Which Nontuberculous Mycobacteria (NTM) organism is negative for Polymerase Chain Reaction (PCR) and positive for Acid-Fast Bacilli (AFB)?

Nontuberculous Mycobacteria Organisms Negative for PCR and Positive for Acid-Fast Bacilli

Several nontuberculous mycobacteria (NTM) species may be positive on acid-fast bacilli (AFB) staining but negative on polymerase chain reaction (PCR) testing, particularly those not targeted by the specific PCR assay being used. This discrepancy typically occurs with less common NTM species that aren't included in standard multiplex PCR panels.

Diagnostic Considerations for NTM

Common NTM Species and Detection Methods

The most clinically significant NTM species include:

• Mycobacterium avium complex (MAC)
• Mycobacterium kansasii
• Mycobacterium abscessus complex
• Mycobacterium massiliense

Standard laboratory methods for identifying NTM include:

1. AFB Staining: Detects all mycobacteria based on acid-fast properties
2. Culture: Gold standard for identification but requires weeks for growth
3. PCR Testing: Typically designed to detect specific NTM species

PCR Limitations in NTM Detection

Most multiplex PCR assays are designed to target only the most common NTM species 1. According to research, one-step multiplex PCR identified specific NTM species in only 49.09% of isolates, while 50.91% were identified only to the genus level (Mycobacteria spp.) despite being AFB-positive 1.

The 2020 ATS/ERS/ESCMID/IDSA guidelines emphasize that "expert consultation should be obtained when NTM are recovered that are either infrequently encountered or that usually represent environmental contamination" 2. This recommendation acknowledges the diagnostic challenges with less common NTM species.

NTM Species Often AFB-Positive but Potentially PCR-Negative

NTM species that may be AFB-positive but missed by standard PCR panels include:

• Mycobacterium species without targeted primers in the PCR assay: About 50.91% of NTM isolates in one study were identified only as Mycobacteria spp. by PCR 1
• Less common pathogenic species: M. xenopi, M. fortuitum, M. chelonae, M. gordonae
• Rare or emerging NTM species: These may not be included in commercial PCR panels

Clinical Significance of AFB-Positive, PCR-Negative Results

When encountering an AFB-positive, PCR-negative result:

1. Consider specimen contamination: Environmental NTM can contaminate specimens 2
2. Evaluate pathogenicity: Some NTM species like M. gordonae rarely cause disease, while others like M. kansasii are usually pathogenic 2
3. Collect multiple specimens: According to guidelines, "in general more than one culture-positive specimen for NTM is necessary for diagnostic purposes" 2

Diagnostic Algorithm for AFB-Positive, PCR-Negative Specimens

1. Confirm AFB positivity with fluorochrome staining (preferred method) 2
2. Culture on both liquid and solid media to allow growth of fastidious species 2
3. Identify to species level using:
   • High-performance liquid chromatography (HPLC)
   • DNA sequencing
   • Extended antibiotic susceptibility testing 2
4. Apply diagnostic criteria for NTM pulmonary disease:
   • Clinical symptoms
   • Radiographic findings (nodular/cavitary opacities or bronchiectasis with nodules)
   • Microbiologic criteria (multiple positive cultures) 2

Important Clinical Considerations

• Pathogenicity varies significantly among NTM species, from rarely pathogenic (M. gordonae) to usually pathogenic (M. kansasii) 2
• Meeting diagnostic criteria doesn't necessitate treatment: A careful assessment of risks and benefits should guide treatment decisions 2
• Geographic variations exist in NTM species distribution and pathogenicity 2

Common Pitfalls to Avoid

1. Misdiagnosing as tuberculosis or MDR-TB: NTM lung disease can appear similar to TB but is resistant to primary anti-TB drugs 1, 3
2. Relying on a single positive culture: Generally insufficient for diagnosis except in specific circumstances 2
3. Assuming all AFB-positive specimens are M. tuberculosis: PCR testing specifically for M. tuberculosis is essential to differentiate from NTM 4
4. Overlooking NTM in patients with previous TB: NTM may colonize damaged lungs secondary to TB 1

When encountering AFB-positive, PCR-negative specimens, comprehensive microbiological workup with species identification is essential for proper diagnosis and management of potential NTM disease.