Tools and Techniques of Microbiology Lab

Question 1

What is the difference between a screening/presumptive test vs a definitive/confirmatory test

Answer 1

Screening/presumptive test= less expensive, rapid, sensitive but less specific test

Definitive/confirmatory test = more expensive, less rapid but more specific test

Question 2

What is sensitivity

Answer 2

• Can it detect it

- the ability of the test to detect low levels of antigen or antibody when present

Question 3

What is specificity

Answer 3

• Is it accurate
• the ability of the test to accurately detect only the correct antigen or antibody while not reacting with incorrect antigen or antibody

Question 4

What are ways to ID microorganisms

Answer 4

1. Gram stain
2. Culture characteristics via microscopy
3. Biochemical test (expression of gene traits)

Question 5

What are the 4 types of general culture media and there characteristics

Answer 5

1. Nutrient= general media that allows bacterial growth
2. Enriched= general nutrients plus extras to grow more fastidious bacteria
3. Selective= contains ingredients to restrict growth of certain type of bacteria
4. Differential= contains ingredients that visually indicate certain chemical reactions by bacterial growth

Question 6

Describe a Blood Agar Plate

Answer 6

moderately enriched medium that permits visual observation of blood cell hemolysis

Question 7

What are the 3 categories of hemolysis in a blood agar plate

Answer 7

1. Alpha=green around the colony, represents partial hemolysis
2. Beta= clear zone around colony, represents complete lysis
3. Gamma= no change in RBCs around the colony

Question 8

Describe a Supplemented Chocolate Agar

Answer 8

highly enriched medium that grows medically significant bacteria, w/o displaying hemolysis

Question 9

Describe a MacConkey Ager (MAC)

Answer 9

selective and differential (pH) media for gram negative bacilli that

Question 10

How are yeasts cultured and identified

Answer 10

• Grow on blood agar plates

- ID= microscopy, biochemical test, antigen test

Question 11

How are molds cultured and identified

Answer 11

-Grow on selective media w/ Abx

-ID= microscopic exam of culture/organism
few antigenic or biochemical test

Question 12

How are viruses cultured and identified

Answer 12

• Grow in living tissue= not usually done anymore because of cost and time
• ID= antibody detection (did you have it), antigen detection (do you have it now), gene probes

Question 13

Summarize Immunoassay Reaction Procedures

Answer 13

1. Antigens bind to specific antibodies
2. Unbound antibodies are washed of, so they will not cross react
3. Antibody-Antigen binding is detected

low to moderate sensitivity

Question 14

What is a Lateral Flow Immunoassay and how does it work

Answer 14

1. assay in which a known molecule and detection molecules are embedded in an artificial membrane
2. specimen is deposited on one end and is wicked through the membrane to react with the detection molecule
3. If the specimen is positive binding becomes visible by deposit of the bound molecules at the test line

Question 15

What is the Agglutination method

Answer 15

The reaction of particle sized antigen with its corresponding antibody resulting in a macroscopic clumping

low sensitivity

Question 16

What is immunofluorescence

Answer 16

fluorescent molecule produces light at a detectable wave length when excited by UV light

Question 17

What is a Enzyme Immunoassay (EIA) or Enzyme Linked Immunosorbent Assay (ELISA) and how does it work

Answer 17

1. A known antibody is labeled w/ color producing enzyme
2. The labeled antibody is then reacted w/ an unknown antigen and unbound antibodies are washed off
3. The antigen-antibody are exposed to a color producing substrate to make the reaction visible

Very sensitive and accurate test

Question 18

What is a gene probe

Answer 18

test kits that are used to detect the presence of significant gene sequences

Question 19

What is gene amplification

Answer 19

the process of increasing the number of copies of genes so they can be detected

Question 20

What 2 gene amplification systems are used in DOD clinical and CBRN labs

Answer 20

BioFire Film Array and Cepheid GeneXpert Omni

Question 21

When do you use oil immersion

Answer 21

when you need high power 90-100x

Question 22

What is an antimicrobial susceptibility test

Answer 22

• used to determine if the pathogen is susceptible or resistant to a set of Abx
• decision to use this test is based on clinical significance of the pathogen (ie pathogen is unlikely to have resistance)

Question 23

Define susceptible in regards to antimicrobial susceptibility test

Answer 23

The microorganism is inhibited by the maximum safe dose of the ABX

Question 24

Define resistant in regards to antimicrobial susceptibility test

Answer 24

The microorganism is affected by the maximum safe dose of the ABX

Question 25

Define intermediate in regards to antimicrobial susceptibility test

Answer 25

The microbe maybe inhibited by a high dosage but is near the resistance point and may not be appropriate to use

Question 26

What is minimum inhibitory concentration (MIC)

Answer 26

test to determine the minimum concentration of each Abx agent that is effective against a pathogen to promote a more precise selection of dosage

Question 27

True or False an Abx should be 2x the minimum inhibitory concentration (MIC)

Answer 27

True to ensure the concentration is always above MIC

Question 28

How is minimum inhibitory concentration (MIC) interpreted

Answer 28

susceptible (2x MIC), intermediate, and resistant (2x MIC can not be achieved

Question 29

What is the disk diffusion testing method

Answer 29

Only used in rapidly growing aerobic and facultative bacteria

Paper disk containing known concentrations of Abx are applied to inoculated agar. A zone of inhibition can then occur, which is then measured and is interpreted using a detailed chart