Abstract
Purpose Corneas are released with ongoing microbiological controls due to their short shelf life. For microbiological tests using automated culture systems, the European guideline (EDQM, 5th edition, 2022) is based on the method described in European Pharmacopoeia. Although the guide recommends 7 to 14 days of cultivation, it is not clear enough when these longer cultivation times are necessary. Our goal is to analyze different microbiological times to guarantee a reliable microbiological result.
Methods 1131 liquid samples from organ-culture medium during corneal storage were analysed for microbial contaminations between March to August 2023 with BACT/ALERT® 3D system. A blood culture BACT/ALERT iF Plus anaerobic and BACT/ALERT iF Plus aerobic was taken from each sample. All microbiological samples were cultured until day 14. In addition, visual inspection of every sample for turbidity was performed on the last day.
Microbiologic tests were performed on the preservation medium on day 6 after the start of corneal culture (n=1131) and after transfer to the deswelling medium (n=1054). Microbiologic testing was also performed 1 day after final evaluaton (n=789) and or the day after any manipulation (n=428) for DSAEK or DMEK.
Results The percentage of corneal contamination was 5.41%. Where 44% were yeasts and filamentous fungi, 27% were aerobic gram-positive cocci, 20% aerobic gram-negative bacilli, 7% strict anaerobes and 2% mycobacteria.
81.7% of the samples were positive before the fifth day. While 18.3% of them tested positive around the 14th. On average, the samples were positive after 2.99 days. The analytical method validation studies are also supportive with these data and equivalent growth times are observed.
We also observed a slight increase of contamination during summer season (June to August) in comparison with spring season (March to May), 1.83 ± 0.76% vs 1.37 ±0.45% respectively.
Conclusions Based on the results and in accordance with the European guideline, moreover the measures already established by most eye banks (such as adequate decontamination and sampling of the conjunctiva or sclera at the recovery time or even storage of empty preservation media), it would also be advisable to analyse samples up to day 14 to ensure a reliable microbiological result. It would even be advisable to continue joining efforts to develop rapid microbiology methods that allow the release the corneal tissue with definitive microbiological results.
