Methods for Dilution Antimicrobial Susceptibility Testing 8th Edition

Methods for Dilution Antimicrobial Susceptibility Testing 8th Edition

Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically outlines standardized procedures for determining the minimal inhibitory concentrations (MICs) of aerobic bacteria. Authored by Matthew A. Wikler and colleagues, this eighth edition provides essential guidelines for clinical laboratories. It covers broth macrodilution, broth microdilution, and agar dilution techniques, ensuring accurate susceptibility testing. The document is crucial for microbiologists and healthcare professionals involved in antimicrobial therapy and resistance monitoring. Updated tables and quality control measures enhance the reliability of susceptibility testing methods.

Key Points

  • Describes broth macrodilution and microdilution techniques for antimicrobial susceptibility testing.
  • Includes updated M100 tables for drug selection and quality control in susceptibility tests.
  • Provides guidelines for testing fastidious organisms and special considerations for specific bacteria.
  • Outlines quality control procedures and responsibilities for accurate susceptibility testing.
  • newtopiccyclegrowin
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January 2009
M07-A8
Vol. 29 No. 2
Replaces M07-A7
Vol. 26 No. 2
Methods for Dilution Antimicrobial
Susceptibility Tests for Bacteria That Grow
Aerobically; Approved Standard—Eighth
Edition
This document addresses reference methods for the determination of minimal inhibitory
concentrations (MICs) of aerobic bacteria by broth macrodilution, broth microdilution,
and agar dilution.
A standard for global application developed through the Clinical and Laboratory
Standards Institute consensus process.
M07-A8
ISBN 1-56238-689-1
Volume 29 Number 2 ISSN 0273-3099
Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria
That Grow Aerobically; Approved Standard—Eighth Edition
Matthew A. Wikler, MD, MBA, FIDSA
Franklin R. Cockerill, III, MD
Karen Bush, PhD
Michael N. Dudley, PharmD, FIDSA
George M. Eliopoulos, MD
Dwight J. Hardy, PhD
David W. Hecht, MD
Mary Jane Ferraro, PhD, MPH
Jana M. Swenson, MMSc
Janet F. Hindler, MCLS, MT(ASCP)
Jean B. Patel, PhD, D(ABMM)
Mair Powell, MD, FRCP, FRCPath
John D. Turnidge, MD
Melvin P. Weinstein, MD
Barbara L. Zimmer, PhD
Abstract
Susceptibility testing is indicated for any organism that contributes to an infectious process warranting antimicrobial chemotherapy, if its
susceptibility cannot be reliably predicted from knowledge of the organism’s identity. Susceptibility tests are most often indicated when
the causative organism is thought to belong to a species capable of exhibiting resistance to commonly used antimicrobial agents.
A variety of laboratory methods can be used to measure the in vitro susceptibility of bacteria to antimicrobial agents. This document
describes standard broth dilution (macrodilution and microdilution [the microdilution method described in M07 is the same methodology
outlined in ISO 20776-1])
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and agar dilution techniques, and it includes a series of procedures to standardize the way the tests are
performed. The performance, applications, and limitations of the current CLSI-recommended methods are also described.
The supplemental information (M100 tables) presented with this standard represents the most current information for drug selection,
interpretation, and quality control using the procedures standardized in M07. These tables, as in previous years, have been updated and
should replace tables published in earlier years. Changes in the tables since the previous edition (M100-S18) appear in boldface type and
are also summarized in the front of the document.
Clinical and Laboratory Standards Institute. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically;
Approved Standard—Eighth Edition. CLSI document M07-A 8 (ISBN 1-56238-689-1). Clinical and Laboratory Standards Institute, 940
West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2009.
The Clinical and Laboratory Standards Institute consensus process, which is the mechanism for moving a document through
two or more levels of review by the health care community, is an ongoing process. Users should expect revised editions of any
given document. Because rapid changes in technology may affect the procedures, methods, and protocols in a standard or
guideline, users should replace outdated editions with the current editions of CLSI/NCCLS documents. Current editions are
listed in the CLSI catalog and posted on our website at www.clsi.org. If your organization is not a member and would like to
become one, and to request a copy of the catalog, contact us at: Telephone: 610.688.0100; Fax: 610.688.0700; E-Mail:
customerservice@clsi.org; Website: www.clsi.org
Volume 29
M07-A8
vii
Contents
Abstract .................................................................................................................................................... i
Committee Membership ........................................................................................................................ iii
Foreword ................................................................................................................................................. x
Summary of Major Changes in This Document ..................................................................................... x
Summary of CLSI Processes for Establishing Interpretive Criteria and QC Ranges ......................... xiii
CLSI Reference Methods vs Commercial Methods and CLSI vs FDA Breakpoints
(interpretive criteria) ............................................................................................................................ xiv
Subcommittee on Antimicrobial Susceptibility Testing Mission Statement ........................................ xv
1 Scope .......................................................................................................................................... 1
2 Introduction ................................................................................................................................ 1
3 Standard Precautions .................................................................................................................. 2
4 Terminology ............................................................................................................................... 2
4.1 Definitions .................................................................................................................... 2
4.2 Abbreviations/Acronyms .............................................................................................. 3
5 Indications for Performing Susceptibility Tests......................................................................... 4
6 Selection of Antimicrobial Agents for Routine Testing and Reporting ..................................... 5
6.1 Routine Reports ............................................................................................................ 5
6.2 Nonproprietary Names .................................................................................................. 5
6.3 Selection Guidelines ..................................................................................................... 8
6.4 Suggested Guidelines for Routine and Selective Testing and Reporting ..................... 8
7 Antimicrobial Agents ................................................................................................................. 9
7.1 Source ........................................................................................................................... 9
7.2 Weighing Antimicrobial Powders ................................................................................. 9
7.3 Preparing Stock Solutions ........................................................................................... 10
7.4 Number of Concentrations Tested .............................................................................. 11
8 Inoculum Preparation for Dilution Tests ................................................................................. 11
8.1 Turbidity Standard for Inoculum Preparation ............................................................. 11
8.2 Direct Colony Suspension Method ............................................................................. 11
8.3 Growth Method ........................................................................................................... 11
9 Agar Dilution Procedure .......................................................................................................... 12
9.1 Reagents and Materials ............................................................................................... 12
9.2 Preparing Agar Dilution Plates ................................................................................... 13
9.3 Preparing the Inoculum ............................................................................................... 14
9.4 Inoculating Agar Dilution Plates ................................................................................ 14
9.5 Incubating Agar Dilution Plates.................................................................................. 15
9.6 Determining Agar Dilution End Points ....................................................................... 15
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End of Document
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FAQs of Methods for Dilution Antimicrobial Susceptibility Testing 8th Edition

