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Credit test 2 Flashcards

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
Which antigenic systems can be used for detection of microbial antigens?
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
26
Myobacterium is stained by which method?
Ziehl – Neelsen staining
27
Which dyes are used in agarose gel electrophoresis and why?
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
What kind of potential changes can be observed by testing by Phenotypic tests?
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
What's typical for Brucella?
- High affinity to alkaline stains | - Not really acid resistant but are resistant to DECOLORIZATION by weak acids
30
Typical features of the endospores?
- 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
Which two bacterias are Catalase negative?
Streptococcus Enterococcus
32
Typical for Triple Sugar Iron, TSI?
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
For what is the Agarose gel electrophoresis?
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
What's typical for the Brucella and Myobacterium?
The they are acid resistant. ``` Brucella = Acid resistant Myobacterium = Acid + Alcohol resistant ```
35
What's typical for the Agarose gel electrophoresis ?
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
What's coagulase test used for?
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
Which two groups can antibodies be divided into?
Monoclonal = Interacting with only one epitope/determinant of one antigen type Polyclonal = Can react with multiple antigen determinants
38
Describe the Giemsa Romanowsky staining method
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
Which method is used for capsule staining of bacteria?
Giemsa - Romanovsky staining method or Burri staining method
40
Common features of the capsule?
- Low ability to take up stains | - Sensitive to fixation (why no heat is used)
41
What's typical for Precipitation method ?
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
What does the ELISA method consist of?
- Ag - Ab - Conjugate: Enzyme linked to Ag or Ab - Substrate
43
The selective staining methods are used for which bacterias?
Myobacterium and Brucella
44
Agglutination can be divided into qualitative and quantitative. Which of the 3 agglutination reactions belong in which?
Qualitative aggl. = Slide (rapid) and Latex agglutination Quantitative aggl. = Tube agglutination (slow)
45
What's typical for Tryptophan degradation to Indole?
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
Which method is used for spore staining?
Wirtz - conclin staining method
47
The PCR mixture must contain what?
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
Typical for ELISA method
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
What is an endospore?
A special resistant and dormant structure
50
What's latex agglutination?
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
What's typical for Catalase test?
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
Which bacterias are forming spores / endospores?
Gram positive
53
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?
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
Typical for Immunoflourescense
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
What's typical for the DNA polymerase / Taq polymerase of PCR reaction?
That it's ONLY active in 72 degrees. | Can also stand higher temperatures; Thermostable!
56
What are the main serological methods and what are they used for?
Agglutination and Precipitation For determination of the antibody or antigen concentration
57
What's the primary function of most endospores?
To ensure survival of a bacteria through periods of environmental stress
58
For what is the special staining method for?
Endospore and capsule staining
59
What do we determine by series of phenotypic tests?
Determine the enzymatic /metabolic activities of microorganisms
60
Which structure is a dormant stage in the life cycle of certain bacterias which are mainly rod-shaped
Endospores
61
Typical for the Burri staining method
``` Applying of Indian ink Air dried Carbolfuchsin is added - Cells shown red/dark pink - Surrounding of the cells are colourless capsules ```
62
Is the capsule an essential part of the cell?
No but have important functions for the cell
63
What's Urease test used for?
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
What's a capsule and what is it built up of?
It's an outer layer situated outside of the cell wall. Consist of Polysaccharides
65
Which are the 3 agglutination reactions?
Slide, tube and latex agglutination

Microbiology (1 decks)

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