The effects of different doses of inhaled bacteriophage therapy for Pseudomonas aeruginosa pulmonary infections in mice

Inhaled phage therapy has been revisited as a potential treatment option for respiratory infections caused by multidrug-resistant Pseudomonas aeruginosa; however, there is a distinct gap in understanding the dose-response effect. The aim of this study was to investigate the dose-response effect of Pseudomonas-targeting phage PEV31 delivered by the pulmonary route in a mouse lung infection model.Neutropenic BALB/c mice were infected with multidrug-resistant P. aeruginosa (2 × 104 colony-forming units) through the intratracheal route and then treated with PEV31 at three different doses of 7.5 × 104 (Group A), 5 × 106 (Group B), and 5 × 108 (Group C) plaque-forming units, or phosphate-buffered saline at 2 hours postinoculation. Mice (n = 5-7) were euthanized at 2 hours and 24 hours postinfection, and lungs, kidneys, spleen, liver, bronchoalveolar lavage fluid, and blood were collected for bacteria and phage enumeration.At 24 hours postinfection, all phage-treated groups exhibited a significant reduction in pulmonary bacterial load by 1.3-1.9 log10, independent of the delivered phage dose. The extent of phage replication was negatively correlated with the dose administered, with log10 titre increases of 6.2, 2.7, and 9 for Groups A, B, and C, respectively. Phage-resistant bacterial subpopulations in the lung homogenate samples harvested at 24 hours postinfection increased with the treatment dose (i.e. 30%, 74%, and 91% in respective Groups A-C). However, the mutants showed increased susceptibility to ciprofloxacin, impaired twitching motility, and reduced blue-green pigment production. The expression of the inflammatory cytokines (IL-1ß and IL-6, and TNF-α) was suppressed with increasing PEV31 treatment dose.This study provides the dose-response effect of inhaled phage therapy that may guide dose selection for treating P. aeruginosa respiratory infections in humans.