on the surface of an agar plate (non-quantitative dilution.)
Emulsify Mix culture in a drop of water to give a smooth, even suspension (used for making smears from solid media; i.e. plate or slant)
Heat Fix
This technique helps to retain the cells on the slide throughout the staining process.
Simple Stain This is a staining technique that employs one basic dye to impart color to cells.
Positive Stain This is the technique in which the cell is stained but the background remains unstained. Examples: Gram Stain, Spore Stain....
Negative Stain
Staining technique in which the background is stained and the structure of interest is not stained. This staining technique may employ an acidic dye to stain the background against which colorless cells can be seen (Example: Nigrosin negative stain.) In the Anthony’s Capsule stain, the basic dye Crystal Violet is used to stain the background and the cell allowing the clear capsule to be observed.
Differential Stain This is a type of staining procedure used to distinguish one group of bacteria from another by taking advantage of the fact that certain bacteria have distinctly different chemical structures in some of their components
Primary Stain
This is the first dye applied in a multi – step differential staining procedure; generally stains all cells.
Mordant This is the substance that increases the affinity of cellular components for a dye
Decolorizer Selectively removes the primary dye from one type of cell or cell structure.
Counter Stain The stain applied to impart a contrasting color to bacteria that do not retain the primary stain.
Endospore Stain This is a differential stain used to detect the presence and location of spores in bacterial cells.
Gram Stain This is used to distinguish between Gram positive and Gram negative cells. This is probably the most important and widely used microbiological differential stain.
Vegetative Cell
A typical, actively growing cell.
Endospore This is a kind of resting cell, characteristic of a limited number of bacterial species (Bacillus, Clostridium), highly resistant to heat, radiation, and disinfectants.
Sporogenesis (Sporulation)
This is in bacteria, a complex, highly ordered sequence of morphological changes during which a bacterial vegetative cell produces a specialized cell greatly resistant to environmental adversity
Germination This is the sum total of the biochemical and morphological changes that an endospore or other resting cell undergoes before becoming a vegetative cell.
Terminal Endospore
A endospore that is located at the end of the cell.
Central Endospore
Central Endospore –A. This is located in the middle of the cell.
Free Spore
A spore that is free of the mother cell after degeneration.
Contaminant A unwanted microorganism.
Lowenstein-Jensen Medium
This is used for the isolation of mycobacterium. Egg supplement enhances the growth of Mycobacterium and malachite green prevents growth of the majority of contaminants.
Mycolic Acids
The waxy substance that is found in cell walls of acid-fast bacteria.
Tubercle A granuloma that is formed in tuberculosis.
Litmus Milk
This is an undefined medium consisting of skim milk and azolitman. Skim milk provides nutrients for growth with lactose as the carbohydrate source and casein as the primary protein source.
Selective Media
This is a culture medium that inhibits the growth of certain microorganisms and therefore favors the growth of desired microorganisms.
Differential Media
This is culture media that allows visual identification of certain types of microorganisms.
Synthetic (Defined) Media
A medium in which the exact chemical composition and quantity of every component is known.
Complex Media A medium that is used to grow bacteria that has some ingredients of uknown chemical composition
All Purpose Growth Media
Complex media that is routinely used to culture bacteria. Usually inexpensive and easy to prepare (Example:TSA).
Enrichment Media
This media is used to isolate organisms which:

Uses compounds or vitamins which have been added to the media. Example: Blood Agar
or
Can grow without certain compounds which have been omitted from the media. Example: Citrus Citrate Media

Antibiotic Sensitivity (Kirby-Bauer Method)
This is testing used to determine the susceptibility of bacteria to various antibiotics. This is typically used to measure the effectiveness of a variety of antibiotics on specific organisms in order to prescribe the most suitable antibiotic therapy.
Why do you avoid turning the petri dish lid upside down when you remove the lid from the petri dish?
This will keep other microbes from contaminating the media.
Why do you incubate the plate inverted? This keeps water from running onto the culture and contaminating our culture
What is the Gram reaction of dead cells? The cells will appear to be gram negative because the degrading cell wall cannot retain the primary stain and it will appear to be red.
Is the endospore stain a differential stain? If so what is the decolorizing agent? A. Yes.
B. Water is used as the decolorizing agent.
Is the acid fast stain a differential stain? If so what is the decolorizing agent? A. Yes.
B. Acetone Alcohol.
Is the capsule stain a differential stain? If so what is the decolorizing agent? A. Yes.
B. Copper Sulfate
What are the two most obvious visual observations that indicate that you do not have a pure culture? A. The culture looks different than the other organisms. (Different Morphology.)
B. The Culture will not be on the streak line.
On an isolation streak plate, what might account for differences in colony size of a pure culture? A. Availability of nutrients.
B. Time of incubation.
In an acid-fast stain, what color are non – acid fast organisms? Blue (Ziehl-Neelsen) or green (Kenyon)
The acid fast stain is important in identifying pathogenic microorganisms, name 2 acid fast pathogens and briefly describe them and the diseases they cause? A. Tuberculosis – (Mycobacterium tuberculosis) Two manifestation of this disease include primary and secondary. Primary stage in a healthy person causes mild flu like symptoms. Necrotic lung inflammation occurs in the secondary stage; secondary is usually associated with a weakened immune system.

