Optimization of preservation procedures for Arthrospira sp. in the frame of the MELiSSA (Micro-Ecological Life Support System Alternative) project

Probiotic cyanobacteria, such as Arthrospira sp., are daily used as nutritional supplements. They are considered for promising applications beyond Earth, where they can play a crucial role in closed miniaturized biological waste recycling systems, like the MELiSSA project, that is currently developed to support future long-term space missions. Cyanobacteria can be used to remove CO2 from the space cabin, to produce oxygen, to remove nitrogen compounds from waste water and to produce edible biomass. Today, Arthrospira strains are maintained in continuous culture, and new batches are inoculated from previous batches. It is presently unknown how to store Arthro-spira sp. in an inactive but reversal state, even for short period of time. Because the several years length of a mission to Mars, particular strain conservation and inoculation strategy to start new cultures have to be developed for space applications. In this study cell integrity and cultivability after various preservation procedures have been assessed by optical density- and pH-measurements, microscopy and flow cytometry. These procedures included preservation in Zar-rouk agar medium, alginate and in presence of clay, all in different storage conditions and over different periods of time. Also, drying on filters, lyophilization and stepwise freezing with cryopro-tectants were assessed. In the results obtained from these experiments, it was observed that preservation in presence of clay was the most successful as an alternative to preservation in Zar-rouk liquid medium. Also preservation in agar could be promising on the condition that a lower concentration of agar is used. Cryopreservation with a mixture of DMSO and trehalose and drying in presence of clay after induced trehalose accumulation also showed to be promising applica-tions.