Microbiology Lab Final Exam Review

Urease Test

Detects microbes that are capable of breaking down urea into ammonia and carbon dioxide with the help of the enzyme urease.

Enteric

The enzyme urease is uncommon in ( ) or intestinal bacteria.

Don’t

Bacterial genera that (do/don’t) exhibit urease activity include salmonella, shigella, and escherichia.

Exhibit

Two species that notably ( ) urease activity are Proteus mirabilis and Helicobacter pylori.

Alkaline/Pink/Red/Positive

In the urease test, when urea is hydrolyzed by urease, ammonia is released and causes the medium to become ( ) and increase the pH. This causes the medium to turn into a deep ( ) to ( ) color, indicating a ( ) test for urease.

Yellow

A ( ) color to the medium indicates a negative urease test result.

Positive

In the Urea/Indole Wee-Tab test, a ( ) Indole test is indicated by the development of a cherry red color in the top layer of the medium.

Triple Sugar Iron (TSI) Test

Utilizes a differential medium that can assess the ability of a microbe to ferment sucrose, glucose, and lactose as well as produce gas and/or hydrogen sulfide. It is mainly used to identify enteric bacteria.

Ten/Hydrogen sulfide/phenol red

TSI medium contains sucrose and lactose as well as glucose in a ( )-fold lower concentration. It also contains ferrous ammonium sulfate, which reveals ( ) production, and ( ) that indicates carbohydrate fermentation.

Aerobic/anaerobic

In the TSI test, ( ) respiration occurs on the agar slant surface while ( ) respiration or fermentation occurs in the agar butt.

Fermentation of the low level of glucose present in the medium

What causes the butt of a TSI slant to be yellow?

No fermentation of the low level of glucose present in the medium

What causes the butt of a TSI slant to be red or pink in color?

Fermentation of sucrose and/or lactose in the medium

What causes the TSI agar surface to turn yellow?

No fermentation of the lactose or sucrose in the medium

What causes the TSI agar surface to turn red or pink?

Bubbles/cracks

Gas production in the TSI test causes ( ) or ( ) to appear in the medium.

Black/sulfur

A ( ) precipitate is produced and indicates hydrogen sulfide production during the TSI test when ( ) is released during protein degradation and combines with the ferrous ammonium sulfate in the medium.

Decarboxylation Test

Detects the use of decarboxylase and dihydrolases, a group of substrate-specific enzymes that remove the carboxyl and amino portions of amino acids. It is mainly used to identify enteric bacilli.

Acidic

In the decarboxylation test, the production of the enzymes responsible for degrading amino acids is induced only in an ( ) environment, but the byproducts of their reactions are alkaline and thus restore the alkaline nature of the medium.

Pyridoxal/glucose

The basal media for the decarboxylation test contains ( ), which is an important cofactor for decarboxylase activity, ( ), and bromocresol purple and cresol red. Arginine, lysine, or ornithine is added to the medium to detect the production of specific amino acid enzymes.

Purple

In the decarboxylation test, the ( ) color indicates alkaline conditions.

To lock out oxygen and prevent false alkalinization at the surface of the medium, promoting fermentation

Why is mineral oil added to each tube in the decarboxylation test?

Acidic/yellow

In the decarboxylation test, if glucose is fermented by the microbe in the tube, an ( ) state will develop and turn the medium ( ). This acidic environment will induce production of the enzymes responsible for breaking down amino acids, so long as the microbe is genetically equipped to do so.

Alkaline/purple

In the decarboxylation test, once enzymes are expressed and reach the amino acids in the medium, ( ) by-products are released and the medium turns back to ( ).

Acidic/yellow

In the decarboxylation test, if a glucose-fermenting bacteria does not produce the appropriate enzyme, the medium will remain ( ) and ( ) in color.

Weak

In the decarboxylation test, non-glucose-fermenting bacteria that also possess enzymes may display ( ) decarboxylase activity, causing a small change in color to the medium.

Same

In the decarboxylation test, non-glucose-fermenting bacteria that don’t possess the enzymes needed will keep the medium nearly the ( ) color.

Phenylalanine Deamination Test

Detects the microbial use of phenylalanine deaminase (PAD), which is an enzyme that removes an amino group from phenylalanine and produces phenylpyruvic acid, water, and ammonium ions.