B. Mycobacterium leprae – Causes Leprosy, characterized by granulomas on face & extremities; loss of nerve function.
What role do capsules play in the disease process Capsules protect bacteria from phagocytosis
Are there any water wash steps in the Anthony’s capsule stain? Explain? A. No.
B. This is because the stain is water soluble, and because some slime layers/capsules are water soluble.
Why do you report sensitivity to an antibiotic instead of just the size of the zone of inhibition? Migration through the agar is dependent on chemical characteristics (molecular weight; hydrophobic/hydrophilic nature) and must be related to clinical effectiveness.
How does a hydrophobic antibiotic influence the size of the zone of inhibition?
This antibiotic will produce a smaller zone of inhibition.
How does a hydrophilic antibiotic influence the size of the zone of inhibition? This antibiotic will diffuse easily in the agar medium and will give a large zone of inhibition.
How does a antibiotic with a small molecular weight influence the size of the zone of inhibition? Antibiotics will diffuse further in a small molecular weight than a large molecular weight, producing a larger zone of inhibition.
How does a antibiotic with a large molecular weight influence the size of the zone of inhibition? Antibiotic will diffuse less than a small molecular weight, producing a small zone of inhibition.
What is the purpose of a simple stain? To detect the presence of bacteria and the morphology of bacteria.
What type of dye is used in a simple stain? Any basic dye
What mordant is used in a simple stain? None

What decolorizing agent is used in a simple stain? None
What is the counter(secondary) stain in a simple stain? None
What color are the cells after a simple stain is performed? All the cells are the color of the primary stain
What is the gram stain? One of the most important stains used in classification.
What color are the cells in a gram positive result? Purple/Blue
What is the color of the cells in a gram negative bacteria? Pink/Red


What is the primary dye in a gram stain? Crystal Violet
What is the mordant used in a gram stain? Iodine
What is the purpose of iodine in a gram stain? It is a mordant that increases the binding of the primary dye in the gram stain
What is the decolorizing agent in a gram stain? Acetone Alcohol
What is the counter(secondary) stain in a gram stain? Safranin

What is the most important step in a gram stain? The decolorizing step
What is the purpose of a spore stain? To detect endospores
What is the primary dye used in a spore stain? Malachite green.

What mordant is used in a spore stain? None
What decolorizing agent is used in a spore stain? Water
What counter(secondary) dye is used in a spore stain? Safranin


What color are the cells and endospores after a spore stain is performed?

The cells are red while the spores are green
What are two important steps in a spore stain? 1. The cells must be heat fixed to the slide
2. The primary stain (malachite green) must be steamed for 10-15 minutes in order to drive the stain into the spore

What are spores located outside of cells called? Free spores

What is the purpose of acid fast stains? To detect acid-fast cells such as mycobacterium tuberculosis
What is the primary stain in a acid-fast stain? Carbol Fushsion
What must be done in an acid-fast stain to the primary dye? The carbol fushion must be steamed for 10 minutes
What is the mordant used in an acid-fast stain? None

What decolorizing agent is used in a acid-fast stain? An acidic alcohol, which removes the dye from the non-acid fast cells.
What counter stain is used in a Ziehl neelsen acid-fast stain? Methylene Blue


What counter stain is used in a kinyoun acid-fast stain? Brilliant Green
What step is sensitive in the acid-fast stain? The decolorizing step
What makes acid-fast cells difficult to stain? Their waxy cell walls
What is the purpose of a nigrosin negative stain?

To determine the morphology and cellular arrangement of bacteria that are too delicate to withstand heatfixing
What is the primary dye in a negative stain? Nigrosin

What mordant is used in a negative stain? None
What decolorizing agent is used in a negative stain? None
What counter stain is used in a negative stain? None


What is the most important concept of a negative stain? Do not heat fix the specimen
What is the purpose of Anthony's capsule stain? To identify capsules
What is the primary dye used in a capsule stain?