What are the key methods for antimicrobial susceptibility testing?
The document details three primary methods for antimicrobial susceptibility testing: broth macrodilution, broth microdilution, and agar dilution. Broth macrodilution involves using a series of tubes with varying concentrations of antimicrobial agents, while broth microdilution uses microtiter plates for smaller volumes. Agar dilution involves incorporating antimicrobial agents into agar plates. Each method aims to determine the minimal inhibitory concentration (MIC) for specific bacteria, which is essential for effective treatment decisions.
What is the significance of the M100 tables in this document?
The M100 tables provide critical information regarding antimicrobial drug selection, interpretive criteria, and quality control ranges. These tables are updated annually and are essential for ensuring that laboratories use the most current standards for susceptibility testing. They help clinicians interpret MIC results accurately, guiding appropriate antimicrobial therapy. The latest updates in the M100 tables reflect ongoing research and changes in resistance patterns, making them a vital resource for healthcare professionals.
How does the document address quality control in susceptibility testing?
Quality control is a significant focus in the document, outlining responsibilities for both manufacturers and laboratory personnel. It emphasizes the importance of using quality control strains to validate testing procedures and ensure accuracy. The document specifies the frequency of quality control testing and corrective actions for out-of-control results. By adhering to these quality control measures, laboratories can maintain the reliability of their susceptibility testing, which is crucial for patient safety and effective treatment.
What organisms require special consideration in susceptibility testing?
The document highlights several organisms that require special consideration, including Staphylococci, Enterococci, and fastidious bacteria like Haemophilus influenzae. These organisms may exhibit unique resistance patterns or require specific testing conditions to ensure accurate results. For instance, Staphylococcus aureus may show inducible clindamycin resistance, which necessitates additional testing methods. Understanding these nuances is essential for microbiologists to provide accurate susceptibility profiles and guide appropriate antimicrobial therapy.
What are the limitations of dilution test methods mentioned in the document?
The document outlines several limitations of dilution test methods, including potential misleading results due to various factors such as inoculum preparation and incubation conditions. It notes that some organisms may not grow well under standard testing conditions, leading to inaccurate MIC determinations. Additionally, the emergence of resistance during testing can complicate interpretations. Recognizing these limitations is crucial for microbiologists to ensure they apply the correct methods and interpret results accurately.

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