Antifungal Susceptibility Testing of Yeasts and Molds

By Dr. Zubair Khalid, DVM, MS, PhD · 2026-08-27

Quick Answer

• Clinical laboratories choose between CLSI reference broth dilution methods and commercial systems such as VITEK 2 and Sensititre YeastOne, balancing accuracy against cost and workflow demands.
• Reference methods provide the most reliable minimum inhibitory concentration results but require skilled personnel and extended incubation, while commercial systems offer faster turnaround with some performance tradeoffs.
• No single method fits every laboratory, so decisions must weigh organism type, testing volume, available expertise, and the need for results that support clinical decisions.

At a Glance

Antifungal susceptibility testing (AST) determines the minimum inhibitory concentration (MIC) of an antifungal drug against a yeast or mold isolate. The Clinical and Laboratory Standards Institute (CLSI) publishes reference methods that define how testing should be performed, and commercial systems adapt these principles for routine laboratory use. The table below compares the main testing approaches available to clinical laboratories.

Each method has a place in laboratory practice. The choice depends on the laboratory's testing volume, the range of organisms encountered, the need for rapid results, and the available budget for equipment and consumables.

The Clinical Need for Antifungal Susceptibility Testing

Antimicrobial resistance is a global public health concern. The World Health Organization defines antimicrobial resistance as the ability of microorganisms to resist the drugs used to treat infections, and it emphasizes that resistance threatens the effective prevention and treatment of an ever-increasing range of infections. Antifungal resistance is a subset of this broader problem, and it has become more important as the population of immunocompromised patients has grown.

Invasive fungal infections are difficult to diagnose and treat. They occur most often in patients with weakened immune systems, including those undergoing chemotherapy, organ transplant recipients, and patients in intensive care units. The most common causes of invasive fungal disease are Candida species and Aspergillus species. Candida auris is a particularly concerning emerging pathogen because it is often multidrug resistant and can spread in healthcare settings.

Antifungal susceptibility testing serves several purposes. It helps clinicians choose the most appropriate drug for a patient with a fungal infection. It also supports surveillance programs that track resistance patterns in hospitals and communities. When resistance is detected, testing results can guide stewardship efforts to preserve the effectiveness of existing drugs.

The CLSI develops and maintains standards for antimicrobial susceptibility testing. These standards describe how to perform testing, how to interpret results, and how to ensure quality. The CLSI microbiology standards are used by laboratories around the world to produce reliable and reproducible results. Laboratories that follow these standards can compare their results with those of other laboratories and with clinical breakpoints established by expert committees.

The Biology of Antifungal Resistance

Fungal organisms develop resistance to antifungal drugs through several mechanisms. Understanding these mechanisms helps laboratory professionals interpret susceptibility testing results and anticipate resistance patterns.

Target Site Alteration

Many antifungal drugs work by binding to a specific target in the fungal cell. For example, azole drugs such as fluconazole and voriconazole inhibit the enzyme lanosterol 14-alpha-demethylase, which is involved in ergosterol synthesis. Ergosterol is an essential component of the fungal cell membrane. Mutations in the gene that encodes this enzyme can change the shape of the enzyme so that the drug no longer binds effectively. This is a common mechanism of azole resistance in Candida species and in Aspergillus fumigatus.

Efflux Pump Overexpression

Fungal cells can pump antifungal drugs out of the cell before they reach their target. Efflux pumps are membrane proteins that recognize a range of substrates, including azole drugs. Overexpression of these pumps reduces the intracellular concentration of the drug, allowing the fungus to survive at higher drug concentrations. This mechanism is important in Candida albicans and other Candida species.

Biofilm Formation

Many fungi can form biofilms on medical devices such as catheters and prosthetic valves. Biofilms are communities of cells attached to a surface and embedded in a matrix of extracellular material. Cells within a biofilm are often more resistant to antifungal drugs than free-floating cells. This is because the matrix can slow drug penetration, and the cells within the biofilm may be in a slow-growing state that makes them less susceptible to drugs that target actively growing cells.

Whole Chromosome Changes

Some fungi can develop resistance through changes in chromosome number or structure. For example, the formation of an isochromosome that carries extra copies of genes involved in drug resistance can increase the expression of those genes. This mechanism has been described in Candida albicans and can lead to high-level fluconazole resistance.

