Tuberculosis Molecular Diagnostic Testing Information for Healthcare Workers and Laboratorians

Mycobacterium tuberculosis Nucleic Acid Amplification Testing (NAAT) Frequently Asked Questions

Which Tuberculosis (TB) Nucleic Acid Amplification Test (NAAT) is used at the Rhode Island State Health Laboratory (RISHL)?

The RISHL utilizes the Cepheid Xpert MTB/RIF assay for NAAT, an automated molecular diagnostic test that is used to detect the presence of Mycobacterium tuberculosis complex (MTB-complex) DNA in respiratory specimens. Once MTB-complex DNA is detected, the Xpert MTB/RIF assay will also determine if mutations associated with resistance to the anti-TB drug rifampin are present. The Xpert MTB/RIF assay has been recommended by the World Health Organization for the diagnosis of TB in pulmonary specimens from both adults and children.

What is the advantage of the TB NAAT?

Conventional culture methods may take up to six weeks to detect growth of M. tuberculosis and several additional weeks for anti-TB drug susceptibility testing, whereas NAAT can be performed directly on sputum and other respiratory specimens and results can be provided to the clinician within 24-48 hours of specimen receipt. In January 2009, CDC published Updated Guidelines for the Use of Nucleic Acid Amplification Tests in the Diagnosis of Tuberculosis.

When should TB NAAT be ordered?

The Centers for Disease Control and Prevention (CDC) recommends in its 2009 guidance that NAAT be performed on at least one respiratory specimen from each patient with signs and symptoms of pulmonary TB for whom a diagnosis of TB is being considered but has not yet been established, and for whom the test result would alter case management or TB control activities. TB NAAT should ONLY be ordered on patients when the clinician’s index of suspicion for pulmonary TB is moderate to high. MORE

What types of specimens are accepted for NAAT?

The Rhode Island State Health Laboratory accepts only the following respiratory specimens for TB NAAT: sputum, bronchoalveolar lavage (BAL), bronchial brushing, bronchial lavage, bronchial washing, and lung washing. The Xpert MTB/RIF assay has been verified in-house for use on these specimen types. Non-respiratory specimen types, such as cerebral spinal fluid, thoracentesis fluid and pleural fluid, gastric lavage/aspirate, lymph node tissue/aspirate, and other tissue types have NOT been validated for testing at the RISHL. more

If a TB NAAT is ordered on a specimen, do I still need to order an AFB smear and culture?

Yes, a TB NAAT does not replace the need for AFB smear and culture (minimum three respiratory specimens collected 8 hours apart, including an early morning specimen). AFB smear and culture must always be ordered on specimens submitted for NAAT. The result of the NAAT is always interpreted in conjunction with the acid-fast bacilli (AFB) smear result and AFB culture remains the “gold standard” for the laboratory confirmation of TB and is required in order to conduct additional anti-TB drug susceptibility testing and genotyping for molecular epidemiological purposes. When a specimen is sent to our laboratory for a TB NAAT, an AFB smear and a culture will also be ordered and performed.

What if AFB smear and culture is performed by my hospital laboratory; will another smear and culture be performed by the State Health Laboratory if a NAAT is ordered?

The Rhode Island State Health Laboratory will require an additional unprocessed specimen be submitted to our laboratory for NAAT. This same specimen will also have an AFB smear and culture done at our laboratory. The method verification that took place in our laboratory was done on specimens that had been through the specimen decontamination process used in our laboratory, so it has not been verified for use on specimens that have been decontaminated by other methods.

How do I order a TB NAAT at the Rhode Island State Health Laboratory?

Complete a Rhode Island State Health Laboratory Test Requisition Form and indicate ‘TB PCR’. Please refer to the Mycobacterium tuberculosis Specimen Submission Guidance and the Guidelines for the Collection and Transport of Specimens for Tuberculosis Testing.

If the State Health Laboratory finds that a specimen is AFB smear positive, how do I order a TB NAAT?

Contact the Rhode Island State Health Laboratory Mycobacteriology Lab at 222-5586/222-5587 if you wish to add on a TB NAAT to a previously received specimen.

What is the expected turn-around time for receiving TB NAAT results?

Our goal is to have NAAT results available within two working days of specimen receipt. Specimens should be sent to the State Health Laboratory as soon as possible after collection.

How will I be notified of the NAAT result?

All positive TB NAAT results will be considered a critical test value and will be immediately phoned to the laboratory that submitted the specimen, the Health Department Tuberculosis Control Program, and the RISE TB Clinic. NAAT results will be routinely reported through our laboratory information system to submitters in the same manner they receive other reports from our laboratory.

Is a negative NAAT result enough to rule out tuberculosis?

No, a single negative NAAT should not be used as a definitive result to exclude TB (refer to NAAT Interpretation Guidelines) especially in a patient where the clinical suspicion of TB is moderate to high. If the clinician is not experienced with the interpretation of NAAT or the diagnosis of TB, consultation with the expert TB clinicians at the RISE TB Clinic is highly recommended (401) 793-2427/2433. more, (Refer to the APHL Consensus statement on the use of Cepheid Xpert MTB/RIF assay in making decisions to discontinue airborne infection isolation in healthcare settings)

I have a patient in respiratory isolation; can this test help me to decide if my patient can be removed from isolation?

A negative NAAT and two independent AFB smear-negative sputum samples (taken at least 8 hours apart, with at least one being an early morning specimen) may be used to determine whether patients may be removed from respiratory isolation, provided that (a) clinical suspicion for infectious TB is low and (b) there is clinical improvement if the patient has been started on multidrug treatment. more (APHL Consensus statement )

Can the Xpert MTB/RIF assay be used to monitor treatment response or to release a newly confirmed TB patient from airborne infection isolation?

No, molecular assays based on DNA detection have not been shown to be suitable for monitoring treatment and the Xpert MTB/RIF assay should not be used to determine when a laboratory confirmed case of pulmonary TB can be released from respiratory isolation. AFB smear microscopy and AFB culture results must be considered in conjunction with clinical data in these cases. MORE (APHL Consensus statement)

Can the Xpert MTB/RIF assay be used to determine the course of anti-TB drug treatment?

No, an Xpert MTB/RIF result for rifampin (Pos or Neg) should be considered preliminary pending confirmation by additional molecular test methods and/or culture-based drug susceptibility testing. An Xpert MTB/RIF result cannot be used to presume susceptibility to other anti-TB drugs, such as isoniazid, ethambutol, pyrazinamide, etc.