Crystal Violet
What mordant is used in a capsule stain? None

What decolorizing agent is used in a capsule stain? Copper Sulfate
What counter stain is used in a capsule stain? None
What color are the cells and capsules after a capsule stain? The cells are purple while the capsules are clear


What accounts for the background staining in a capsule stain? A litmus milk protein
What is a important concept with a capsule stain?

The cells are not heat fixed.
What is the purpose of a purple broth and phenol red broth? To detect the ability of a bacterium to ferment a specific carbohyrate.

What are the possible results of a purple broth and what do they indicate? Yellow = pH < 6.8
Purple = pH > 6.8

What are the possible results of a PR broth and what do they indicate?
Yellow = pH < 6.8
Red = pH 6.8 - 7.4
Pink = pH > 7.4
What are the principles behind a purple and phenol red broth? Fermentation yields acid products = Yellow
Deamination of amino acids yields NH3 = Pink
A pink result in a PR broth indicates what? Deamination



A yellow result with bubbles in the durham tube indicates what?
Fermentation with gas end products
A yellow result outside urham tube, but purple inside the durham tube indicates what? Acid Produced Aerobically


When must you read the results of PB/PR broth test? Within 18 hours of inoculation
What is the function of a bile esculin agar? To select and differentiate among Group D Streptococci
What are two important ingredients in a bile esculin agar and what do they inhibit? Bile - Inhibits most G+ other than group D streptococci
Sodium Azie - Inhibits G-
What does the bile esculin agar test for?

The ability to hydrolyze esculin in the presence of bile
How do you interpret the results of a bile esculin test? Positive Result = more than half the tube has turned black or brown within 48 hours
How should the tubes in a bile esculin agar be inoculated? They should be streaked AND stabbed.


What is the function of litmus milk test?
To differentiate among the enterobacteriaceae based on the ability to ferment lactose and/or reduce litmus
In a litmus milk test, what are the possible results? Lactose Fermentation = Pink with cracks in the clot
Casein Coagulation = Pink with solid clots
Deamination = Blue
In a litmus milk test, what indicates reduction or oxidized ilitmus? If the bottom of the tube is...
Purple = Litmus Oxidized
White = Reduced litmus
What is the function of nitrate reduction test?


To detect the ability to reduce nitrate to nitrite
In step 1 of nitrate reduction, what indicates a positive result?
Gas and a non-fermenting organism
In step 2 of nitrate reduction, what is added? Sulfanilic acid and a-naphthylamine
In step 2 of nitrate reduction, what indicates a positive result? If the tube turns red
In step 3 of nitrate reduction, what is added? Powdered zinc
In step 3 of nitrate reduction, what indicates a positive result? Positive = No red color
Negative = Red Color
What is the function of a SIM Media?

To simultaneously test for sulfur reduction, indole production, and motility.

What is SIM Media used to differentiate among?
Enterobacteriaceae
What are the possible test results in sulfur reduction? Positive = Black Precipitate
Negative = None
What does the indole production test test for? The enzyme tryptophanase, which converts tryptophan to indole
What indicates a positive result in indole production?

A red color in the alcohol(top) layer.
What does the methyl red(MR) test test for? Glucose fermentation to produce large amounts of stable acids.
What are the possible results of a methyl red test? Red = Positive for acid production
Yellow = Negative for stable acids
Orange = Inconclusive
What does the Vogues-proskauer (VP) test test for? 2,3-butanediol and acetoin production
What are the possible result of a VP test?



Red = Positive
Brown = Negative
What is the function of Simmons citrate agar? To differentiate among the enterobacteriaceae based on the ability to utilize citrate as a sole carbon source.
What are the possible results of a Simmons citrate test?

Blue = Positive (Growth)
Green = Negative (No Growth)
What tests are standard for differentiating among the enterobacteriaceae?

IMViC
Indole, Methyl Red, Voges-Proskauer, Citrate Utilization
What is the function of gelatinase test? To determine if the bacterium is able to hydrolyze gelatin.
What is very important when performing a gelatinase test? After incubation, the tubes must be maintained and examined cold!
What are the possible results of a gelatinase test? If the media is liquid, then the result = positive
If the media is solid, then the result = negative
What does the urease test test for?


The ability to hydrolyze urea
What are the possible results for a urease test? Positive = All pink within 24 hours (rapid hydrolysis)
Weak Positive = Partially pink between 1-6 days (slow hydrolysis)
Negative = Orange or yellow after 6 days (no hydrolysis)
What does starch hydrolysis test for? The ability of an organism to hydrolyze starch
What are the possible results for starch hydrolysis? Positive = Clearing around the growth
Negative = No clearing