Comparative evaluation of long-term preservation methods for morphologically distinct bacteriophages.

The rapid increase in antibiotic resistance has led to a renewed interest in phage therapy, which has created a need for the establishment of phage collections in order to preserve diverse phages and to reduce the delivery time to patients. However, there are currently no standard methods for the long-term preservation of phages. We assessed the stability of four different phages under distinct storage conditions, including different temperatures, storage solutions, concentrations, and with or without cryoprotectant. We found that the type of storage buffer has a significant impact on phage stability, followed by the storage temperature. Phages demonstrated higher viability in lysogeny broth (LB) than saline-magnesium (SM) buffer without gelatin. We also observed a higher sensitivity to freezing in tailed phages with longer tails, such as T4. Ultimately, we found that all four phages maintained high stability after snap freezing, followed by storage at -80°C using LB as a storage buffer without cryoprotectant. IMPORTANCE: Phage therapy, which involves treating bacterial infections using bacteriophages (phage), has shown promise as an alternative to antibiotics and can offer a solution for treating infections caused by antibiotic-resistant bacteria. However, phages are not conventional drugs and can lose their viability when stored under unsuitable conditions. Their high diversity makes finding a standard storage method for long-term preservation challenging. Here, we studied the stability of phages under different storage conditions and identified key factors affecting their viability. We have also identified a specific storage condition that can effectively preserve a wide range of phage morphotypes for over 2 years.