Validation of a device for elimination of antibiotics from liquid samples undergoing microbiological analysis

Abstract: Our previous studies indicated that the use of antibiotic cocktails during tissue processing can lead to an antibiotic carry-over effect, which in turn can generate false negative results in microbiological analysis.

Purpose: To validate a single-use device for elimination of antibiotics from liquid samples undergoing microbiological analysis to prevent interference with microbial growth.

Methods: The efficacy of the ResEP device was tested on 5 ml of BASE.128 (AL.CHI.MI.A.S.r.l.) and determined by HPLC after 5, 10, 20, 30 min. The recovery of S. Aureus, P. Aeruginosa, C. Albicans, B. Subtilis, A. Niger and C. Sporogenes was determined by dilution plating of inoculants containing 1-10, 10-100 and 100-1000 CFU, 20 min after treatment.

The device was further tested by three cardiovascular and skin Italian banks. Standard bacteriological tests were performed on transport, rinsing and cryopreservation liquids (BACT-ALERT Biomerieux or thioglycollate/TSB media) of twenty tissue samples decontaminated with BASE.128; additional bacteriological tests were performed on liquid samples using ResEP device, after antibiotic removal. The presence of antibiotic residues in all samples undergoing the microbiological analyses was evaluated by HPLC.

Results: The HPLC analysis of the samples treated with ResEP device showed complete antibiotic removal from the BASE.128 within 20 min of treatment. A total bacterial recovery was obtained for all investigated inoculants.
HPLC analysis on processing liquids showed that important antibiotic residues were present in rinsing and cryopreservation liquids, after decontamination of skin and cardiovascular tissues. The ResEP device removed efficiently antibiotic residues from all liquid samples prior to microbiological analysis allowing the detection of 10 % of false negative results.

Conclusion: The ResEP device was validated for easy and quick removal of antibiotic residues from liquid samples undergoing microbiological analysis, showing complete microbial recovery and high specificity for BASE.128 medium and allowing the detection of false negative results in microbiological analyses.