Susceptibility testing on a QC organism with a new media lot produced zones too large for all antibiotics. Repeating with a previously used lot yielded acceptable zones. The unacceptable zone sizes are best explained by which factor?

Susceptibility testing on a QC organism with a new media lot produced zones too large for all antibiotics. Repeating with a previously used lot yielded acceptable zones. The unacceptable zone sizes are best explained by which factor?

• A. Antibiotic disks were not stored with the proper desiccant
• B. Depth of media was too thick
• C. Depth of media was too thin
• D. Antibiotic disks were not properly applied to the media

Answer: (D)

The key idea is how disk diffusion results depend on the procedure, not just the materials. In Kirby-Bauer testing, zone sizes come from how well the antibiotic diffuses from each disk into the agar and how the organism responds. The technique must be precise: the disks must be placed flat on the surface, in proper contact with the agar, with correct spacing, and the plate must meet standard depth. Here, a new media lot produced zones that were too large for all antibiotics, but re-testing with the previously used lot gave acceptable zones. If the issue were the media depth, you’d expect the abnormal, larger zones to appear with both lots, since diffusion is governed by the agar regardless of the disk. The fact that only the run with the new media lot showed aberrant, uniformly enlarged zones points to a procedural error specific to that run. The most plausible factor is that the antibiotic disks were not properly applied to the media in the new test, leading to anomalous diffusion and larger apparent zones. When proper technique was used with the established media lot, zones returned to normal. Other factors like desiccant issues or disk potency would not explain the run-to-run difference as cleanly; and if the media depth were the problem, the issue would persist across lots.

The key idea is how disk diffusion results depend on the procedure, not just the materials. In Kirby-Bauer testing, zone sizes come from how well the antibiotic diffuses from each disk into the agar and how the organism responds. The technique must be precise: the disks must be placed flat on the surface, in proper contact with the agar, with correct spacing, and the plate must meet standard depth.

Here, a new media lot produced zones that were too large for all antibiotics, but re-testing with the previously used lot gave acceptable zones. If the issue were the media depth, you’d expect the abnormal, larger zones to appear with both lots, since diffusion is governed by the agar regardless of the disk. The fact that only the run with the new media lot showed aberrant, uniformly enlarged zones points to a procedural error specific to that run. The most plausible factor is that the antibiotic disks were not properly applied to the media in the new test, leading to anomalous diffusion and larger apparent zones. When proper technique was used with the established media lot, zones returned to normal.

Other factors like desiccant issues or disk potency would not explain the run-to-run difference as cleanly; and if the media depth were the problem, the issue would persist across lots.