CLSI Reference Methods for Antifungal Susceptibility Testing

The CLSI has published reference methods for antifungal susceptibility testing of yeasts and molds. These methods are the basis for most commercial systems and are used to establish breakpoints for interpreting MIC results.

Broth Microdilution for Yeast

The CLSI reference method for yeast is a broth microdilution method. The method uses a standardized inoculum of the test organism, a defined growth medium, and a range of drug concentrations. The test is performed in a 96-well microtiter plate, with each well containing a different concentration of the antifungal drug. After incubation, the MIC is read as the lowest drug concentration that inhibits visible growth.

The method requires careful attention to detail. The inoculum must be prepared from a fresh culture, and the density must be adjusted to a standard turbidity. The medium must be prepared according to the CLSI specification, and the plates must be incubated at the correct temperature for the correct time. For most yeast, the incubation time is 24 hours, but some species may require 48 hours.

Broth Microdilution for Molds

The CLSI reference method for molds is also a broth microdilution method, but it has some differences from the yeast method. Mold inocula are prepared from sporulating cultures, and the inoculum is adjusted to a standard concentration of spores. The medium is the same as that used for yeast, but the incubation time is longer. Most molds require 48 hours of incubation, and some may require up to 72 hours.

The endpoint for mold testing is different from that for yeast. For molds, the MIC is often defined as the lowest concentration that produces a significant reduction in growth compared with the growth control. The definition of significant reduction can vary depending on the drug and the organism. For example, the endpoint for the azole drugs is often a 50 percent reduction in growth, while the endpoint for amphotericin B is often a 100 percent reduction.

Quality Control for Reference Methods

Quality control is essential for any susceptibility testing method. The CLSI standards specify the use of quality control strains with known MIC ranges. These strains are tested alongside clinical isolates to ensure that the test is performing correctly. If the quality control strain produces a result outside the expected range, the test is invalid and must be repeated.

The American Type Culture Collection (ATCC) maintains authenticated strains that are used for quality control. These strains are characterized and tested to ensure that they produce consistent results. Laboratories should use authenticated strains from a recognized culture collection to ensure the validity of their quality control program.

Commercial Systems for Antifungal Susceptibility Testing

Commercial systems offer laboratories a way to perform antifungal susceptibility testing with less manual effort and faster turnaround times. Two of the most widely used systems are the VITEK 2 and the Sensititre YeastOne.

VITEK 2

The VITEK 2 is an automated system that performs identification and susceptibility testing for bacteria and yeast. The system uses a card that contains a series of wells with different drugs and concentrations. The card is inoculated with a standardized suspension of the organism, and the system monitors growth in each well over time. The results are interpreted automatically, and the system reports MIC values for each drug.

The VITEK 2 antifungal susceptibility card is designed for testing yeast. It includes a panel of antifungal drugs that are commonly used to treat candidiasis. The system is fully automated, which reduces the hands-on time required for testing. Results are typically available in 30 hours or less, which is faster than the reference method.

The VITEK 2 system has some limitations. It is designed for yeast and does not cover molds. The drug panel is fixed, so laboratories cannot customize the drugs that are tested. The system requires a significant capital investment, and the cards are relatively expensive. Laboratories that do not have a VITEK 2 system for bacterial testing may not find it cost-effective to purchase one solely for antifungal testing.

Sensititre YeastOne

The Sensititre YeastOne is a broth microdilution system that uses a colorimetric endpoint. The system uses a 96-well plate that is pre-coated with a range of antifungal drugs. The plate is inoculated with a standardized suspension of the organism, and the plate is incubated for 24 to 48 hours. The wells contain a color indicator that changes color when the organism grows. The MIC is determined by the lowest concentration that shows no color change.

The Sensititre YeastOne system covers a broad range of antifungal drugs, including azoles, echinocandins, and amphotericin B. It can be used for both yeast and molds. The system is less automated than the VITEK 2, but it does not require a large capital investment. The plates are relatively inexpensive, and the system can be used in laboratories that do not have a full automation platform.

The Sensititre YeastOne system has been shown to have good agreement with the CLSI reference method for many drug-organism combinations. However, the agreement can vary depending on the drug and the organism. Laboratories should verify the performance of the system for the organisms and drugs that they test.

