Smear, Culture and Identification of Acid Fast Bacilli

Test Code

AFB3

Description

This test includes AFB smear, culture, and organism identification from a clinical specimen.

The AFB smear is a direct fluorescent stain from the specimen and can be used to quickly establish the presence of mycobacteria. The AFB culture is the inoculation of a clinical specimen onto multiple media types followed by monitoring for six weeks. Growth of any AFB (i.e. mycobacteria) or partially acid fast organisms (e.g. aerobic actinomycetes) is identified. Identification will occur to the greatest extent possible, which is usually to the species level but can be to the subspecies level for some organisms (e.g. Mycobacterium abscessus subsp. abscessus).

This test is typically used when there is low risk of tuberculosis since no direct test for Mycobacterium tuberculosis is performed from the specimen (although it is fully identified if it is seen). If there is an unknown or higher risk of M. tuberculosis infection, order AFB1.

Collect

Specimen	Preferred Volume	Unacceptable Conditions	Collection Container	Total stability time, including transport
Sputum (expectorated or induced)	5-10 mL	<3 mL	Sterile, leak-proof container.	7 days refrigerated; 72 hours ambient.
Lower respiratory tract (e.g. bronchoalveolar lavage, bronchial washes etc.)	5-10 mL	<3 mL	Collect washing and aspirates in a sputum trap. Place brushes in a sterile, leak-proof container with up to 5 mL sterile non-bacteriostatic saline.
Body Fluids (e.g, pericardial, synovial, pleural etc.)	2-4 mL	<2 mL	Sterile, leak-proof container.
Cerebrospinal Fluid (CSF)	2-4 mL	<2 mL refrigerated/frozen CSF.	Sterile, leak-proof container.	72 hours ambient. These sources should not be refrigerated.
Fresh Tissue	1-2 g	<1 g or tissue submitted on a dry swab or fixed in formalin or other preservatives.	Sterile, leak-proof container with tissue submerged in sterile non-bacteriostatic saline.
Blood	5-10 mL	< 5 mL or blood collected in a red-top, EDTA (purple-top), or ACD (yellow-top) tubes.	Sterile, leak-proof blood collection green (heparin) or yellow (SPS) top tube.
Gastric lavage or wash	10 mL	<10 mL or a specimen that has not been neutralized within 1 hour of collection.	Sterile, leak-proof container. Neutralize specimen within 4 hours of collection with 100 mg of sodium carbonate.
Urine	5-10 mL required	<3 mL, Pooled urine specimen, or a urine specimen that has been mixed with preservatives..	Sterile, leak-proof container.	7 days refrigerated.
Stool	1-2 g	<1 g or a stool specimen in PVA, formalin, or other preservative, rectal swabs.	Sterile, leak-proof container.	7 days refrigerated.

Unacceptable Conditions

Samples collected in wax containers are not acceptable. Wax containers may produce false-positive smear results.
Swabs are not recommended for the isolation of mycobacteria, since they provide limited material and the hydrophobicity of the mycobacterial cell envelop often compromises a transfer from the swabs onto solid media or into broth media.
Specimen received in the lab 14 days post-collection

Notes

Stool cultures for mycobacteria are not encouraged.
AFB smear is not performed for urine, stool, or blood.

Performed

Typically all days

Methodology

When appropriate, specimens are processed using a standard digestion/decontamination procedure. An acid fast smear is performed from the processed specimen using a florescent auramine stain. Specimens are cultured on three solid and one liquid media then incubated for 6 weeks. Mycobacteria or partially acid fast organisms are identified to the full extent possible, such as to the species or subspecies level (see AFB4 for further details).

Reported

AFB Smear reported within 24 hours. Note that AFB smears are not performed for urine, stool, or blood.
The final report for the AFB culture is issued at 6 weeks. Solid media cultures are read at 1, 3, and 6 weeks. Liquid media cultures are continuously monitored. Positive cultures are reported at the time of positivity and negative cultures are reported at 1, 3, and 6 weeks. In some cases, media may be incubated for up to 8 weeks and if positive, a supplemental report will be issued.
Identification is reported within 3 days to 2 weeks, depending on the method(s) used.

Department

Infectious Disease Laboratory, Mycobacteriology

Synonyms

AFB3, mycobacterial smear, culture and identification

Related Tests

AFB7, AFB4

License Comments

Smear and culture: Reference method, no special licensing required
Identification of AFB: NY State waiver is required.

Reference Interval

Negative for acid fast bacilli.

Interpretation

Test	Interpretations
AFB Smear	Fluorochrome stain read at 400X. No AFB found Doubtful: Repeat specimen recommended. 1 to 2 AFB/ 70 fields observed. 1+ = 2-18 AFB/ 50 fields observed. 2+ = 4-36 AFB/ 10 fields observed. 3+ = 4-36 AFB/ field observed. 4+ = >36 AFB/ field observed.
AFB Culture	Negative cultures are reported as negative for growth Positive cultures are reported as organisms recovered with a colony count where possible (CFU/mL)
AFB Identification	Organisms are identified to the species or subspecies level, where relevant.

CPT Code

This is a test algorithm. When it is ordered, smear and culture are typically performed. If an organism is recovered, a reflex identification test will be ordered. The identification method used will depend on the organism.

Test	Method	CPT Code	Billing Code	When is this applied?
AFB Smear	Fluorochrome Stain	87206	CXAFB1	All samples, except blood, stool and urine
AFB Culture	Specimen processing	87015	CXAFB2	All samples
AFB Culture and presumptive ID	Specimen processing	87116	CXAFB3	All samples
AFB Identification	PCR/MALDI-TOF mass spectrometry	87798x9	MASID (1st organism), MASI2 (2nd organism), MASI3 (3rd organism)	If appropriate organisms are recovered
AFB Identification	Line probe assay	87798x7	NTMID (1st organism), NTMD2 (2nd organism), NTMD3 (3rd organism)	If appropriate organisms are recovered
MTB Identification	Line probe assay	87556	MTB7T	If an appropriate organism is recovered
rpoB Identification	Sequencing	87153	RPOB	If an appropriate organism is recovered
16S Identification	Sequencing	87153	16S	If an appropriate organism is recovered