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Microbiology > Credit test 2 > Flashcards

Credit test 2 Flashcards

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1
Q

What’s Simmons citrate?

A

Determines which bacterium that can utilize Citrate as the only source of carbon and energy.
- Bacteria can break the conjugate base salt of citrate into organic acids + CO2, whereby the CO2 bind with Na from the base salt and forming Sodium carbonate

End-product: Sodium carbonate which can change the colour according to the pH indicatior

pH indicator: Bromthymol blue
- Detecting presence of Sodium carbonate.

If positive;

  • Presence of Sodium carbonate
  • The color changes from green to blue

If negative;
- If remains green. Shows that bacteria is not able to utilize the sodium carbonate.

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2
Q

What are the divisions of endospores?

A

Size
- large / small
Location
- Terminal / central / subterminal

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3
Q

What’s typical for carbohydrate fermentation with gas production?

A
  • If it’s able to able to ferment a specific carbohydrate/use sugar the metabolic end-product will be acid and gas

pH indicator = Bromthymol blue
- Changes colour from green –> yellow when pH is low. Shows a positive result.

Sometimes ONLY the organic acid is produced and not gas. But both still positive reactions

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4
Q

Procedure of Stamp staining

A

For acid resistant Brucella bacterium

Fixed by flame!
Primary dye; carbolfuchsin added 
Decolorization: Acetic acid
Rinse under water 
Apply malachite green as counter stain

Brucella: Red
Other bacteria: Green

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5
Q

Typica for methyl red test, MRT?

A

Broth contain peptone, buffers and sugars(dextran or glucose)
- Different bacteria convert these sugars into Pyruvate by using DIFFERENT METABOLIC PATHWAYS.

Some pathways produce unstable acidic products: quickly converted to Neutral compounds
- Some organisms uses the Butylene glycol pathway –> Producing Neutral END PRODUCTS.

Sometimes stable acid products like lactic, acetic and formic acid. Will remain acidic.

pH indicatior = Methyl red

  • If indicator added and color remains red = Acidic and positive
  • If acidic products are changed neutral compounds it changes to yellow
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6
Q

Typical for Ziehl - Neelsen staining method?

A
  • For acid/alcohol resistant Mycobacterium
  • Primary stain is Carbolfuchsin which also contains Phenol
  • Addition by heat
  • Decolorizer: Acidic alcohol
  • Non-acid fast cells take up Malachite green

Myobacterium: Red
Other bacteria: Green

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7
Q

What’s typical for Wirtz - Conclin staining method?

A
  1. Usage of H20 / hot vapers since spores are very resistant to catch dye, these help with penetrate the dye into the bacteria / endospore
  2. Dry preparation in air and then fix with flame.
  3. Apply Malachite green = Primary stain
  4. Add water for decolorizing
  5. Apply of carbolfuchsin / Safranin = Counter stain
  6. Rinse preparation under running water, let it dry.
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8
Q

What are the DNA amplification by PCR method?

A

Cyclic processes with changing temperature. Each cycle consists of 3 steps

  1. Denaturation
    - Highest temperature! –> 95 degrees
    - Denature template of DNA so get two single strands
  2. Hybridization / Annealing
    - Lowest temperature –> 40 – 60 degrees
    - Depends on primer length
    - Primers anneal (hybridize) complementary “stick”, to either side of target DNA sequence and thus flank “mark”, the target sequence to be amplified
  3. Extension (Elongation of Primers)
    - Free nucleotides are added by the polymerase enzyme / Taq DNA polymerase to the 3´ end of each primer at it’s optimal temperature to get 2 double stranded DNA’s. 2,4,6,8…..
    - Temperature optimum –> 72 degrees
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9
Q

What’s the function of the capsule?

A

1) To protect the bacterial cell and is often associated with pathogenic bacteria because it serves as a barrier against Phagocytosis by white blood cells, drying and harmful agents.
2) Attachment for adhesion to the surface.