Comparing Reference Methods and Commercial Systems

The choice between reference methods and commercial systems depends on several factors. The following table summarizes the key differences.

Accuracy and Agreement

The CLSI reference methods are the gold standard for antifungal susceptibility testing. They are used to establish breakpoints and to evaluate the performance of commercial systems. Commercial systems are expected to have a high level of agreement with the reference method, but the agreement is not always perfect.

The agreement between a commercial system and the reference method is usually expressed as the percentage of results that are within one doubling dilution of the reference MIC. A doubling dilution is a two-fold change in the drug concentration. For example, if the reference MIC is 2 micrograms per milliliter, a result of 1 or 4 micrograms per milliliter would be within one doubling dilution.

The acceptable level of agreement depends on the drug and the organism. For some drugs, a high level of agreement is expected. For others, the agreement may be lower, and the laboratory may need to use the reference method for certain organisms or drugs.

Categorical Agreement and Errors

In addition to the MIC value, the clinical interpretation of the result is important. The MIC is interpreted as susceptible, intermediate, or resistant based on clinical breakpoints. The breakpoints are established by the CLSI and are based on a combination of pharmacokinetic and pharmacodynamic data, clinical outcomes, and the distribution of MIC values in the population.

The categorical agreement is the percentage of results that are in the same category as the reference method. A very major error occurs when a commercial system reports a resistant result as susceptible. A major error occurs when a commercial system reports a susceptible result as resistant. A minor error occurs when the result is in a different category but not a major error.

The acceptable rates of errors are defined by the CLSI and other organizations. Laboratories should monitor the performance of their commercial systems and investigate any results that are outside the expected range.

Practical Implementation of Antifungal Susceptibility Testing

Implementing antifungal susceptibility testing in a clinical laboratory requires careful planning. The following steps can help laboratories choose and implement a method that meets their needs.

Step 1: Assess the Laboratory Workload

The first step is to assess the number of antifungal susceptibility tests that the laboratory expects to perform. Laboratories that test only a few isolates per month may find that the reference method is sufficient. Laboratories that test many isolates may benefit from a commercial system that reduces the hands-on time.

The workload also affects the choice of system. The VITEK 2 is most cost-effective when the laboratory already has a VITEK 2 system for bacterial testing. The Sensititre YeastOne is a good option for laboratories that do not have a VITEK 2 system.

Step 2: Determine the Organism and Drug Coverage

The laboratory must determine which organisms and drugs it needs to test. The most common organisms are Candida species and Aspergillus species. The most common drugs are fluconazole, voriconazole, amphotericin B, and the echinocandins.

The VITEK 2 covers yeast only, while the Sensititre YeastOne covers both yeast and molds. If the laboratory tests molds, the Sensititre YeastOne or the reference method is required.

Step 3: Evaluate the Cost

The cost of antifungal susceptibility testing includes the cost of reagents, the cost of labor, and the cost of equipment. The reference method has a low reagent cost but a high labor cost. The VITEK 2 has a high reagent cost but a low labor cost. The Sensititre YeastOne is in between.

The laboratory should calculate the total cost per test for each method, including the cost of quality control and the cost of repeat testing. The cost of repeat testing can be significant if the method has a high rate of errors.

Step 4: Consider the Turnaround Time

The turnaround time is the time from the receipt of the isolate to the reporting of the result. The reference method takes 24 to 72 hours. The VITEK 2 can provide results in 24 hours or less. The Sensititre YeastOne takes 24 to 48 hours.

The turnaround time is important for patient care. A faster result can help the clinician choose the right drug sooner. However, the laboratory must balance the need for speed with the need for accuracy.

Step 5: Train the Staff

The laboratory staff must be trained to perform the testing correctly. The training should include the preparation of the inoculum, the use of the equipment, and the interpretation of the results. The training should also include the use of quality control strains and the handling of errors.

The American Society for Microbiology provides educational resources for laboratory professionals. These resources can be used to train staff and to stay current with the latest developments in the field.

Step 6: Establish a Quality Control Program

A quality control program is essential for the accuracy of the testing. The program should include the use of quality control strains, the monitoring of the results, and the investigation of any results that are outside the expected range.

