Agar diffusion refers to a class of microbiological assays in which a substance placed on or in a layer of solid agar diffuses into the medium and affects the growth of microorganisms surrounding it. The most common application is measuring the susceptibility of bacteria or fungi to antibiotics by observing zones where growth is inhibited around impregnated discs or wells.
Explanation
In a typical agar diffusion test, a standardized inoculum of a test organism is spread over the surface of a plate containing nutrient agar. Small paper discs impregnated with known concentrations of antibiotics, or wells filled with solutions of antimicrobial agents, are placed on the surface. As the plate is incubated, the compound diffuses radially into the agar, establishing a concentration gradient. If the organism is sensitive to the agent, growth will be suppressed in an area surrounding the source, forming a clear zone. After incubation, the diameter of this zone is measured and compared to interpretive standards to determine whether the organism is susceptible, intermediate or resistant to the antibiotic. Factors such as inoculum density, agar depth, diffusion rate and incubation conditions must be controlled to produce reproducible results.
Uses and Considerations
The disc diffusion method, often called the Kirby–Bauer test, is widely used in clinical laboratories because it is simple and inexpensive and does not require specialized equipment. Variants include the agar well diffusion assay, used to evaluate natural products or chemicals, and assays for enzyme activity or toxin production. While agar diffusion provides qualitative or semi‑quantitative information on antimicrobial activity, it is less precise than broth dilution for determining minimum inhibitory concentrations. Slow‑growing organisms or those requiring special conditions may not be suitable for this method.
Agar diffusion assays have played a key role in monitoring antimicrobial susceptibility and screening new compounds. By observing the relationship between diffusion and microbial inhibition, researchers and clinicians can make informed decisions about treatment and the potential of novel agents.
Related Terms: Kirby–Bauer test, Zone of inhibition, Minimum inhibitory concentration, Antibiotic sensitivity test, Agar well diffusion