Clinical and Laboratory Standards Institute standard M24—Susceptibility Testing of Mycobacteria, Nocardia spp., and Other Aerobic Actinomycetes includes susceptibility testing procedures for Mycobacterium tuberculosis complex (MTBC), clinically significant slowly and rapidly growing mycobacterial species, Nocardia spp., and other aerobic actinomycetes. Also included in this standard are recommendations for selecting agents for first-line and second-line drug testing, organism group–specific methodologies, reporting recommendations, and organism quality control criteria. Recommendations regarding agent selection for testing mycobacteria are based primarily on published guidelines. For testing MTBC, M24 recognizes agar proportion as the reference methodology on which all other methodologies are based. In addition, this standard includes recommendations for using commercial broth susceptibility methods with shorter incubation times, which are now in widespread use for MTBC susceptibility testing, and information on molecular methods for detecting drug resistance and their integration with culture-based methods.

This standard replaces the previous edition of the approved standard, M24-A2, published in 2011. Several changes were made in this edition, including: 
• Removed information related to the short-incubation, liquid-radiometric testing system, because this system is no longer available

• Expanded the description of molecular testing for both MTBC and NTM to determine antimicrobial susceptibility or resistance

– For MTBC, Table 3 (Considerations for Molecular or Repeat Testing After Initial Testing on MTBC Using a Commercial Short-Incubation Broth System) and text are included to describe the integration of molecular and culture-based test results for the best possible prediction of the expected drug efficacy.

– For NTM, text is included to describe integration of molecular techniques to assist in determining efficacy of macrolides and amikacin in the treatment of infections caused by MAC and various RGM.

• Added a description of recently discovered challenges to MTBC AST accuracy with use of rapid broth systems and/or the agar proportion method, particularly limited sensitivity in detection of low-level resistance to rifampin and ethambutol 

• Added information in Appendix A regarding the relationship of pharmacokinetics and pharmacodynamics in determining breakpoints and interpretive criteria 

• Updated all breakpoint and quality control tables and moved them to a newly created informational supplement, CLSI document M62

M24 includes antimicrobial susceptibility testing (AST) protocols for three major categories of mycobacterial species: 
• Mycobacterium tuberculosis complex (MTBC) 
• The slowly growing nontuberculous mycobacteria (SGM) 
• The rapidly growing mycobacteria (RGM) 

Also provided are: 
• AST recommendations for Nocardia spp. and other aerobic actinomycetes 
• Guidance on selecting first-line and, for some organisms, second-line antimicrobial agents for testing and reporting 
• Instructions for performing the standard agar proportion (AP) method for MTBC and broth microdilution for mycobacteria and aerobic actinomycetes 
• Molecular methods for detecting mutations associated with MTBC drug resistance 
• QC protocols for each organism category 

Testing and reporting recommendations and QC procedures apply to both reference methods and commercial shorter-incubation broth systems that have been regulatory organization cleared or approved for testing MTBC. This standard does not cover identification methods, nor does it provide an in-depth discussion of molecular test procedures. 

This standard is intended for use by hospital, public health, and referral laboratories that perform AST on MTBC, nontuberculous mycobacteria (NTM), Nocardia spp., and/or other aerobic actinomycetes.