The quality control program should be documented, and the results should be reviewed regularly. The laboratory should have a process for investigating and correcting any problems that are identified.

Records and Measurements in Antifungal Susceptibility Testing

Accurate records are essential for the quality of the testing and for the interpretation of the results. The following records should be maintained for each test.

Patient and Isolate Information

The laboratory should record the patient identifier, the date of the collection, the source of the specimen, and the organism that was isolated. This information is used to link the result to the patient and to track the epidemiology of the infections.

Test Method and Conditions

The laboratory should record the test method that was used, the drug that was tested, and the MIC value that was obtained. The laboratory should also record the incubation time and temperature, the quality control results, and any deviations from the standard procedure.

Quality Control Results

The quality control results should be recorded for each batch of tests. The results should be compared with the expected range for the quality control strain. If the result is outside the expected range, the test is invalid and must be repeated.

Interpretation and Reporting

The laboratory should record the interpretation of the result, including the category (susceptible, intermediate, or resistant) and the clinical breakpoint that was used. The result should be reported to the clinician in a timely manner.

Common Failure Patterns in Antifungal Susceptibility Testing

Laboratories can encounter several common problems when performing antifungal susceptibility testing. Understanding these problems can help the laboratory avoid them.

Inoculum Preparation Errors

The inoculum is the suspension of the organism that is used to inoculate the test. The inoculum must be prepared to a standard concentration. If the inoculum is too high, the MIC will be too high. If the inoculum is too low, the MIC will be too low.

The inoculum is prepared by adjusting the turbidity of the suspension to match a standard. The turbidity is measured using a spectrophotometer or a McFarland standard. The laboratory should verify the turbidity of the inoculum before use.

Medium Preparation Errors

The medium is the growth medium that is used for the test. The medium must be prepared according to the CLSI specification. If the medium is not prepared correctly, the results may be inaccurate.

The medium should be checked for the correct pH and for the correct concentration of the components. The medium should be stored correctly and used before the expiration date.

Incubation Errors

The incubation time and temperature are important for the test. The incubation time is 24 to 48 hours for yeast and 48 to 72 hours for molds. The incubation temperature is 35 degrees Celsius for most organisms.

If the incubation time is too short, the growth may not be sufficient for the result to be read. If the incubation time is too long, the growth may be too heavy, and the MIC may be too high.

Reading Errors

The reading of the MIC is a subjective step. The laboratory professional must determine the lowest concentration that inhibits the growth of the organism. The reading can be affected by the growth pattern of the organism and the type of the drug.

The reading should be performed by a trained professional who is familiar with the method. The reading should be done in a consistent manner, and the results should be recorded immediately.

Quality Control Failures

Quality control failures occur when the quality control strain produces a result outside the expected range. This can be caused by a problem with the inoculum, the medium, the incubation, or the reading. The laboratory should investigate the cause of the failure and correct the problem before testing patient samples.

Biosafety and Laboratory Safety

Antifungal susceptibility testing involves the handling of live fungal cultures. These cultures can be hazardous to laboratory workers, especially if the organism is a mold that produces spores. The laboratory must follow the biosafety guidelines for the handling of these organisms.

The Centers for Disease Control and Prevention (CDC) and the National Institutes of Health (NIH) publish the Biosafety in Microbiological and Biomedical Laboratories (BMBL) manual. This manual provides guidance on the risk assessment, the containment, and the decontamination of biological materials. The laboratory should follow the BMBL guidelines for the handling of fungal organisms.

Risk Assessment

The laboratory should perform a risk assessment for the organisms that it handles. The risk assessment should consider the pathogenicity of the organism, the route of transmission, and the laboratory procedures that are used. The risk assessment should be used to determine the biosafety level for the laboratory.

Most yeast species are handled at biosafety level 2. Some molds, such as Aspergillus species, are also handled at biosafety level 2. The laboratory should follow the BMBL guidelines for the biosafety level that is appropriate for the organisms that it handles.

Containment

The laboratory should use the appropriate containment equipment to prevent the release of the organisms. This includes the use of a biological safety cabinet for the handling of the organisms. The biological safety cabinet should be certified and maintained according to the manufacturer's instructions.

