CLSI 2026: Antibiotic Selection and Zone Interpretation Criteria for Acinetobacter spp.

Antimicrobial susceptibility testing (AST) measures an antimicrobial agent’s ability to inhibit the growth of a microorganism. Several AST methods are available to determine bacterial susceptibility to antimicrobials. The disk diffusion method is an inexpensive and very common method.

AST is carried out using standardized laboratory methods and is interpreted according to guidelines established by organizations such as the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST).

Breakpoints are MIC values or zone diameters that classify an organism as susceptible (S), intermediate (I), or resistant (R) to a specific antimicrobial agent. Breakpoints vary not only according to the antimicrobial but also to the organism. This document summarizes the updated zone diameter breakpoints and interpretation criteria for Acinetobacter spp. in the disk diffusion method as published in the CLSI M100, Performance Standards for Antimicrobial Susceptibility Testing, 36^th Edition (2026). It replaces CLSI M100, 35^th Edition, published in 2025, and includes revised disk diffusion and MIC breakpoints for amikacin, gentamicin, and tobramycin.

CLSI 2026 Breakpoints for Acinetobacter spp.

The following table lists the CLSI 2026 zone diameter breakpoints for Acinetobacter spp. All values represent zones of inhibition (ZOI) measured in millimeters (mm).

Selection of Antibiotics for AST of Acinetobacter spp.

• Tier 1: Appropriate for routine, primary testing and reporting.
• Tier 2: Appropriate for routine, primary testing. Reporting may follow institution-specific cascading rules.
• Tier 3: Introduced for institutions with high levels of multidrug-resistant organisms. Appropriate for routine, primary testing and reporting but must follow institution-specific cascade rules.
• Tier 4: Reserved for testing/reporting on request. Used if other antimicrobial agents in other tiers are not optimal because of various factors.
• Others (-): Antimicrobial agents not listed in a table that contains antimicrobial agents that should be considered for testing and reporting by microbiology laboratories, but for which established clinical breakpoints are available.

Testing conditions for Acinetobacter spp.

• Medium: Muller Hinton Agar (MHA)
• Inoculum: Colony suspension, equivalent to a 0.5 McFarland standard
• Number of antibiotic discs: Maximum 12 discs on 150 mm plate, no more than 6 discs on 100 mm plate
• Distance between discs: Discs should be placed no less than 24 mm apart (center to center)
• Incubation: 35C±2°C, ambient air, 20-24 hours

Intrinsic Resistance of Acinetobacter spp.

Acinetobacter spp. (and all other non-lactose-fermenting gram-negative bacteria) are intrinsically resistant to the following antibiotics:

• Penicillins
• First and second generation cephalosporins
• Clindamycin/Daptomycin
• Glycopeptides (vancomycin)
• Linezolid
• Macrolides (e.g., Erythromycin, Azithromycin)
• Quinupristin-dalfopristin
• Rifampin

In addition to the above antibiotics, Acinetobacter spp. also shows intrinsic resistance to the following antibiotics:

• Ampicillin, amoxicillin
• Amoxicillin-clavulanate
• Ertapenem
• Aztreonam
• Trimethoprim
• Chloramphenicol, and
• Fosfomycin

References

1. CLSI. Performance Standards for Antimicrobial Susceptibility Testing. 36th ed. CLSI supplement M100. Clinical and Laboratory Standards Institute; 2026.