Introduction

2023 Breakpoint Implementation Toolkit

Clinical laboratories performing antimicrobial susceptibility testing (AST) should use breakpoints currently
recognized by Clinical and Laboratory Standards Institute (CLSI) or US Food and Drug Administration (FDA).
CLSI, Association of Public Health Laboratories (APHL), American Society for Microbiology (ASM), College of
American Pathologists (CAP), and Centers for Disease Control and Prevention (CDC),have jointly developed
this toolkit to assist clinical laboratories in updating minimal inhibitory concentration (MIC) breakpoints. It
is provided in a streamlined format and designed to guide performance of a verification or validation study
required to update breakpoints. There are links to other resources that explain the rationale behind breakpoint
updates, regulatory requirements for updating breakpoints, and detailed instructions for performing an AST
breakpoint validation or verification. Manufacturers of AST systems can provide guidance on breakpoints used
and clearance status with their systems.

Contents
Introduction
How to Use 2023 Breakpoint Implementation Toolkit
Definitions/References/Resources
Pathway for Updating Breakpoints (Figure 1)

Part A. Breakpoints in Use

Template for Documenting Breakpoints in Use

Part B. CLSI vs FDA Breakpoints

Listing of all current CLSI breakpoints (M100-Ed33) and corresponding FDA breakpoints

Part C. Breakpoint Implementation Summary

Template for documenting results of a validation study to update breakpoints

Part D. CDC and FDA AR Bank BIT Isolate Sets

CDC and FDA Antibiotic Resistance Isolate Bank isolate sets available for use in breakpoint verification and
validation studies

Part E. Instructions for Use of 2023 BIT Part F

Instructions for using the prefilled Excel workbook

Part F. AR Bank Data Entry and Calculations

Excel workbook prefilled with the AR Bank BIT set isolates and their results

Part G. Blank Form for Data Entry

Template for validations or verifications using isolates other than those listed in the AR Bank BIT sets included
in this toolkit

CLSI Version 1.0. This was last updated on 15 March 2023 and has been approved by CLSI’s Outreach
Working Group.

Toll Free (US): 877.447.1888 | P: +1.610.688.0100 | F: +1.610.688.0700 | E: [email protected]

How to Use 2023 Breakpoint Implementation Toolkit
It is assumed that those using this toolkit have some knowledge of the need to update AST breakpoints. Please
check references in the list below for more detailed information about updating breakpoints.
The APHL-ASM CRO Breakpoint Implementation Toolkit published in 2022 and accessible here contains detailed
instructions, as well as worksheets and forms for validating updated carbapenem breakpoints, and is based
on the CAP Breakpoint Implementation Toolkit (2012, no longer available). These instructions can be adapted
to verification or validation of other breakpoints when following guidance included in the 2023 Breakpoint
Implementation Toolkit.
In brief, it is suggested that you proceed as follows:
1. Determine what breakpoints are in use in your laboratory.
2. Determine which of these breakpoints are old or out of date (eg, no longer recognized by CLSI or FDA) and
would require updating for continued reporting.
3. Develop a priority list and a plan for updating breakpoints.
Refer to the flowchart in Figure 1 that highlights a detailed pathway for updating breakpoints.
Refer to Part A here to assist you in documenting breakpoints in use.

IMPORTANT NOTES:
1. Laboratories are encouraged to implement updated CLSI breakpoints as listed in M100, 33rd ed.
2. If CLSI breakpoints differ from FDA breakpoints, (see Part B) a laboratory can elect to use current CLSI or
FDA breakpoints.
3. Manufacturers of commercial AST must use FDA breakpoints that are current at the time they submit a
test for FDA clearance.
4. A laboratory should NOT use breakpoints that are no longer recognized by CLSI or FDA.
5. As of January 2024, CAP-accredited laboratories may be penalized if they use breakpoints that are no
longer recognized by CLSI or FDA. US laboratories must at least adopt breakpoints within three years of
the date of official publication by FDA accepting the revised breakpoints.

Definitions/References/Resources
In the context of updating to current CLSI breakpoints:
Verification is used to evaluate the performance of breakpoints which have been FDA cleared for use on a
device manufacturer’s AST system (ie, FDA recognizes the CLSI breakpoints, and the manufacturer has obtained
clearance by FDA for the current CLSI/FDA breakpoints on their AST system).
Validation refers to any other scenario not covered by verification and when the laboratory is modifying an
FDA-cleared test (eg, using breakpoints that are different from those that are FDA cleared for use on the device
manufacturer’s AST system).
NOTE: Details regarding how AST device manufacturers implement updated breakpoints for their system can
be found in Table 3.

CLSI Version 1.0. This was last updated on 15 May 2023 and has been approved by CLSI’s Outreach
Working Group.

Toll Free (US): 877.447.1888 | P: +1.610.688.0100 | F: +1.610.688.0700 | E: [email protected]

Table 1. Situations Where Breakpoint Verification or Validation Is Required to Use Updated CLSI Breakpoints
Updated Performance
Breakpoint Status Commercial Antimicrobial Susceptibility Testing System Status Assessment Requireda
CLSI=FDA CLSI breakpoints are FDA cleared and available on panel/software. • Verificationb
• 10 to 15 isolates/drug
CLSI=FDA Device manufacturer has notified customers that the device has • Verificationb
received FDA clearance with updated CLSI/FDA breakpoints and • 10 to 15 isolates/drug
has advised customers how to implement breakpoints with their
panels/software.
CLSI=FDA Device manufacturer has not received FDA clearance of the device • Validation (if desire to
with updated CLSI/FDA breakpoints. use CLSI breakpoints)
• 30 isolates/drug
CLSI≠FDA Manufacturer must provide FDA breakpoints; use of CLSI • Validation (if desire to
breakpoints would be off label. use CLSI breakpoints)
• 30 isolates/drug
a
Consensus suggestions from authors of 2023 Breakpoint Implementation Toolkit
b
If no change to the test has been made by the AST manufacturer (eg, no reformulation of drug dilutions), a verification of reporting
may be sufficient. This would involve ensuring MIC results are interpreted correctly on patient reports.