Decontamination

The laboratory should decontaminate the work surfaces and the equipment after the testing is complete. The decontamination should be done using an appropriate disinfectant. The waste should be disposed of according to the laboratory's waste management plan.

The Role of Antifungal Susceptibility Testing in Stewardship

Antifungal susceptibility testing is an important component of antimicrobial stewardship. Stewardship programs aim to optimize the use of antimicrobial drugs to improve patient outcomes and reduce the development of resistance.

The World Health Organization emphasizes the importance of stewardship in the fight against antimicrobial resistance. The WHO recommends that countries develop national action plans to address antimicrobial resistance, including the surveillance of resistance and the promotion of the appropriate use of antimicrobial drugs.

The laboratory can support stewardship by providing accurate and timely susceptibility results. The results can be used to guide the choice of the drug and the dose. The results can also be used to identify resistance patterns and to monitor the effectiveness of the stewardship program.

Limitations of Antifungal Susceptibility Testing

Antifungal susceptibility testing has several limitations that the laboratory and the clinician should be aware of.

Correlation with Clinical Outcome

The MIC value is a measure of the activity of the drug against the organism in the laboratory. The MIC does not always correlate with the clinical outcome. The clinical outcome depends on many factors, including the immune status of the patient, the site of the infection, and the pharmacokinetics of the drug.

The breakpoints are established to predict the clinical outcome, but the correlation is not perfect. The clinician should use the MIC value in the context of the patient's clinical condition.

The Complexity of the Organism

Some organisms are difficult to test. Molds are more difficult to test than yeast because they grow more slowly and the inoculum is more difficult to prepare. The results for molds are also more difficult to read because the endpoint is less clear.

The Cost of the Testing

The cost of the testing can be a barrier to the use of the testing. The cost of the commercial systems can be high, and the reference method requires a significant amount of labor. The laboratory should consider the cost of the testing when deciding which method to use.

The Availability of the Testing

The availability of the testing can be a barrier to the use of the testing. Some laboratories do not have the equipment or the expertise to perform the testing. The laboratory should consider the availability of the testing when deciding which method to use.

Professional Escalation Criteria

The laboratory should have a process for escalating the results that are unusual or that require a clinical decision. The following are the criteria for the escalation.

Unusual Resistance Patterns

If the laboratory detects a resistance pattern that is unusual for the organism, the laboratory should escalate the result to the clinician. The unusual resistance pattern may indicate a problem with the testing or a new resistance mechanism.

The Need for a Clinical Decision

If the result is in the intermediate or resistant category, the laboratory should escalate the result to the clinician. The clinician may need to change the drug or the dose. The laboratory should provide the clinician with the MIC value and the interpretation.

The Need for a Confirmatory Test

If the result is unexpected or if the quality control fails, the laboratory should repeat the test. If the repeat test produces the same result, the laboratory should escalate the result to the clinician.

The Future of Antifungal Susceptibility Testing

The field of antifungal susceptibility testing is evolving. New methods are being developed, and the existing methods are being improved. The laboratory should stay current with the developments in the field.

Molecular Methods

Molecular methods are being developed for the detection of resistance. These methods can detect the genes that are responsible for the resistance. The molecular methods can be faster than the phenotypic methods, but they are not yet widely available.

The Use of the Whole Genome Sequencing

Whole genome sequencing is being used to understand the mechanisms of resistance. The sequencing can identify the mutations that are responsible for the resistance. The sequencing can also be used to track the spread of the resistant organisms.

The Development of the New Drugs

New antifungal drugs are being developed. The new drugs may have a different mechanism of action and may be active against the resistant organisms. The laboratory should be prepared to test the new drugs.

A Practical Decision Framework for Selecting an Antifungal AST Method

Choosing between CLSI reference methods and commercial systems requires a structured approach that goes beyond simple cost comparisons. The following framework helps laboratory directors and clinical microbiologists make a defensible decision based on their specific operational context.

Step 1: Define the Testing Population

The first decision point is the patient population served by the laboratory. A laboratory supporting a hematology-oncology unit will have different requirements than one serving a general hospital or an outpatient clinic. The laboratory should document the expected annual volume of antifungal susceptibility tests, the predominant Candida species encountered, and whether Aspergillus or other molds are part of the testing menu.

