3.1 Biochemical test strips

Multiple biochemical tests in a single kit form, come as a strip with micro wells and reagent. The testing process is straightforward; inoculate, incubate, add some more reagents then read and compare the result to a standard. Examples of testing kits include Analytical Profile Index (API, Biomerieux) and Microbact (Thermofisher).

Activity 7: API identification of Gram-negative pathogens

Timing: Allow 15 minutes

In this activity you will first watch a video showing how to set up an API test for the identification of Gram-negative pathogens, and then answer some related questions.

First, watch the video ‘Performing an API20E strip test’ (2016).

Note that in the video Enterbacterales are referred to by the old term ‘Enterobacteriaceae’.

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Transcript

Add water to 3/4 fill all of the honeycomb wells of the tray

Flame a loop until it is red hot then allow it to cool for 20 seconds

Select a single isolated colony and emulsify in 10 mils of sterile distilled water

Use a new sterile pipette

For CIT, VP and gel tests fill both the tube and cupule

Fill only the tubes for the remaining tests

Add one or two drops of paraffin oil two tests: ADH, LDC, ODC, H2S and URE

Incubate the strip at 37°C overnight

After incubation add biochemical reagents to the required tests

One drop each of VP1 and VP2 to the VP test

One drop of Kovacs reagent to the IND test and

One drop of the TDA reagent to the TDA test

After 10 minutes compare the colours against the chart and record on an API record sheet

End transcript
 
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Now, answer the following questions:

  1. What temperature is the test kit incubated at overnight?
  2. Would the same API kit be suitable for all of the following: E. coli, Shigella, Salmonella, Klebsiella, Haemophilus influenzae?
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Discussion

  1. The test kit should be incubated at 37°C overnight.
  2. No. ‘E’ in API20E stands for Enterobacterales (formerly known as Enterobacteriaceae), so all of these organisms are identified by API 20E except Haemophilus influenzae. Haemophilus influenzae, as a non-glucose fermenter, is not classified as Enterobacterales. A different kit (API 20NE – ‘Non-Enterobacterales’) would be needed to identify Haemophilus influenzae.

Activity 8: Individual biochemical tests versus test kits

Timing: Allow 10 minutes

What are the pros and cons of using individual biochemical tests rather than test kits such as API or similar?

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Discussion

Test type Advantages Disadvantages
Individual
  • Lower cost
  • Only need to stock essential tests
  • Gives enough information to identify most isolates to guide treatment
  • Interesting for laboratory technologist as requires skill
  • Hard to get an accurate identification unless you have all of the required reagents
  • Time consuming
  • Quality control of all reagents is required. Different reagents may need different storage conditions.
  • Requires a high level of expertise
Test strip kits
  • Easier and less time-consuming to perform
  • Gives a numerical answer which can be used to give a relatively accurate identification
  • Rapid 4-hour versions available for some kits
  • Higher cost
  • Most need 24-hour incubation so may take longer to get a result
  • Refrigerated storage required
  • Online access to a Standards database is desirable, though not essential
  • Leads to a loss of laboratory interpretive skills and expertise

3 Commercially available and more advanced tests

3.2 Chromogenic media