Can bacteriophage be stabilised by lipid encapsulation when nebulised for inhalation delivery against Pseudomonas aeruginosa?

Inhaled bacteriophage (phage) therapy is emerging as a promising approach to combat multidrug-resistant (MDR) respiratory pathogens such as Pseudomonas aeruginosa. Aerosol delivery by nebulization poses challenges for maintaining phage stability, often resulting in titer losses due to mechanical stresses. This study evaluated the use of liposomal encapsulation to protect phages during nebulization. Two P. aeruginosa phages, PEV2 (short-tail) and PEV40 (long-tail), were selected for this work. Liposomes were prepared using DSPC, cholesterol, Tween 80, and cationic lipid DOTAP. Encapsulation efficiencies were 78 % for PEV2 and 90 % for PEV40, with mean particle sizes of 300 nm and 650 nm, respectively. Nebulization by jet and vibrating mesh devices showed that the liposome-encapsulated phages were able to preserve viability, with titer losses below 0.4 log (PEV40) and 0.07 log (PEV2). In contrast, non-encapsulated phages experienced titer reductions of up to 1.23 log, especially by jet nebulization. Vibrating mesh nebulization generated slightly larger droplets (∼5.6 µm) but with better phage recovery (> 90 %) and respirable fractions (> 70 %) for both types of phages encapsulated in liposomes. These results demonstrate that the approach of lipid encapsulation effectively protects phages from mechanical damage during nebulization, maintaining bioactivity for aerosol delivery to enhance the success of inhaled phage therapy.