Advanced Diagnostic Laboratories

tNGS for Mycobacteria

Test Code

TNGS

Description

Identification and susceptibility testing of Mycobacterium tuberculosis complex (MTBC) and nontuberculous mycobacteria (NTM) has a long turnaround time because the organisms grow so slowly. Targeted next-generation sequencing (tNGS) is a molecular method that allows our laboratory to identify mycobacterial species and predict resistance to 13 anti-tuberculosis drugs within days instead of months. This method can be used on both isolates and sputum. The tNGS assay has been validated to:

Identify MTBC and NTM from isolates
Identify MTBC directly from sputum. Do not sputum if only NTM is suspected.
Predict resistance in MTBC for up to 13 anti-tuberculosis antibiotics in isolates or sputum. These include rifampin, isoniazid, pyrazinamide, ethambutol, ethionamide, streptomycin, fluoroquinolones, kanamycin, amikacin, capreomycin, linezolid, clofazimine, and bedaquiline.

Collect

This test can be ordered on a clinical sputum specimen.

Processed sputum: 1-2 mL. Less than 1 mL of processed sputum will be rejected.
For unprocessed sputum, submit 5-10 mL. If between 3-5 mL is submitted, a disclaimer of low sample volume will be added. Samples <3 mL will be rejected.

This test can also be ordered on organisms recovered elsewhere.

Pure mycobacterial isolates should be sent in any routine solid or liquid media. Isolates can be tested after transfer and/or subculture in 7H9.
Culture volume: Laboratory must receive visible growth of an isolate on solid media or an isolate in broth media (≥3 mL).

Unacceptable Conditions

Specimens that do not meet the minimum volume requirement
Specimens from a source not acceptable for this testing
Samples received outside of temperature/stability
Specimens that are excessively leaking
Mislabeled or unlabeled specimen

Stability

See collection requirements

Performed

Once every 1-2 weeks

Methodology

Targeted next-generation sequencing

Turnaround Time

2-3 weeks. Turnaround time may be longer if isolate is not pure, or if isolate is submitted in other media and needs to be cultured in 7H9 broth.

Department

Infectious Disease Laboratory, Mycobacteriology

Synonyms

Sequencing for TB
Sequencing for mycobacteria
Direct sequencing

Study Offerings

CAP/CLIA

Related Tests

TNGSI, TNGSR

Reference Interval

Possible results:

Test	Isolates	Sputum	Notes
Identification	Genus and species of mycobacteria. No mycobacteria detected More than one mycobacteria detected	Mycobacterium tuberculosis complex, with species identification where possible. No Mycobacterium tuberculosis complex detected	NTM cannot be identified directly from sputum. Samples must have sufficient bacterial load and must meet quality metrics for a valid result This assay does not adequately differentiate between M. bovis and M. bovis BCG.
Resistance genes (resistotype)	Susceptible Resistant Variant of unknown significance	Susceptible Resistant Variant of unknown significance	Resistance genes are only available for MTBC. These include rifampin, isoniazid, pyrazinamide, ethambutol, ethionamide, streptomycin, fluoroquinolones, kanamycin, amikacin, capreomycin, linezolid, clofazimine, and bedaquiline. Variants of unknown significance are often correlated with a susceptible phenotype, but NOT always. Additional testing should be performed. Samples must have sufficient bacterial load and must meet quality metrics for a valid result.

Interpretation

MTBC or NTM can be identified from an isolate. MTBC can be identified directly from sputum while NTM are not validated from sputum. Detection of MTBC from a sputum is a critical result.

MTBC resistance genes can be identified from both isolates and sputum. Mutations are analyzed according to the WHO's 2nd edition catalogue of mutations and reported as resistant or susceptible. Mutations detected that do not correlate clearly with either will be reported as "variants of unknown significance."