Enteric

( ) bacteria do not produce phenylalanine deaminase, while species of Morganella, Providencia, and Protesus do.

Ferric Chloride/Positive

Production of phenylalanine deaminase (PAD) is detected by combination with ( ), which forms a green complex. This indicates a ( ) PAD test.

Motility Test

Detects whether a species contains flagella based on its movement through a medium.

TTC/red/motile/non-motile

Incorporation of triphenyltetrazolium chloride ( ) into the semi-solid medium for a motility test produces a ( ) pigment when reduced by bacteria. After stabbing bacteria through the media, two outcomes can be observed.

(1) A ( ) organism will produce a pinkish-red color that diffuses from the stab line

(2) A ( ) microbe will exhibit a pinkish-red pigment confined to the stab line

Hemolysis Test

Determines whether bacteria are capable of lysing red blood cells (RBCs). It is commonly used to identify pathogenic cocci.

Alpha-Hemolytic

Bacteria are described as this when they only partially lyse red blood cells and generate a zone of greenish coloration around colonies. This greenish tint is produced from the oxidation of iron released by lysed red blood cells.

Beta-Hemolytic

Bacteria are described as this when they completely lyse red blood cells, forming a clear zone surrounding colonies.

Gamma-Hemolytic

Bacteria are described as this when they don’t digest red blood cells and thus don’t produce clearing of any type around colonies.

Oxidase Test

Detects the activity of cytochrome oxidase or indophenol oxidase with the use of an oxidase swab containing TMPD, which reduces these electron carriers.

Dark purple/red

In the oxidase test, if cytochrome c oxidase or indophenol oxidase is present in the organism, it will be reduced by TMPD, forming a product that is ( ) to ( ) in color within 30 seconds of exposure.

Positive/negative

In cocci, Micrococcus, Neiserria, and Moraxella are oxidase ( ), while most species of Staphylococcus, Streptococcus, and Enterococcus are oxidase ( ).

Negative/positive

Pseudomonas are oxidase ( ) bacillus, while enterobacteria are generally oxidase ( ).

Gelatin

A soluble mixture of polypeptides derived from the collagen of animal connective tissue.

Gelatinase Test

Detects the presence of gelatinase, which hydrolyzes the polypeptides of gelatin, prohibiting its solidification at low temperatures. Because of gelatinase, the amino acids released by its hydrolysis of gelatin are a nutritional source for the microbe. Gelatinase also aids in the microbe’s pathogenicity because it prevents solidification and allows the microbe to spread throughout the body.

DNAase Test

Detects a microbe’s use of DNAase, an enzyme that degrades DNA. It uses media that incorporates DNA along with dyes like methyl green or toluidine blue sometimes.

Acidity

In a DNAase test with dyes, the blue-green DNA complex is disrupted if a microbe grown on the plate hydrolyzes DNA. This will cause a change in color around the bacterial growth due to the change in ( ).

Colorless/light pink

If methyl green were used for the DNAase test, the zone around the bacterial colonies that hydrolyze DNA will be ( ). If toluidine blue medium is used instead, a ( ) zone will form as the medium becomes LESS acidic.

Hydrochloric acid

In a DNAase test where dyes are not used, the addition of ( ) is used to dissolve any oligonucleotides in the medium into DNA salts that are insoluble. This means that ( ) will cause unhydrolyzed DNA to precipitate, making the medium opaque while DNAase makes the medium around DNAase-containing bacteria clear.

Positive

These are examples of DNAase ( ) bacteria

1) Staphylococcus aereus

2) Streptococci

3) Serratia

4) Moraxella

Negative

These are examples of DNAase ( ) bacteria

1) Enterobacter

2) Neiserria

3) Escherichia coli

4) Klebsiella

Coagulase Test

Detects the presence of coagulase, a type of enzyme that binds to fibrinogen. Fibrinogen is a soluble blood protein involved in the clotting of blood, so coagulase thus promotes binding of pathogenic bacteria to blood clots and damaged tissues. Coagulase can also convert fibrinogen to fibrin, which coats bacteria and protects it from the immune system.