CLSI Version 1.0. This was last updated on 15 May 2023 and has been approved by CLSI’s Outreach
Working Group.

Toll Free (US): 877.447.1888 | P: +1.610.688.0100 | F: +1.610.688.0700 | E: [email protected]

Table 2. Resources for Updating Breakpoints
Resource Location Comments
Current CLSI Breakpoints Performance Standards for Antimicrobial Updated annually, usually in
Susceptibility Testing, 33rd ed. CLSI supplement February
M100. Clinical and Laboratory Standards
Institute; 2023.
Current FDA Breakpoints Susceptibility Test Interpretive Criteria (STIC) Updated as new information
is obtained from CLSI and/or
pharmaceutical company and
reviewed by FDA
CLIA regulations for verification CLIA §493.1253(b)(1); No specific details for
and performance specifications verification of AST systems
CAP requirements for updating Contact CAP to obtain complete checklist; see (Available to CAP-accredited
Breakpoints below for specific breakpoint requirements laboratories)
CLSI. Verification of Commercial Purchase from clsi.org 1st edition is currently under
Microbial Identification and revision; describes verification
Antimicrobial Susceptibility in detail; validation is
Testing Systems. 1st ed. CLSI discussed briefly in
guideline M52. Wayne, PA: Appendix B
Clinical and Laboratory Standards
Institute; 2015.
APHL-ASM CRO Breakpoint APHL Implementation Toolkit Contains extensive
Implementation Toolkit instructions, worksheets, and
forms for validating updated
carbapenem breakpoints

Table 3. Considerations for Updated Breakpoint Implementation in Commercial AST Devices

• FDA clearance status of a test for an AST device may not be synonymous with up to date FDA breakpoints (ie,
clearance may have been granted with previously published FDA breakpoints).
• If a test was FDA cleared, and breakpoints were subsequently updated by CLSI, the manufacturer must wait
for FDA to review rationale documentationa and update the FDA Susceptibility Test Interpretive Criteria (STIC)
website to reflect FDA acceptance of the updated CLSI breakpoints.
• The AST manufacturer can re-submit performance data to FDA for authorization to update the breakpoints for
the test for that device only when the FDA STIC website update has occurred.
a
Rationale documents prepared by CLSI when breakpoints are updated can be found here with a companion webinar here.

CLSI Version 1.0. This was last updated on 15 May 2023 and has been approved by CLSI’s Outreach
Working Group.

Toll Free (US): 877.447.1888 | P: +1.610.688.0100 | F: +1.610.688.0700 | E: [email protected]

References
References Addressing Potential Negative Impact of Using Obsolete Breakpoints
Ambrose PG, Bhavnani SM, Andes DR, et al. Old in vitro antimicrobial breakpoints are misleading stewardship
efforts, delaying adoption of innovative therapies, and harming patients. Open Forum Infectious Diseases.
2020;7(7). doi: https://doi.org/10.1093/ofid/ofaa084.
Humphries RM, Hindler JA, Epson E, et al. Carbapenem-resistant Enterobacteriaceae detection practices in
California: what are we missing? Clin Infect Dis. 2018; 66(7):1061-1067. doi: https://doi.org/10.1093/cid/cix942.
Johnson WM, Clark JA, Olney K, et al. Changing times: The impact of gram-negative breakpoint changes over
the previous decade. Antimicrob Stewardship & Healthcare Epidem. 2022;2(1). doi:
https://doi.org/10.1017/ash.2022.301.
Yarbrough ML, Wallace MA, Potter RF, et al. Breakpoint beware: reliance on historical breakpoints for
Enterobacteriaceae leads to discrepancies in interpretation of susceptibility testing for carbapenems and
cephalosporins and gaps in detection of carbapenem-resistant organisms. Eur J Clin Microbiol Infect Dis.
2020;39(1):187-195. doi: https://doi.org/10.1007/s10096-019-03711-y.

References Addressing Understanding Breakpoint Updating

Humphries RM, Abbott AN, Hindler JA. Understanding and addressing CLSI breakpoint revisions: a primer for
clinical laboratories. J Clin Microbiol. 2019;57(6). doi: https://doi.org/10.1128/JCM.00203-19.
Newitt VN. AST and safety at core of microbiology checklist changes. CAP Today. October 2021.
https://www.captodayonline.com/ast-and-safety-at-core-of-microbiology-checklist-changes/
Accessed March 16, 2023.
Simner PJ, Rauch CA, Martin IW, et al. Raising the Bar: Improving Antimicrobial Resistance Detection by Clinical
Laboratories by Ensuring Use of Current Breakpoints. Open Forum Infectious Diseases. 2022;9(3). doi:
https://doi.org/10.1093/ofid/ofac007.

To access the entire BIT Toolkit, visit https://clsi.org/bit-toolkit/.

CLSI Version 1.0. This was last updated on 15 May 2023 and has been approved by CLSI’s Outreach
Working Group.

Toll Free (US): 877.447.1888 | P: +1.610.688.0100 | F: +1.610.688.0700 | E: [email protected]