The World Health Organization emphasizes that antimicrobial resistance surveillance depends on reliable laboratory data. Laboratories that participate in regional or national surveillance programs may need to use a method that produces results comparable to reference methods. This requirement can influence the choice of system because some commercial methods have documented agreement levels that are acceptable for clinical use but may not be suitable for research or surveillance purposes.

Step 2: Score the Method Against Laboratory Capabilities

A scoring matrix helps laboratories compare methods against their specific constraints. The matrix should include the following criteria with a weight assigned to each based on local priorities.

Each laboratory assigns a score from 1 to 5 for each criterion, multiplies by the weight, and sums the total. The method with the highest total score is the best fit for that laboratory. This scoring approach is transparent and can be documented for the laboratory's quality management system.

Step 3: Verify Performance Before Implementation

Before adopting a commercial system, the laboratory should verify its performance against the reference method. The verification plan should include a minimum of 20 to 30 clinical isolates for each drug-organism combination that the laboratory intends to test. The isolates should include organisms with known resistance mechanisms and quality control strains from a recognized culture collection such as the American Type Culture Collection.

The verification should measure the essential agreement, which is the percentage of results within one doubling dilution of the reference MIC. The laboratory should also calculate the categorical agreement and the rates of very major errors, major errors, and minor errors. The acceptable error rates are defined by the CLSI and should be reviewed before the verification begins.

Step 4: Establish a Method Change Protocol

When the laboratory changes from one method to another, the change must be documented and communicated to the clinical staff. The laboratory should notify the clinicians of the change, the date of the change, and any differences in the drug panel or the interpretation. The laboratory should also review the historical results to identify any changes in the resistance patterns that could be due to the method change instead of a true change in the organism population.

The laboratory should maintain a record of the method change in the quality management system. The record should include the reason for the change, the verification data, and the approval of the laboratory director.

Step 5: Use a Decision Tree for Isolate Testing

A decision tree can help the laboratory determine which method to use for each isolate. The decision tree should consider the organism, the source of the specimen, and the clinical urgency.

For a yeast isolate from a sterile site in a critically ill patient, the laboratory may choose the fastest method available. For a yeast isolate from a non-sterile site in a stable patient, the laboratory may use the reference method or a commercial method with a longer turnaround time. For a mold isolate, the laboratory must use a method that covers molds, which excludes the VITEK 2.

The decision tree should be documented and reviewed regularly. The laboratory should also document the reasons for the choice of the method for each isolate.

Step 6: Integrate the Method into the Laboratory Workflow

The chosen method must fit into the laboratory workflow. The laboratory should consider the time of day when the isolates are received, the availability of the staff, and the need for the results. The laboratory should also consider the integration of the method with the laboratory information system.

The laboratory should develop a standard operating procedure for the method. The procedure should include the preparation of the inoculum, the inoculation of the test, the incubation, the reading of the results, and the reporting of the results. The procedure should also include the quality control and the troubleshooting.

Step 7: Review the Decision Annually

The laboratory should review the choice of the method annually. The review should consider the changes in the workload, the changes in the organism distribution, the changes in the drug panel, and the changes in the cost. The review should also consider the performance of the method, including the error rates and the quality control failures.

The annual review should be documented and approved by the laboratory director. The review should be used to make any changes to the method or the workflow.

Records and Measurements for the Decision Framework

The laboratory should maintain the following records for the decision framework.

Verification Records

The verification records should include the isolates that were tested, the results of the reference method, the results of the commercial method, and the calculations of the agreement and the errors. The records should also include the quality control results and the interpretation of the results.

Monitoring Records

The monitoring records should include the results of the quality control testing, the results of the proficiency testing, and the results of the internal audits. The records should also include the investigation of any errors or failures.

Annual Review Records

The annual review records should include the data that was reviewed, the decision that was made, and the approval of the decision. The records should also include the implementation of the decision.

Common Failure Patterns in the Decision Process

The decision process can fail in several ways. The laboratory may choose a method based on the cost alone, without considering the accuracy or the workflow. The laboratory may not verify the method before the implementation. The laboratory may not monitor the performance of the method after the implementation. The laboratory may not review the decision annually.

