Development of a Lyophilization Process for Bacteriophage Storage and Transport.

Bacteriophages are a sustainable alternative to control pathogenic bacteria in the post-antibiotic era. Despite promising reports, there are still obstacles to phage use, notably titer stability and transport-associated expenses for applications in food and agriculture. In this study, we have developed a lyophilization approach to maintain phage titers, ensure efficacy and reduce transport costs of bacteriophages. Lyophilization methods were adopted with various excipients to enhance stabilization in combination with packaging options for international transport. Lyophilization of CP30A using tryptone formed a cake that limited processing titer reduction to 0.35 ± 0.09 log PFU mL. Transmission electron microscopy revealed the initial titer reduction was associated with capsid collapse of a subpopulation. Freeze-dried phages were generally stable under refrigerated vacuum conditions and showed no significant titer changes over 3 months incubation at 4 °C ( = 0.29). Reduced stability was observed for lyophilized phages that were incubated either at 30 °C under vacuum or at 4 °C at 70% or 90% relative humidity. Refrigerated international transport and rehydration of the cake resulted in a total phage titer reduction of 0.81 ± 0.44 log PFU mL. A significantly higher titer loss was observed for phages that were not refrigerated during transport (2.03 ± 0.32 log PFU mL). We propose that lyophilization offers a convenient method to preserve and transport phages, with minimal titer reduction after the drying process.