M. tuberculosis complex DNA Detection

This test replaces MTB complex PCR [TBCPCR]

General Information

Lab Name

Lab Code

TBCDNA

Epic Name

M. tuberculosis complex DNA Detection

Description

Detection of Mycobacterium tuberculosis complex DNA

Tuberculosis (TB) is caused by a bacterium called Mycobacterium tuberculosis. The bacteria usually attack the lungs, but TB bacteria can attack any part of the body such as the kidney, spine, and brain. Not everyone infected with TB bacteria becomes sick. As a result, two TB-related conditions exist: latent TB infection (LTBI) and TB disease. If not treated properly, TB disease can be fatal. In addition to Mycobacterium tuberculosis, the related Mycobacterium africanum and Mycobacterium bovis are considered members of the Mycobacterium tuberculosis complex (MTBC) and also can cause TB disease in human and animal hosts. The diagnosis and control of Mycobacterium tuberculosis complex is a very significant problem in global health.

The identification of Mycobacterium tuberculosis complex influences treatment, and the potential need for patient isolation. Due to the slow growing nature of Mycobacterium tuberculosis complex, and the need for a rapid and accurate diagnosis, molecular methods have replaced phenotypic methods for the identification of MTBC.

Although an acid fast smear (AF smear) can provide an important clue to the nature of the causative agent for a patient's disease, a positive result does not necessarily indicate the presence of Mycobcaterium tuberculosis complex (MTBC). The presence of Acid Fast Bacillis (AFBs) in clinical specimens can be simply due to the growth of common water contaminants such as M. chelonae and M. fortuitum. On the other hand, a negative smear result also does not rule out the presence of M. tuberculosis complex at low pathogen load. Recent developments in PCR based amplification and detection of organism specific DNA offers an alternative that can be more specific and sensitive than a smear test.

The insertion sequences IS6110 and IS1081 have been considered as useful phylogenetic markers for the identification of Mycobacterium tuberculosis complex. It is also known that almost all members of Mycobacterium tuberculosis complex have multiple copies of IS1081 and/or IS6110, providing inherent signal amplification. Internal validations showed that sequence specific primers and molecular beacon probes targeting IS6110 and IS1081 can be utilized to detect Mycobacterium tuberculosis complex while not amplifying the most commonly isolated Nontuberculous Mycobacteria species (M. abscessus, M. avium, M. chelonae, M. fortuitum, M. haemophilum, M. intracellulare, M. kansasii, M. marinum). This has allowed us to design an assay with great sensitivity and specificity for detecting the presence of Mycobacterium tuberculosis complex DNA in patient clinical specimens.

Forms & Requisitions

Molecular Microbiology Order Form

Synonyms

AFB PCR, broad range AFB PCR, broad range Mycobacteria PCR, IS1081, IS6110, M. tuberculosis complex, Molecular AFB, Molecular Mycobacteria, Molecular Mycobacterium, MTB, MTB PCR, MTBC, multiplex PCR, Mycobacteria PCR, Mycobacterial identification, Mycobacterium identification, Mycobacterium tuberculosis complex identification, Mycobacterium tuberculosis complex PCR, TB PCR, TBC, Tuberculosis PCR, universal AFB PCR, universal Mycobacterium PCR

Components

Code	Name
TBCSUM	MTB complex Summary
TB6110	MTB complex PCR: Detection of Insertion Sequence IS6110
TB1081	MTB complex PCR: Detection of Insertion Sequence IS1081
TBCSI	MTB complex PCR: Specimen Description
TBCSPI	MTB complex PCR: External Identifier
TBCSR	MTB complex PCR: Special Requests
TBCSC	MTB complex PCR: Specimen Comments
TBCNAE	MTB complex PCR: Specimen DNA Extraction
TBCREV	MTB complex PCR: Pathologist Review
TBCME	MTB complex PCR: Method Note

Interpretation

Guidelines

Method

DNA extraction, nucleic acid purification, polymerase chain reaction (PCR), sequencing

Reference Range

See individual components

Ordering & Collection

Specimen Type

Tissue (Fresh frozen or paraffin-embedded), Fluid (see Acceptable Specimens for details). When possible, a mycobacterial culture is recommended regardless of the molecular test result to aid in clinical diagnosis.

Collection

Acceptable specimens are listed below. Please see Molecular Microbiology Specimen Submission for complete specimen collection and handling instructions.

Shipping/Handling

Fresh frozen tissue/fluid specimens should be collected into a DNA free container labeled with at least two identifiers and be submitted and maintained on dry ice.
Formalin Fixed Paraffin-embedded tissues (FFPE, PET) can be sent ambient or with ice packs during warmer summer months to prevent melting.

