Antimicrobial Susceptibility Testing (AST) is a laboratory procedure that determines how effectively specific antimicrobial agents, such as antibiotics, can inhibit the growth of a microorganism causing an infection. AST is performed in medical laboratories, often using culture methods where bacteria are exposed to antimicrobials to observe their effects.
The Purpose of Antimicrobial Susceptibility Testing
The primary purpose of antimicrobial susceptibility testing is to guide healthcare providers in making informed decisions about treating bacterial infections. When a patient presents with an infection, clinicians often start with broad-spectrum antibiotics, which target a wide range of bacteria, as they await test results. AST helps transition from this initial broad approach to a more focused, targeted therapy once the specific microorganism and its vulnerabilities are known.
This personalization of treatment can significantly improve patient outcomes by ensuring the most effective drug is used, thereby reducing the duration of illness and potential side effects. For instance, AST can differentiate between common Staphylococcus aureus and more challenging methicillin-resistant Staphylococcus aureus (MRSA), guiding appropriate antibiotic selection. Choosing the correct antibiotic at the outset also plays a role in preventing the development and spread of antibiotic resistance, a growing global health concern.
Common Methods for Testing
Antimicrobial susceptibility testing involves various laboratory methods to observe how bacteria react to different antimicrobial agents. The techniques commonly employed provide either qualitative results, indicating resistance presence, or quantitative results, such as the minimum inhibitory concentration.
Disk Diffusion (Kirby-Bauer test)
The Disk Diffusion test, also known as the Kirby-Bauer method, is a widely used technique. In this method, a standardized amount of bacteria from a patient’s sample is evenly spread across the surface of a special agar plate, typically Mueller-Hinton agar, to create a uniform “lawn” of growth. Small paper disks, each impregnated with a specific concentration of a different antibiotic, are then placed onto the inoculated agar surface. The antibiotic diffuses outwards from each disk into the agar, creating a concentration gradient.
After an incubation period, usually 18-24 hours at 37°C, the plate is examined for zones of inhibition. A zone of inhibition is a clear, circular area around an antibiotic disk where bacterial growth has been prevented. The diameter of this clear zone is measured in millimeters, and its size correlates with the bacteria’s susceptibility to that particular antibiotic; a larger zone indicates greater susceptibility.
Broth Dilution (MIC testing)
Broth Dilution is considered a gold standard for phenotypic susceptibility testing, providing a quantitative measure called the Minimum Inhibitory Concentration (MIC). This method involves preparing a series of tubes or wells, typically in a microtiter plate, each containing a liquid growth medium with progressively lower concentrations of a specific antibiotic. A standardized bacterial suspension is then added to each well.
After incubation, 16-20 hours at 35-37°C, each well is examined for visible bacterial growth, indicated by turbidity or cloudiness. The MIC is determined as the lowest concentration of the antibiotic that completely inhibits visible bacterial growth. This numerical MIC value provides a precise measure of the antibiotic’s potency against the tested microorganism.
Automated Systems
Modern clinical laboratories use automated systems to perform antimicrobial susceptibility testing more rapidly and efficiently. These sophisticated machines can automate various steps of the testing process, including inoculum preparation, antibiotic dispensing, incubation, and result reading. Automated systems utilize technologies based on broth microdilution or gradient diffusion methods, reading changes in turbidity or optical density to determine MIC values.
Examples of such systems include Vitek 2, Phoenix, and Microscan WalkAway Plus, which can process multiple samples simultaneously and provide results within 4 to 18 hours. They enhance laboratory efficiency by reducing manual labor and turnaround times, and some can even offer advanced expert systems to interpret complex resistance patterns.
Interpreting Test Results
After antimicrobial susceptibility tests are performed, the results are interpreted to inform treatment decisions. The outcomes are categorized into three main groups, based on established clinical breakpoints: Susceptible (S), Intermediate (I), and Resistant (R). These categories indicate the likelihood of therapeutic success with a given antimicrobial agent.
A “Susceptible” (S) result means there is a high likelihood that the infection will respond to treatment with the antibiotic at a standard dose. This indicates that the concentration of the antibiotic achievable in the body at the site of infection will be sufficient to inhibit or kill the bacteria. Clinicians prefer to use antibiotics falling into this category when possible.
An “Intermediate” (I) result suggests that the antibiotic may be effective, but only under specific circumstances. This category requires careful clinical judgment, considering factors like the infection’s severity and the patient’s overall health.
A “Resistant” (R) result indicates a high likelihood of therapeutic failure, even if increased doses of the antibiotic are used. This means the bacteria are unlikely to be inhibited or killed by the antibiotic, and alternative treatment options should be explored. Doctors consider these results alongside the patient’s medical history, the specific infection site, and other clinical factors to determine the most appropriate course of action.
The Role in Monitoring Antibiotic Resistance
Antimicrobial susceptibility testing extends beyond individual patient care to play a broader role in public health surveillance. The data collected from these tests are important for monitoring the emergence and spread of antibiotic resistance. Hospitals and public health organizations aggregate and analyze AST results from many patients over time.
This cumulative data is used to create an “antibiogram,” which is a summary chart illustrating the collective susceptibility patterns of local bacterial isolates to various antimicrobial drugs. An antibiogram shows the percentage of tested organisms that are susceptible to a particular antibiotic, compiled annually. For example, it might show that 99% of Escherichia coli isolates are susceptible to a certain drug, while only 50% are susceptible to another.
Antibiograms are important tools for tracking shifts in resistance patterns within a healthcare facility or region, helping to identify the rise of “superbugs”. This information informs hospital treatment guidelines and local antibiotic prescribing policies, guiding empirical therapy when specific culture results are not yet available. Public health agencies use this population-level data to develop strategies for combating widespread antibiotic resistance and promoting responsible antibiotic use.