CDST_LT: MGIT DST quality control

MGIT DST Quality Control

Quality control of MGIT DST is critical to laboratory testing, as it ensures the accuracy and consistency of the test processes and the results reported. 

Frequency: 

It is important to perform quality control (QC) of drug susceptibility testing periodically. This testing must be performed:

• For each new batch of reagents (MGIT drug kits, other drugs, media, etc.) 

•  Weekly, in a DST run, when patient tests are run weekly.

• With each batch of patient isolates, when DST is performed less frequently.

• If the batch QC fails, all the results obtained within that batch and the new batch of a reagent should be thoroughly reviewed, and the testing should be repeated.

Strains used of MGIT DST Quality control: 

• M. tuberculosis H₃₇Rv (ATCC [American Type Culture Collection] number 27294) is used as a QC strain which is susceptible to all anti-tuberculosis drugs. If the ATCC reference strain cannot be obtained, a well-characterized strain derived from a patient’s isolate that is completely susceptible to first-line anti-TB agents may be used instead. It is preferable to use a strain that has been fully sequenced and shown to have a wild-type pattern for all known genes associated with TB drug resistance.

• It is not necessary to include a resistant strain, as most of the resistant strains against a drug which are available from ATCC and other culture collections are highly resistant and do not give any added benefit in quality control.

Procedure: 

The test procedure for QC organisms is the same as for clinical isolates. The inoculum should be from a freshly grown culture in the MGIT medium or on the Löwenstein-Jensen medium (LJ) slant. After a standardized suspension has been made, it should be frozen at -70° C ±10° C. It may be stored for up to 1 year with no significant decrease in viable counts.

Resources

1. 1. STANDARD OPERATING PROCEDURE FOR MYCOBACTERIOLOGY LABORATORY

2. Revised National TB Control Programme Training Manual for Mycobacterium tuberculosis Culture & Drug susceptibility testing. Central TB Division, Ministry of Health and Family Welfare, GOI. 

3. Technical manual for drug susceptibility testing of medicines used in the treatment of tuberculosis. World Health Organization 2018.

Assessment

The definite diagnosis of tuberculosis demands that M. tuberculosis be recovered on culture media and identified using differential in vitro tests. Hence using laboratory standards is an important aspect of ensuring quality processes in all aspects of MGIT DST. 

These include the following:

1. Standard laboratory biosafety measures as applicable to TB containment laboratories for handling culture isolated and performing DST

2. Trained staff to use standard operating procedures for equipment and laboratory methods 

3. Standard stains for ZN microscopy, reagents and culture media as positive and negative controls

4. Using standard strains with every batch of the medium as a check on drug concentration in drug susceptibility tests

5. Using known standard positive and negative control in all biochemical tests for identification

6. Using standard Laboratory Performance Indicators for Mycobacterial Culture and DST

7. Using standard NTEP registers and formats for day-to-day work and monthly/quarterly abstracts to summarize laboratory activities

Resource

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Quality controls procedures are essential to generate quality DST results. Quality procedures are essential when using QC strains and reagents and preparing and using drugs for DST. These include:

QC strains
​1. Use well-characterized pan-susceptible M. TB strain/ H37Rv/ M. tuberculosis ATCC 27294 
2. Colonies from solid media less than 14 days old​
3. MGIT 960 tube 1–5 days after flagged positively by instrument ​
4. Additional organisms may be tested to supplement BD QC recommendations ​
5. Test with a known mono-resistant strain of M. tuberculosis

Drug and Inoculum Preparation​
1. Ensure proper reconstitution of lyophilized drugs​ by following the given guidelines:

• Rehydrate drugs with sterile distilled water​
• Thaw aliquots of prepared drugs from freezer​
• Check the expiry date before use​

2. Avoid wrong drug concentrations. ​
3. Avoid improper drug preparation.​
4. Proper dilution of inoculum for drug and growth control tube is critical.
5. Suspension must be well mixed and homogeneous without clumps.
6. Prepare QC inoculum (H37Rv) suspension in the same way as patient isolate suspension.

7. Use positive and negative controls.

8. Follow standard operating procedures to perform SIRE/ Second-line and PZA DSTs​.

Resource

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Quality control of MGIT DST inoculum is essential for quality DST results. The different aspects of inoculum preparation include:

1. Using the right inoculum​​
2. Using pure cultures only​
3. Using fresh cultures​
4. Using slant < 14 days

Quality aspects for Positive MGIT tube ​include:
1. Time period should be plus 1 day to 5 days from the date of positivity​
2. Homogeneous inoculum​
3. Using sterile glass beads and vortex to break up organism clumps​
4. Too high or too low an inoculum may give error results or un-interpretable results
5. Preparing dilutions according to the procedure​
6. Using an accurate pipette to add inoculum to tubes​

Resource

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Quality Control of DST is critical to ensure the test is functioning properly so that reliable results can be interpreted. ​

Steps in Quality Control in DST result interpretation include: 

1. SIRE/ Second- line and PZA QC

• Carefully entering the tubes into MGIT 960​
• Carefully scanning the DST set carrier into the BACTEC MGIT 960 instrument.

2. Interpretation of results
 

• The time period to interpret results is crucial. Interpret between 4–13 days for first and second-line drugs and 4-21 days for PZA. ​
• Results should read susceptible for all drugs.
• If proper results are not obtained, repeat the test.

3. First and Second- line drugs and PZA QC Records​
 

• Record lot numbers of MGIT 960 tubes, drugs and drug supplements. ​
• Record QC results.
• Maintain records for a minimum of five years.

Resource