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10
Q

What’s the tube agglutination?

A

Saline + antiserum

Diluting antibodes by constantly transferring 1/2 of saline from tube to tube.

  • In the tubes from the start are saline and antigen
  • The last tube where agglutination can be seen indicates a positive test
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11
Q

What are the spore coat composed of?

A

Several protein layers

Impermeable to many toxic molecules

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12
Q

What’s the difference between antigen and antibody?

A

Antigen =
A substance penetrating into the body and induces a production of antibodies.
Can almost be anything; polysaccharides, protein, sugars, toxins, bacteria etc

Antibody =
A protective substance that’s produced by the organism
Responsible for Ag-Ab complex

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13
Q

What’s typical for Agglutination?

A

Corpusclar antigens.
Visible changes with the eye.

Agglutinogene reacts with specific antibodes called Agglutinins.
1st step: Specific. Agglutinins bind to antigen
2nd step: Non - specific. Ag- Ab complex forms an insoluble complex, Agglutinate that is VISIBLE

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14
Q

Brucella is stained by which method?

A

Stamp staining

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15
Q

Method for detection of microbial antigens?

A

Detection of bacterial antigens, serological reactions. Base on in VITRO qulitative and quantitative evaluation of antigen-antibody reaction

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16
Q

What are the virulence factors of bacteria?

A

Enzymes and toxins

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17
Q

What are endospores resistant to?

A
  • Ultraviolet and gamma radiation
  • Desiccation
  • Lysozyme
  • Temperature
  • Starvation
  • Chemical disinfectants
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18
Q

What’s typical for both Myobacterium and Endospores?

A

That both of them require water above boiling point for the dye to be able to penetrate into the cell

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19
Q

What cause the thermal thermostability of spores?

A

It’s high content of Dipicolinic and Calcium

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20
Q

Which two bacterias are Catalase positive?

A

Staphylococcus and Micrococcus

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21
Q

Describe the Hydrolysis of Gelatin

A

Detection of Proteolytic activity of positive bacteria

If positive test;
If bacteria can produce the enzyme gelatinase which can break down the gelatin and cause it to remain in liquid form.

Negative test;;
If no production of gelatinase and gelatin remains intact, the media can solidify in the refrigerator

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22
Q

What’s important to think about before when it comes to Giemsa - Romanowsky staining method?

A

That the stained preparation should NOT be fixed by flame!

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23
Q

What’s Burri staining used for?

A

Capsule staining

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24
Q

What’s typical for Myobacterium

A
  • Acid/Alcohol resistant
  • Wax - like cell walls that’s nearly impermeable
  • Contain Mycolic acid, FA, waxes and complex lipids
  • Gram positive
  • Needs to be stained above boiling point by Phenol; helps the dye to penetrate into the cell
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25
Q

Which antigenic systems can be used for detection of microbial antigens?

A

O - antigens: Lipo-polysaccharide . Somatic or corpuscular antigens
H - antigens: Situated on tails, flagellas. Composed of proteins.
K - antigens: Capsular antigens. Composed of polysaccharides
F - antigens: Fimbrial antigens on bacterial pilli. Composed of proteins

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26
Q

Myobacterium is stained by which method?

A

Ziehl – Neelsen staining

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27
Q

Which dyes are used in agarose gel electrophoresis and why?

A

Ethidium bromide and Bromohpenol blue.

Ethidium bromide for:
- Makes DNA or RNA strands visible after agaration under UV light

Bromophenol blue for:
To see MIGRATION of DNA.
Will see movement from - to + of the DNA strand

28
Q

What kind of potential changes can be observed by testing by Phenotypic tests?

A

Observing changes or presences of some things.

  • Make one or more specific chemicals
  • Has certain enzymes
  • Fermentation ability of one or more specific carbohydrates
  • Fermentation ability of individual substrates
  • If require specific nutrients for it to grow
29
Q

What’s typical for Brucella?

