Mycobacteriology is of utmost importance due to the prevalence of Multiple Drug Resistant Tuberculosis (MDRTB) seen in the last few years.  Proper collection, quick turn-around time and drug sensitivities are important in the plan to control the spread of this disease.  Clinical specimens may be sent to the laboratory for isolation, identification and drug sensitivity.  We are currently using the MB/BacT broth system for isolation.  We use the Auramine-Rhodamine stain in our fluorescent antibody technique to determine presence of AFB on smears.

Specimen Requirements

• Multiple samples should be collected on consecutive days and not on the same day.
• All samples must be submitted in the 50 ml conical tubes provided with the TB kit and the TB form.
• The Submitter will be notified by phone of any unusual delays or any problems that may arise with your specimens.
• Specimens for AFB Smear & Culture are processed twice a week, usually Tuesday & Friday.

Sputum, bronchial washings
Collect three separate sputum samples in the early morning on three consecutive days.  Sputum volume MUST be between 5 ml and 10 ml.  Specimens must be submitted in the 50 ml conical tubes provided by the laboratory.
Urine
An early morning clean catch or catheterized specimen is acceptable.  Specimen volume MUST be between 5 ml and 10 ml. Urine specimens must be received by the laboratory within 24 hours of collection and should be mailed on ice.
CSF
Spinal fluid may be submitted in the CSF collection tube without being transferred to our 50 ml collection tube.  Place the CSF tube inside the 50 ml collection tube if it will fit.
Other body fluids
Pleural, pericardial, synovial, and ascitic, aseptically collect fluid and add a small amount of heparin to prevent clotting.  This should be submitted in the 50 ml containers provided in the TB kit.
Tissue
Tissue suspected of containing mycobacteria should be collected aseptically, placed in a sterile container and protected from drying by adding 0.85% sterile saline.  Swabs are not acceptable.

Shipment
Specimens should be shipped to the Public Health Laboratory using the mailing kit provided by our laboratory central services.  Care should be taken to make sure the caps are securely tightened and laboratory forms are not in the same inner container as the specimen in case of leakage.  Please send specimens the day they are collected.  If a delay in shipment is unavoidable, specimen should be refrigerated until shipment.

Criteria for Rejection
1. Pooled specimens
2. Sputum <2 ml
3. Improper collection containers
4. Improper specimen

Reporting/Interpretation

Smear results

• Results of specimen smears will be reported the same day they are tested
• Smear results will be reported as positive or negative and graded as 1+ to 4+ depending on the number of organisms seen.
  0 = Negative for AFB
  1-2 / 70 fields = + / - resubmit
  2-18 / 50 fields = 1+
  4-36 / 10 fields = 2+
  4-36 / field = 3+
  >36 / field = 4+
• Positive smear results will be phoned the same day they are read, and a hard copy of all results will be sent to the submitting physician and to the TB program.

Culture Results

• Cultures must be held for 8 weeks before a final report is available.
• Cultures are identified by DNA probe. The probes that are available are M. tuberculosis complex and M. avium complex.   If the culture is other than one of these it will be identified by sequencing at the WPHL. 
• Only confirmed M. tuberculosis species will be forwarded to the National Jewish Hospital for drug sensitivity testing.
• Additionally, only the first confirmed isolate from a given patient will receive the drug sensitivity.  Only upon special request, and with approval of the Laboratory Director, will any other isolates be forwarded to NJH for drug sensitivity.
• A hard copy of the culture results will be sent to the submitting physician and the TB program.

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