The laboratory should be aware of these failure patterns and should take steps to avoid them. The decision framework described above can help the laboratory make a sound decision and avoid the common pitfalls.

Professional Escalation Criteria for the Decision

The laboratory should escalate the decision to the laboratory director or the medical director in the following situations.

When the Verification Fails

If the verification of the commercial method produces an unacceptable level of agreement or an unacceptable error rate, the laboratory should escalate the issue to the laboratory director. The director should decide whether to continue the verification, to modify the method, or to choose a different method.

When the Quality Control Fails

If the quality control results are outside the expected range, the laboratory should escalate the issue to the laboratory director. The director should investigate the cause of the failure and decide whether to repeat the test, to adjust the procedure, or to stop the testing.

When the Annual Review Identifies a Problem

If the annual review identifies a problem with the method, the laboratory should escalate the issue to the laboratory director. The director should decide whether to change the method, to modify the workflow, or to take other corrective action.

Frequently Asked Questions

What is the difference between the CLSI reference method and a commercial system?

The CLSI reference method is a standardized broth microdilution method that is used to determine the MIC of an antifungal drug. A commercial system such as the VITEK 2 or the Sensititre YeastOne is a product that is designed to perform the testing with less labor and faster turnaround. The commercial systems are evaluated against the reference method to ensure that they produce accurate results.

How long does antifungal susceptibility testing take?

The turnaround time depends on the method. The CLSI reference method takes 24 to 48 hours for yeast and 48 to 72 hours for molds. The VITEK 2 can provide results in 24 hours or less. The Sensititre YeastOne takes 24 to 48 hours.

What is a minimum inhibitory concentration?

The minimum inhibitory concentration is the lowest concentration of a drug that inhibits the visible growth of the organism. The MIC is expressed in micrograms per milliliter. The MIC is used to interpret the susceptibility of the organism to the drug.

What is a clinical breakpoint?

A clinical breakpoint is the MIC value that is used to classify the organism as susceptible, intermediate, or resistant. The breakpoints are established by the CLSI and are based on the clinical outcomes, the pharmacokinetics of the drug, and the distribution of the MIC values in the population.

What is the difference between a yeast and a mold?

A yeast is a fungus that grows as a single cell. A mold is a fungus that grows as a multicellular filament. The testing of the yeast and the mold is different because the inoculum preparation and the endpoint are different.

What is the role of the quality control in the antifungal susceptibility testing?

The quality control is used to ensure that the testing is performing correctly. The quality control strain is a strain with a known MIC range. The quality control strain is tested with the clinical isolates. If the quality control strain produces a result outside the expected range, the test is invalid and must be repeated.

What is the role of the laboratory in the antimicrobial stewardship?

The laboratory provides the results that are used to guide the use of the antimicrobial drugs. The laboratory can also provide the surveillance data that is used to monitor the resistance patterns. The laboratory is an important part of the antimicrobial stewardship program.

What is the biosafety level for the antifungal susceptibility testing?

The biosafety level for the antifungal susceptibility testing depends on the organism that is being tested. Most yeast and molds are handled at biosafety level 2. The laboratory should follow the BMBL guidelines for the biosafety level that is appropriate for the organisms.

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References and Further Reading

• Biosafety in Microbiological and Biomedical Laboratories. CDC and NIH.
• Microbiology Resources. American Society for Microbiology.
• Antimicrobial Resistance. World Health Organization.
• Bacteriological Analytical Manual. U.S. Food and Drug Administration.
• Antimicrobial Susceptibility Testing. Clinical and Laboratory Standards Institute.
• Culture Collections. American Type Culture Collection.
• NCBI Pathogen Detection. National Center for Biotechnology Information.
• Antifungal Susceptibility Testing: Current Approaches.. Clinical microbiology reviews, 2020.
• Antifungal susceptibility testing: applicability of methods and strategies for improving access in resource-constrained settings.. The Lancet. Infectious diseases, 2024.
• Current status of antifungal susceptibility testing methods.. Medical mycology, 2007.
• Antifungal susceptibility testing.. Clinical microbiology reviews, 1993.

This article is educational and does not replace validated laboratory procedures, clinical diagnosis, institutional biosafety review, or public-health guidance.