Zones of Inhibition

What is a Zone of Inhibition?

Zones of inhibition are areas where no bacteria can grow around an antibiotic compound. A bacterial 'lawn' most often appears as a light-colored and opaque layer over a darker field of supportive agar jelly. Gaps in this lawn of represent areas where growth has failed.

What do zones mean?

Most commonly associated with antibiotic susceptibility tests such as diffusion disc assays, a zone can appear around different test components depending on whether a component (like a doped filter disc) contains a compound that can prevent bacteria from growing, replicating or simply being alive.

Figure 1 below displays sample images of how a disc can appear on a plate. Only one of the example images represents a recordable, true zone of inhibition. The others are examples of zones where the bacteria is overcoming whatever the disc is doped with.

Four images of zones of inhibition. Image ‘A’ presents the zone with fairly big radius but almost non-transparent. Image ‘B’ presents the zone with big radius and transparency, however within that transparency radius, small spheres of bacterial colonies are present in irregular fashion. Image ‘C’ presents the zone with a very small, almost unnoticeable radius. Image ‘D’ presents the zone with big radius and full transparency.

Figure 1: Examples of zones of inhibition.

Image A: No antimicrobial effect

In this image, we see that although the background lawn of bacteria is slightly lightened, there are many bacteria growing right up to the disc. There is no recordable antimicrobial effect.

Image B: Potential resistance or regrowth

We can see a radius that appears to have been cleared, however, there are several colonies growing within this radius. This could represent colonies of resistant bacteria, or regrowth of bacteria following the decomposition of the antimicrobial agent. Either way, this test would normally be repeated in the lab, as there is no recordable clear zone of inhibition.

Image C: Ambiguous outcome

There does appear to be a zone here, however, the zone is very small and there does appear to be a few colonies growing next to the plate. This ambiguity would lead to a repeat test if certainty was required. There is no clear zone here.

Image D: True zone of inhibition

There is a very clear zone here with only a couple of small irregularities around the very edge of the zone. The bacteria-free area would be measured across its diameter and the distance recorded as the size of the zone.