A
  • High affinity to alkaline stains

- Not really acid resistant but are resistant to DECOLORIZATION by weak acids

30
Q

Typical features of the endospores?

A
  • Have a resistant and dormant structure
  • Helping the bacteria to survive = NOT a virulence factor
  • Resistant to catching dyes (heat usually needed or water vapors)
31
Q

Which two bacterias are Catalase negative?

A

Streptococcus Enterococcus

32
Q

Typical for Triple Sugar Iron, TSI?

A

For identification of Gram negative ENTERIC RODS

To see the bacterias ability to utilize sugars; Glucose, sucrose and lactose

If fermentation:

  • pH indicator changes colour from red -> yellow
  • Acid - end product
  • Positive fermentation of sugar

If NO fermentation

  • Color remains red
  • Alkaline surrounding
  • Red indicates a negative fermentation of sugar
  • The Iron of the sugar can catch hydrogen sulfide
    If black color = Bacteria producing hydrogen sulfide + occurence of gas/CO2 can occur.
    = FeS positive
33
Q

For what is the Agarose gel electrophoresis?

A

For evaluation of basic DNA methods by detection of Nucleic acid bacteria

Can also determine the SIZE of the separated strands by comparison to DNA ladder - a mixture of DNA fragments

34
Q

What’s typical for the Brucella and Myobacterium?

A

The they are acid resistant.

Brucella = Acid resistant
Myobacterium = Acid + Alcohol resistant
35
Q

What’s typical for the Agarose gel electrophoresis ?

A

It separates the DNA according to it’s size

a) Separates and distinguish DNA fragments by movement in agarose gel
- Will move from NEGATIVE to POSITIVE
pole
b) Strength of electric field also plays in
c) Smaller molecules move quicker than larger
d) Moves slower if there’s more agarose

36
Q

What’s coagulase test used for?

A

For distinguishing if Staphylococcus are pathogenic or non-pathogenic .

Determines if the organism can produce the Coagulase enzyme or not.
Fibrinogen –> Fibrin –> Clot = Positive test

Pathogenic = Coagulative positive
Non - pathogenic = Coagulative negative

37
Q

Which two groups can antibodies be divided into?

A

Monoclonal = Interacting with only one epitope/determinant of one antigen type

Polyclonal =
Can react with multiple antigen determinants

38
Q

Describe the Giemsa Romanowsky staining method

A
  1. Usually spleen or liver organ
  2. Usually GR dye
  3. Distally water added.
  4. Rinse under water and allow to dry.
    NOT HEAT for fixation.
  • Bacterial bodies will be stained blue
  • Capsules are pink coloured
39
Q

Which method is used for capsule staining of bacteria?

A

Giemsa - Romanovsky staining method or Burri staining method

40
Q

Common features of the capsule?

A
  • Low ability to take up stains

- Sensitive to fixation (why no heat is used)

41
Q

What’s typical for Precipitation method ?

A

Soluble antigens; Precipitonogens reacting with specific antibody’s(Precipitins) and producing a VISIBLE Ag-Ab complex

Visible in that way that it’s forming a precipitation ring in the test tube!
= A zone equivalence between the antigen and antibody.
- Antigen at the bottom of test tube

42
Q

What does the ELISA method consist of?

A
  • Ag
  • Ab
  • Conjugate: Enzyme linked to Ag or Ab
  • Substrate
43
Q

The selective staining methods are used for which bacterias?

A

Myobacterium and Brucella

44
Q

Agglutination can be divided into qualitative and quantitative. Which of the 3 agglutination reactions belong in which?

A

Qualitative aggl. = Slide (rapid) and Latex agglutination

Quantitative aggl. = Tube agglutination (slow)

45
Q

What’s typical for Tryptophan degradation to Indole?

A

The broth contains a high concentration of the AA Trp.

Using of Kovac’s reagent for detection of color differences.

