API 20 E

Question 1

Why don’t we use biochemical assays

Answer 1

*Biochemical assays using a dichotomus key
*Inexpensive
*Time consuming
*Creates a lot of waste

Question 2

Why don’t we use sequencing?

Answer 2

*Sequencing
*Requires specialized equipment
*Can be expensive

Question 3

Identification testing systems

Answer 3

*Identification testing systems
*Does multiple tests at once
*Can be more expensive, but faster

Question 4

Enteropluri kit

Answer 4

*Tests for Enterobacteriaceae
*Does 15 biochemical tests
*Test results can be converted to a code which then can be used to identify microorganisms according to a codebook

Question 5

*Microgen kits

Answer 5

*12 or 24 test system
*Tests for Enterobacteriaceae and Oxidase positive, Non-fastidious, Gram-Negative bacilli (using 24 test system)
*Gives a numerical code which can beentered into an online database to identify the unknown

Question 6

API suite kits

Answer 6

*API suite
*Multiple test strips available to test for over 700 bacteria
*Typically do 20-22 tests
*The results of the strip are interpreted to give a numeric code, which can then be entered into an online database to determine the unknown
*Depending on the strip used, it can identify:
*Enterobacteriaceae
*Streptococcus species
*Staphylococcus species
*Listeria
*Protozoa and fungi

Question 7

Explain the API 20E system

Answer 7

*Gives a numeric code based on the results of 21 biochemical tests

*20 tests are done on the strip, and 1 test (the oxidase test) is done separately

*Every 3 tests are interpreted and given a numerical value, which are added to give one digit for a the 7 digit code

*The code is then input into an online database called APIWeb which will be linked to a microorganism

*You can also use a dichotomos key to manually interpret the results.

Question 8

What are the limitations of the API 20E system

Answer 8

*The API 20E system only tests for Enterobaceriaceae and Oxidase positive, Non-fastidious, Gram-Negative bacilli

*Gram-Positive bacilli and Gram-Positive or Gram-Negative Cocci will NOT be detected by this system!

Question 9

How to use the API 20E: Inoculating your bacteria steps

Answer 9

1.Distribute 5mL of sterile water into the honeycomb try to keep a humid environment for the test strip

2.Using a sterile loop, pick a colony and inoculate to 5mL of sterile water. Then use a sterile pipette to mix the water and bacteria, creating a homogenous mixture

3.Using the same pipette, fill the tube and cupule of the Citrate (CIT), VP, and Gelatinase (GEL) wells with your bacterial suspension. Fill only the tube of the other 17 wells.

4.Using a new sterile pipette, add sterile mineral oil to the cupules ADH, LDC, ODC, H2S, and URE tubes to create an anaerobic environment

5.Close the incubation box and place in the incubator overnight

Question 10

4 tests requiring reagents will be done after incubation: List the positive rnxs

Indole text
VP test
TDA test
Oxidase test

Answer 10

*The indole test (add 1 drop of Kovak’s reagent. A red color is +)
*The VP test (add 1 drop KOH and 1 drop 1-napthol. A red/rose color is +)
*The tryptophane daminase (TDA) test (add 1 drop TDA reagent. A reddish-brown color is +)
*The oxidase test will also be done using the filter paper method
*Add oxidase reagent to the filter paper. Then grab a colony of your microorganism off the plate and wipe onto the filter paper. Purple is +