S.aereus

The presence of coagulase is only associated with ( ) bacteria.

Plasma/clots

The reagent used in the coagulase test contains rabbit ( ), which tests for the presence of coagulase. In a positive test, the reagent ( ) or gels up.

Dismutase/catalase

As a result of metabolism, toxic byproducts like hydrogen peroxide and superoxide ions are produced. Superoxide ( ) converts superoxide ions into hydrogen peroxide, which is then broken down into water and molecular oxygen by ( ) or peroxidase.

Catalase Test

Detects the use of catalase and its function in bacteria by adding hydrogen peroxide directly to a sample of bacteria.

Positive/negative

In the catalase test, if bubbles of oxygen are released, this indicates a ( ) catalase test result. The absence of any bubbles indicates a ( ) result.

Staphylococcus/Micrococcus

Though Streptococcus, Enterococcus, Staphylococcus, and Micrococcus are morphologically similar, only ( ) and ( ) are catalase positive.

Positive

Among Clostridium and Bacillus species, Bacillus is catalase ( ).

Esculin/PYR Test

Detects the use of esculinase by bacterial species. Esculinase hydrolyzes esculin and its derivatives or similar compounds into several different byproducts. It is mainly used to distinguish between Staphylococcus, Streptococcus, and Enterococcus bacteria.

Positive/yellow

A ( ) Esculin test before addition of PEP will turn the medium ( ).

B-naphthylamine/Positive/Negative

The Esculin/PYR test also detects bacteria who hydrolyze PYR into ( ) with the help of the enzyme pyrolidonyl arylamidase. Adding PEP produces a color change that indicates whether PYR was hydrolyzed. A ( ) test will turn the medium red or pink while a ( ) test will keep the medium yellow in color.

45/NaCl

Enterococcus, Staphylococcus, and Streptococcus all differ in their ability to grow at ( ) degrees C and in the presence or absence of 6.5% ( ).

Blacken

Group D streptococci like Enterococci, S. bovis, and S, gallolyticus will ( ) the BEA agar, while non-group D streptococci will not.

Ferric citrate/positive

BEA agar contains esculin, ( ), bile, and azide. Group D streptococci while hydrolyze esculin to form dextrose and esculetin. Esculetin will react with the ferric ions in the medium and turn the medium black, which is a ( ) result.

Axide/bile

BEA agar is a selective medium because ( ) in the medium prevents the growth of gram-negative bacilli. The presence of ( ) also inhibits the growth of non-group D bacteria.

Citrate Utilization Test

Detects the use of citrate by Gram-negative bacilli. It uses medium containing ammonium dihydrogen phosphate as a nitrogen source and sodium citrate as a carbon source as well as bromothymol blue as a pH indicator.

Alkaline/blue

In the citrate test, when citrate is metabolized, an ( ) carbonate is produced, turning the pH indicator in the medium from green to ( ). This is a positive result.

Methyl Red-Voges Proskauer (MR-VP) Test

Differentiates between the two types of fermentation products produced by two kinds of enteric bacteria called “mixed acid” fermenters.

Methyl red/Positive/Yellow

The MR-VP test uses ( ) as a pH indicator, which under acidic conditions appears red, unlike in most other tests that use phenol red as the pH indicator. In a ( ) test, the medium would remain red if added to a culture of mixed acid fermenters. A non-mixed-acid fermenter would turn the medium ( ).

Negative

Typically, if a species is MR positive, it will be VP ( ). And vice versa.

VP/A/B/Red

Non-mixed-acid fermenters are detected using the Voges-Proskauer test. It uses Barritt’s ( ) reagent containing alpha-napthol and Barritt’s ( ) reagent containing sodium hydroxide. The B reagent reacts with acetoin to form diacetyl, which reacts with the A reagent to form a ( ) complex.

Indole Test

Detects the production of indole by bacteria that use tryptophanase to digest tryptophan into Indole, pyruvic acid, and ammonia.

Kovac’s/positive

In the indole test, when indole combines with ( ) reagent, a red band forms at the top of the medium, indicating a ( ) result.

Hektoen Enteric Agar

A selective and differential media that inhibits gram positive bacteria and helps differentiate between different gram-negative enteric pathogens.