Generation of reactive oxygen species upon red light exposure of cyanobacteria from Roman hypogea

Subterranean archaeological sites in Rome (Italy) are threatened by phototrophic biofilms predominated by cyanobacteria and associated microorganisms. They damage the frescoes, mortar, marble, and tufa rock wherever artificial lighting is installed. During the past two decades, the conservation strategies have evolved gradually; rather than restricting the illumination time and intensity, the latest approach is to use strong light to reduce their growth. Since cyanobacterial cells are abundant in phycobilisomes and chlorophyll a, which produce reactive oxygen species (ROS) upon irradiation, strong red light (620–650 nm) was applied to generate high amount of ROS in a rate beyond the quenching capacity of the organism. After 25 h of irradiation, the photosystem II quantum yields of seven cyanobacterial isolates in culture were reduced by 65–94%. Conversely, blue light (460–480 nm) promoted photosystem II activity by up to 35%. δ-Aminolevulinic acid (D-ALA) was introduced to enhance the treatment, as it can be transformed into protochlorophyllide by cyanobacteria and then excited by red light to generate ROS inside the cells. Since the natural photosynthetic pigments as well as the endogenous protochlorophyllide exist only within the cyanobacterial cells, they are unlikely to contaminate or damage the underlying stone substrata. Electron spin resonance spectroscopy confirmed that D-ALA treatment caused the formation of ROS; spin trap experiments indicated that radicals were produced in the system.