(P-QU-12) Quantification of UVC-induced DNA damages in Escherichia coli by UVC treatment of platelet concentrates

Pathogen inactivation (PI) technologies for blood components have been established to reduce the residual risk of known and emerging infectious agents. THERAFLEX UV-Platelets system (Macopharma), a novel PI technology for platelet concentrates (PC), works without photoactive agents by UVC illumination of PC. It is known that the inactivation of pathogens is based on the UVC-induced damage of nucleic acids. According to the literature, the absorption of UVC light produces two predominant types of DNA damage: Cyclobutane pyrimidine dimers (CPD) and pyrimidine (6-4) pyrimidone photoproducts (6-4PP). Aim of the current study was to quantify the amounts of UVC-induced photoproducts in the DNA of Escherichia coli (E. coli) introduced by pathogen inactivation treatment of bacterially contaminated PCs using the UVC-based THERAFLEX UV-Platelets system.

Study

Design/Methods:

Plasma reduced PCs from 4 buffy coats (35% plasma in additive solution SSP+ (Macopharma)) were inoculated with E. coli (PEI-B-P-19) to a final concentration of approx. 10⁶ colony forming units (CFU)/mL. PCs (n=3, 350 mL) were UVC-treated on the MacoTronic UV illumination device (Macopharma) with increasing doses until the full UVC dose of the THERAFLEX -UV Platelet procedure was achieved. Samples were taken for determination of the bacterial concentration by plating on agar plates and for quantification of the amounts of photoproducts by ELISA (OxiSelect UV-induced DNA Damage ELISA Kit, Cell Biolabs).

Results/Findings:

The results are shown in Table 1. E. coli was inactivated by UVC treatment down to the detection limit resulting in a reduction factor of ≥ 6.1 log steps. UVC-induced damage in the DNA of E. coli was monitored by the quantification of CPD and 6-4PP photoproducts. The amount of CPD and 6-4PP increased dose dependently to a final CPD concentration of 32.9 ± 7.4 ng/µg DNA and 6-4PP concentration of 8.2 ± 2.7 ng/µg DNA at 100% UVC dose, respectively.

Conclusions:

The results of this study suggest that the mode of action of the THERAFLEX UV-Platelets system is based on dose-dependent, UVC-mediated alterations in nucleic acids including CPD and 6-4PP photoproducts.