Eye Bank microbiological analyses: antibiotic residues versus antibiotic elimination

Purpose: aim of the study was to compare the direct inoculum method of corneal preservation media in the presence of antibiotics with the same method after elimination of antibiotics with the RESEP device.

Methods: 42 samples of cold storage media (EUSOL-C, AL.CHI.MI.A. S.r.l.) were obtained from the Eye Bank of Rome after donor cornea storage for up to 7 days. Aliquots of 3 ml of the media were either inoculated directly in trypton soy broth and fluid thioglycollate medium or treated with RESEP to eliminate antibiotic residues and then inoculated in growth media. 10 samples of sterile broth, treated with RESEP and tested by direct inoculum method, served as controls. To evaluate the initial contamination, bioburden was performed for each of the tested EUSOL-C by membrane filtration (MILLIFLEX SENSOR II – MILLIPORE). Positive samples were identified genetically.

Results: 19 out of 42 tested samples (45.2%) resulted contaminated when assessed by the direct inoculum method without RESEP treatment. The same samples treated with RESEP showed 25 of 42 (59.5%) positive samples. The RESEP positive samples, which were not detected without antibiotic elimination, were contaminated with Staphylococcus spp, Bacillus spp, Propionibacterium acnes. Turbidity was detected up to 5 days in advance using RESEP. None of the control samples was contaminated.

Conclusions: The elimination of antibiotic residues from samples allowed to reveal an additional 14% of contaminated samples as compared to the direct inoculum method without elimination of antibiotics. Our study demonstrated that the antibiotics present in the corneal preservation media may interfere with the microbial growth during microbiological analyses by direct inoculum method and lead to false negatives.