Acceptable Specimens

Fresh frozen tissue
Fresh frozen fluid: any body fluid is acceptable if it is not listed under Unacceptable Specimens.
- Common examples include: cerebrospinal fluid, pleural fluid, pericardial fluid, urine, bronchial lavage, joint fluid, bone marrow, vitreous fluid, etc.
Formalin Fixed Paraffin-embedded tissues (FFPE, PET): blocks, scrolls, and unstained slides. Note that fresh tissue is considered the optimal specimen of choice, as it reduces the chance of introducing exogenous DNA templates or microorganisms during embedding/fixation, and because formalin fixation dramatically reduces the sensitivity of molecular assays due to reduced template yield and quality.
Sputum: acceptable - except for Bacterial PCR reflex NGS [BCTDNA]/Bacterial DNA Detection by PCR (without reflex to NGS) [NRBDNA], Fungal PCR reflex NGS [FUNDNA]/Fungal DNA Detection by PCR (without reflex to NGS) [NRFDNA], and Nontuberculous Mycobacteria DNA Detection [NTMDNA]* assays
eSwabs**, UTM (universal transport media), body fluid/bone marrow in EDTA (not including blood)
Sodium polyanethol sulfonate (SPS, Wampole Isolator Tubes) acceptable with disclaimer

*Mycobacterium avium complex DNA Detection [MAVDNA] can be ordered on sputum

**Fungal PCR reflex NGS [FUNDNA] and Fungal DNA Detection by PCR (without reflex to NGS) [NRFDNA] may have interference due to some lots of eSwabs which have been found to contain Saccharomyces cerevisiae DNA, resulting in false positive detection. Clinical correlation and/or retesting with a different collection method is advised. The detection of S. cerevisiae from eSwab specimens can interfere with our ability to rule out other fungal DNA.

Unacceptable Specimens

Blood, serum, plasma, stool/rectal swabs
No citrated or heparinized solutions
Tissues floating in formalin
Swab/fluid collected in tube containing agar

Optimal Quantity:

Fresh Tissue: 0.3-1.0 cm^3
Fluid: 0.2-1 mL
Formalin Fixed Paraffin-embedded Tissue (FFPE/PET): blocks are preferred over scrolls/unstained slides and will be sent back to client upon completion of testing
Scrolls/unstained slides: cross-sectional area >1cm^2 send 10 sections of 10µm thickness, if <1cm^2 send 20 sections if available

Please note: We do not need a separate specimen aliquot for each test ordered. Only a single specimen aliquot or block of optimal quantity is necessary for performing multiple tests. If multiple aliquots or blocks of optimal quantity are sent, up to 2 will be pooled.

Fresh tissue is the optimal specimen of choice, as it reduces the chance of introducing exogenous DNA templates or microorganisms during embedding/fixation. Formalin fixation dramatically reduces the sensitivity of the assays due to reduced template yield and quality.

When possible, a mycobacterial culture is recommended regardless of the molecular test result to aid in clinical diagnosis.

Forms & Requisitions

Molecular Microbiology Order Form

Handling Instructions

Please see Molecular Microbiology Specimen Submission for complete specimen collection and handling instructions.

Quantity

Requested: See "Collection" for Optimal Quantity
Minimum: Specimens below optimal quantity are acceptable for testing, however, diagnostic yield is generally proportional to specimen size.

Processing

Receiving Instructions: UWMC/HMC: Store and send fresh tissue/fluid specimens refrigerated, if specimen storage and transport will exceed 8 hours, freeze at -20°C. Freeze all fresh tissue/fluid specimens at -20°C upon arrival in UW Molecular Microbiology.

Misc Sendout

Performance

Lab Department

Micro Molecular Diag(MMD)

Frequency

Fresh frozen tissues/fluids result in 2-3 business days after receipt of specimen. Formalin Fixed Paraffin-embedded tissues result in 3-4 business days after receipt of specimen.

Available STAT?

Performing Location(s)

UW-MT	Microbiology, Molecular Diagnostics 206-520-4600 ---------------------------------------- Shipping Address Attn: Molecular Microbiology UW CLSPS 1601 Lind Ave SW Room 117 Renton, WA 98057 Phone: 206-520-4600 Alternate phone: 206-598-6147 Performing Lab Address Clinical Microbiology Lab, NW177 University of Washington Medical Center 1959 NE Pacific Street Seattle, WA 98195 Phone: 206-598-5735 Alternate phone: 206-598-6147	Contact Information Please e-mail us with any questions or comments you may have. Your inquiry will be answered as soon as possible. email: molmicdx@uw.edu The Molecular Microbiology lab is open from Monday-Friday, 7am-4pm PDT. Billing inquiries and requests for faxed reports can be made to our Client Services Department at (206) 520-4600 or (800) 713-5198. For results or other inquiries, we can be reached by phone at the following numbers: Phone: (206) 598-5735 Alternate phone: (206) 598-6147 FAX: (206) 520-4903 For assistance during weekends, holidays and after hours, please contact Lab Medicine Resident at (206) 598-6190

Billing & Coding

CPT Codes: 87556
LOINC: 38379-4
Interfaced Order Code: UOW4350