Box III.

Essentials for Identification of non-tuberculous mycobacteria (NTM)

Sample collection and transportation to the laboratory

For respiratory specimens, individuals should not rinse their mouths with tap water or other fluids before submitting the specimen.

Use a sterile, leak proof, disposable plastic container. Avoid waxed containers. Swabs are not recommended for the isolation of mycobacteria.

Collect specimens aseptically, reducing contamination with indigenous microbiota.

Collect initial specimens before antimicrobial therapy is started.

Three early morning specimens collected on three consecutive days are ideal.

For induced sputum, sterile hypertonic saline (3-5%) should be used. Avoid contamination with nebulizer reservoir water.

In case of BAL or bronchial wash, bronchoscope should be sterile, cleaned with suitable disinfectant not with tap water and saline used should be devoid of any micro-organism growth. (Lidocaine used during BAL procedure may inhibit growth of NTM).

While collection of extrapulmonary specimens, surgical instruments should be cleaned cautiously avoiding tap water or stored water. Formalin should not be used as transfer medium.

Once samples stored in container, it should not be opened until it reaches to the laboratory.

Store at 2-8°C (do not freeze) if transport is delayed more than one hour; should not be kept more than one week

Precautions in the laboratory

Effect of disinfectant depends on concentration of the disinfectant, duration of disinfection and mycobacterial load in solution or on surface.

Avoid use of chlorine, benzalkonium chloride, cetylpyridinium chloride, quaternary ammonium compounds, and phenolic- or glutaraldehyde-based disinfectants as NTM are resistant to these chemicals.

Use of tap water or stored distilled water should be avoided.

Use of 70% alcohol and 5% phenol as disinfectant is recommended for bench surface cleaning and biosafety filters.

Autoclaving (at 131°C under 15 psi pressure) of plasticware and glassware used in laboratory is strongly recommended.

Laboratory workers should look for contamination by other micro-organism such as Pseudomonas aeruginosa, Staphylococcus aureus, Nocardia, Aspergillus, etc.

Incubation temperature for every species may vary between 27-45°C and requires constant monitoring.

Selective drug susceptibility testing should be done.

Laboratory workers should be aware about the patient’s disease status and must co-ordinate the treating physician while reporting NTM species and subspecies.

BAL, bronchoalveolar lavage. Source: Ref. 130