An encapsulation-dehydration method for the long-term storage of cyanobacterial strains in the BCCM/ULC collection.

Long-term genetic and functional stability is a fundamental requirement for the maintenance of microorganisms. Cryopreservation is the preferred method for the long-term storage of many micro-organisms, including cyanobacteria. The BCCM/ULC collection currently holds over 200 cyanobacterial strains as living cultures, but only 62 are maintained in a cryo-preserved state. The main limiting factors are the low levels of survival of some strains, as well as the long periods required to recover from cryopreservation, and thus the inability to rapidly deliver strains to clients..
The BRAIN-be project PRESPHOTO (preservation of photosynthetic micro-algae in the BCCM collections) (www.presphoto.ulg.ac.be) aims to improve the preservation of cyanobacterial and diatoms in the BCCM/ULC and BCCM/DCG collections, respec-tively. The encapsulation-dehydration is often considered as a promising alternative to the traditional cryopreservation method for recalcitrant microalgal strains. In this technique, cyanobacterial cultures are entrapped in calcium-alginate beads, osmotically dehydrated (traditionally with sucrose solutions), then evaporatively desiccated in a sterile air flow, or over silica gel and subsequently cryopreserved.
In this study, the encapsulation-dehydration method was evaluated as potential long-term preservation technique of cyanobacterial strains. The effects of several factors on the viability of 4 strains have been investigated to determine which are the most important for the successful cryopreservation. In particular, several cryoprotectants (i.e. sucrose, PEG or mannitol), methods of beads dehydration (i.e. under a laminar flow or with silica gel) were tested. We have also compared storage at -70°C and in liquid nitrogen (-196oC). A pre-cultivation step in KCl was also tested for its ability to enhance desiccation tolerance. In addition, the use of alginate dissolution buffer following thawing was also considered.
In the final phase of the project, the selected cryopreservation protocols will be tested on a large set of strains. In addition, an independent validation of the protocols will be also performed by both partners of the project (BCCM/DCG and the Culture Collection of Algae and Protozoa).