Enzyme Tryptophanase enzyme degrade Trp -> Indole

If a red ring on top of broth;

  • Indole is present, positive reaction
  • Able to use Trp from the medium /Positive for the Tryptophanase enzyme

If a yellow ring on top of broth;
- No indole is present, a negative reaction

46
Q

Which method is used for spore staining?

A

Wirtz - conclin staining method

47
Q

The PCR mixture must contain what?

A
  1. Template DNA or RNA
  2. Primers -> 2 obligonucleotides for flanking/marking the target sequence
  3. DNA polymerase / Taq polymerase
  4. Free nucleotides
  5. Reaction buffer: With a suitable concentration of Mg2+ ions
48
Q

Typical for ELISA method

A

Not visible with eye
Bind to different enzyme; Most often to horse radish peroxidase and Alkaline phosphatase

Direct / Indirect method
Substrate detect presence of antigen or not.
If colour change in Ab-enzyme binding = Been a substrate bounded to the Ag-Ab complex

49
Q

What is an endospore?

A

A special resistant and dormant structure

50
Q

What’s latex agglutination?

A

For determination of antigen or antibody where one of these in the reaction is binded to latex particles .

Only in Staphylococcus oreos!!!

If granules visible on surface, agglutination, specific binding = Positive test

51
Q

What’s typical for Catalase test?

A

Imp for classification of bacteria!
Some bacteria uses Catalse to fight with toxic compounds like below.
- For detection of Hydrogen peroxide and Superoxide.

Using enzyme for conversion of these toxic compounds –> Diatomic oxygen and water.

If the bacteria can produce Catalse; This transformation mentioned above occurs
- Oxygen gas causes bubbles immediately = Positive test

  • If no bubbles = Negative test
52
Q

Which bacterias are forming spores / endospores?

A

Gram positive

53
Q

Acid fast organisms generally require a special staining method since the cell wall is so resistant to most compounds.
Which primary stain is used and why?

A

Use generally acid-fast staining Carbolfuchsin since it’s lipid soluble and also contains phenols which helps the stain to penetrate through the cell wall after addition of heat

54
Q

Typical for Immunoflourescense

A

Use fluoroscent dyes and NOT enzymes
- Reacting with proteins and doesn’t cause changes in their biological characteristics

Direct: Ab to tissue Ag LABELED with Fluorocrome

Indirect: Ab to tissue Ag UNLABELED

55
Q

What’s typical for the DNA polymerase / Taq polymerase of PCR reaction?

A

That it’s ONLY active in 72 degrees.

Can also stand higher temperatures; Thermostable!

56
Q

What are the main serological methods and what are they used for?

A

Agglutination and Precipitation

For determination of the antibody or antigen concentration

57
Q

What’s the primary function of most endospores?

A

To ensure survival of a bacteria through periods of environmental stress

58
Q

For what is the special staining method for?

A

Endospore and capsule staining

59
Q

What do we determine by series of phenotypic tests?

A

Determine the enzymatic /metabolic activities of microorganisms

60
Q

Which structure is a dormant stage in the life cycle of certain bacterias which are mainly rod-shaped

A

Endospores

61
Q

Typical for the Burri staining method

A 
Applying of Indian ink
Air dried
Carbolfuchsin is added 
- Cells shown red/dark pink
- Surrounding of the cells are colourless capsules
62
Q

Is the capsule an essential part of the cell?

A

No but have important functions for the cell

63
Q

What’s Urease test used for?

A

Differentiation especially of Enterobacteriacae

Bacteria utilizes urea and form ammonia during incubation

Phenol red indicator in medium for detection of the enzyme production

Causes an alkaline reaction, pH increase –> colour change

  • From yellow to pink = Positive reaction
  • Stays yellow = Bacteria is not producing the Urease enzyme
64
Q

What’s a capsule and what is it built up of?

A

It’s an outer layer situated outside of the cell wall.

Consist of Polysaccharides

65
Q

Which are the 3 agglutination reactions?

A

Slide, tube and latex agglutination

Microbiology (1 decks)

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