Optimization of cryopreservation method for toxin-producing cyanobacteria

Aims:The process of isolating cyanobacteria from various environments and ecosystems can be challenging, consuming a lot of time and laborious.Therefore, to maintain cyanobacterial culture, including toxin-producing cyanobacteria, for a longer period, the best preservation method should be developed and established.This study was conducted to assess the effectiveness of cryopreservation of isolated toxin-producing cyanobacterial strains from aquatic environments using deep freezing with methanol as the cryoprotectant.Methodology and results: Twelve strains of cyanobacteria were isolated from various locations in Malaysia and inoculated in BG11 media supplemented with 0%, 5%, and 10% methanol and kept in -20 °C freezer for one, two and three months.The strains' viability was observed for one month at room temperature after being preserved in the freezer.The evaluated cyanobacteria exhibited different responses to the cryopreservation protocols following their classification and group.Storage in the -20 °C freezer was not suitable for the filamentous cyanobacterial strain, Leptolyngbya frigida ANT.L52B.3,while five out of the twelve strains tested were still viable only when cryopreserved in the presence of methanol.Meanwhile, a total of five strains (Synechococcus sp.EO68, Synechococcus sp.M1, Nodosilinea cf.nodulosa LEGE 10377, Cephalothrix komarekiana SAG 75.79 and Oscillatoria sp.OF9) responded well with methanol, showing high post-thaw viability even after three months of preservation.Conclusion, significance and impact of study: Our results demonstrated that methanol is suitable for preserving most toxin-producing cyanobacterial strains tested in this study, offering a practical alternative to costly and time-consuming maintenance processes while conserving valuable genetic resources.