MYCOBACTERIOLOGY

Mycobacteriologist         Jody Fleming BS MT (ASCP)      (307) 777-3609

Microbiologist                 Valerie O'Neill                                (307) 777-3327 

 

Tuberculosis (TB) or Mycobacterium tuberculosis complex (MTBc) is still a serious public health threat in the United States.  Screening for TB in Wyoming is important to control the human-to-human transmission of this deadly disease.  To quickly control the spread of TB, rapid turn around times (TAT) from collection of the specimens to identification is required.  At WPHL, the Mycobacteriology Laboratory can culture a variety of specimen types, perform fluorescent Acid Fast Bacillus (AFB) smears, identify the organism that causes TB, and facilitate drug susceptibility testing.

 

TYPE OF TEST

AFB smear, Nucleic Acid Amplification Testing (NAAT), 16s rDNA sequencing, and Drug Susceptibility Testing (DST).

SPECIMEN REQUIREMENTS

All specimens must be submitted in propertly labeled sterile 50 ml conical tubes provided with WPHL's TB mailer.  Carefully follow packing instructions provided with mailer to prevent leakage during transit.

Respiratory Specimens - Sputums and Bronchial Washings.  Sputums should be collected in the early mornings on 3 consecutuve days.  Volume collected should be approximately 5 - 10 mls.  We prefer to have respiratory specimens shipped daily, but it is acceptable to ship the three collections together.

Urine - An early morning clean catch or catherized specimen is acceptable.  Specimen volume should be at least 10 mls and must be received by WPHL within 24 hours of collection.

Cerebral Spinal Fluid - Spinal fluid may be submitted in a CSF collection tube.  The collection tube can be placed inside the sterile 50 ml conical tube for shipment if it will fit inside securely.

Body Fluids - Pleural, pericardial, synovial, ascitic fluid should be aseptically collected and submitted in a sterile 50 ml conical tube.

Tissues - Tissues suspected of containing mycobacterium should be collected aseptically and placed in a sterile container and protected from dehydration by adding sterile physiological saline solution.

Specimens not accepted for TB testing - Specimens not accepted for testing are swabs, blood, and stool.

SHIPPING REQUIREMENTS

Use the mailing container supplied by WPHL.  Ship according to regulations for infectious substances.  Make sure caps are securely tightened on the 50 ml conical tubes and wrap with parafilm to prevent leakage.  Complete and enclose WPHL TB test request form.  The test request form should not be placed in the inner container.  Specimens should be submitted the day they are collected from the patient.  If a delay in shipment is unavoidable, refrigerate the specimens until shipment occurs.  Submitters will be notified by phone of any problems associated with their specimen.

DAYS TEST PERFORMED

AFB smears are done on Tuesdays and Fridays.

RESULTS AVAILABLE

Upon completion of testing.  All positive results are called to the submitter and a hard copy is mailed.  Mycobacteriology cultures are held for 6 weeks before a final negative report is released.

REPORTING & INTERPRETATION

AFB smear results are reported as "POSITIVE" or "NEGATIVE".  Positive AFB smears are graded from 1+ to 4+ depending on the number of AFB observed.  See Table 1 below.

Reflex testing:  Cultures that grow AFB are identified using DNA probes for MTB complex and M. avium complex.  Cultures that are negative for these 2 probes are further identified using 16s rDNA sequencing.  Only confirmed isolates of MTB complex will be forwarded to our reference laboratory for drug susceptibility testing (DST).  The first confirmed isolate from a patient will receive DST and additional approval is required to forward other isolates for DST.

ADDITIONAL COMMENTS

Rejection criteria: 

   Pooled specimens.

   Specimens > 5 days old.

   Sputum volume < 2 mls.

   Improper specimen (swab, blood, or stool).

   Inappropriate collection container.

   Improper or incomplete labeling on specimen container or WPHL test request form. 

Positive specimens:  AFB smears that are positive will be sent to our reference lab for NAAT testing.

 

Table 1.  Grades for positive AFB smear results.  

 
0 AFB                            Negative for AFB
1-2 per slide              "Rare or few AFB".  Order a repeat specimen.

1-9 AFB / 100 fields    1+
1-9 AFB / 10 fields       2+
1-9 AFB / field              3+
> 9 / field                      4+

 

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