Specimen Collection and Submission Guidelines for Pathologic Evaluation of Possible Mycobacterial Infections

Tissue samples are typically submitted for mycobacterial testing when there is strong clinical suspicion of tuberculosis, a bacterial infection caused by species within the Mycobacterium tuberculosis complex. Nontuberculous mycobacteria, such as Mycobacterium avium-intracellulare complex (MAC) and Mycobacterium marinum, can also cause clinically significant pulmonary and cutaneous infections, which may be particularly severe in the immunocompromised host.

Mycobacterial infections often cause granulomatous inflammation in tissues, which can be observed in routine hematoxylin and eosin (H&E) stained slides. The H&E dyes are used to stain tissue slides and serve as a key foundation of histopathology.  The findings typically prompt further routine testing in the pathology laboratory for acid-fast bacilli (AFB) using special stains, in addition to further clinical laboratory testing.

At IDPB, molecular testing for Mycobacterium species, including M. tuberculosis complex can be performed as indicated according to clinical and epidemiologic information provided by the submitter, and the histopathologic findings in the submitted tissue specimens. Testing by immunohistochemical assays and for other pathogens may be performed as indicated.

CDC’s Tuberculosis Reference Laboratory can perform Molecular Detection of Drug Resistance (MDDR) testing. DNA will only be transferred for samples in which M. tuberculosis complex is detected in IDPB.

Additional Required Information for Specimen Submission*

Provide a clinical summary, copies of pertinent recent clinical notes or a cover letter detailing:

• The patient’s initial presentation
• TB or other antimicrobial treatment provided
• Risk factors for mycobacterial infection, including:
  • Underlying medical conditions or immunosuppressive therapy
  • History of travel or residence outside of the United States
  • Other pertinent exposures like exposure to an active TB case, and for non-tuberculous mycobacteria, any water, outdoor, or animal exposures
  • Any prior mycobacterial diagnoses and treatments, including prior history of latent tuberculosis infection (LTBI).

Provide a copy of the surgical pathology or autopsy report, or summary of these findings with results of special stain testing for AFB, for example, Ziehl-Neelsen, performed prior to submission.

Provide results of all pertinent clinical laboratory testing such as sputum AFB smear and culture, interferon gamma release assays (IGRA), and PCR assays.

*Please note that if any of the above information is not provided, there will be no further processing of the case. Because of the high volume of mycobacterial cases tested at IDPB, we suggest ensuring your submission is complete, so we can provide results in a timely manner.

Submission of Specimens

Formalin-fixed paraffin-embedded (FFPE) tissue blocks

Preferred specimen type for histopathology, histochemistry (special stains), immunohistochemistry, PCR and sequencing, or nucleic acid extraction for transfer to other CDC laboratories. FFPE tissue blocks must be less than 10 years of age.

Biopsy and autopsy tissue specimens are only acceptable if embedded within 2 weeks after being placed in formalin.

The distribution of bacteria in tissues can be minimal and patchy; therefore, we suggest the provision of adequate tissue blocks that demonstrate pathology like granulomas. For scant specimens, submission of the original H&E and acid-fast stained slides in addition to the tissue blocks may be requested.

Formalin-fixed wet tissue

Biopsy tissues: NOT acceptable for testing.
Autopsy tissues:  only acceptable if the duration of formalin-fixation has been within 2 weeks, or if tissues have been transferred to 70% ethanol within 2 weeks after initial placement in formalin. Brain autopsy tissue may take longer than 2 weeks for full formalin-fixation, but fixation cannot exceed 4 weeks.

Unstained paraffinized tissue slides

Because evaluation for mycobacterial infections requires molecular (PCR) testing, unstained slides are NOT acceptable for testing.

Formalin-fixed paraffin-embedded (FFPE) tissue scrolls

This specimen type is NOT acceptable for testing.