When we monitored infection of P. PF-01367338 mouse aeruginosa PAO1 in ASM we noticed a 50-fold lower concentration of phage particles. This indicates a reduced efficiency of phage infection by JG024 under simulated chronic infection using the artificial sputum medium. In parallel we tested a P. aeruginosa CF-isolate, strain BT73, for susceptibility to phage infection in LB and ASM. Unexpectedly, we noticed only a 1.9-fold lower phage number in ASM compared to LB (Figure 4). We noticed that phage JG024 was less effective against the CF isolate under both conditions, since approximately tenfold less phage particles were produced under both conditions compared to PAO1. However, while strain BT73 is less susceptible to selleck chemicals llc phage lysis, the

efficiency does not decrease dramatically under ASM growth conditions. Figure 4 Infection assay of JG024 in ASM medium. Phage growth during infection assay in LB medium (dark grey bars) and ASM medium (light grey bars). Changes in phage concentration are described as n-fold. In contrast to the P. aeruginosa PAO1 strain the CF-isolate BT73 is mucoid and secretes

the exopolysaccharide alginate. We wondered if alginate overproduction could explain the observed results. It was recently published that even non-mucoid strains like the wild type PAO1 express the exopolysaccharide alginate in response to oxygen-limiting conditions [33]. We also observed that cultures of PAO1 in ASM, which mimics the CF lung, were highly viscous compared to the cultures in LB medium, suggesting a high production of alginate by the wild type PAO1 in this medium. If alginate is the factor in our experimental setup which decreases phage infection efficiency,

N-acetylglucosamine-1-phosphate transferase a mucoid Selleck Compound C variant of strain PAO1 should show a similar result as the clinical isolate BT73. Therefore, we repeated the phage infection experiments in LB and ASM with a P. aeruginosa mucA mutant strain. We observed again only a 1.6-fold decrease in ASM and an overall approximately tenfold reduction in phage particles when compared with P. aeruginosa PAO1 (Figure 4). These results are in agreement with our hypothesis that alginate overproduction reduces phage infection efficiency. Moreover, they point to alginate as the dominant factor for the decrease in phage infection efficiency in ASM. To verify this result, we performed the same experiment with P. aeruginosa PAO1 in LB medium and increasing alginate concentrations. We chose alginate concentrations of 50, 100, 200, 500 μg/ml up to 1000 μg/ml, since non-mucoid P. aeruginosa strains have been reported to produce 50-200 μg/ml alginate, while mucoid isolates produce up to 1000 μg/ml alginate [34–36]. In accordance with our hypothesis, the presence of alginate reduced phage multiplication in our test assay. A concentration of 50 to 200 μg/ml alginate resulted in an almost 20-fold reduction of phage particles compared to LB medium alone in accordance with the 50-fold reduction of phage particles observed in ASM compared to LB.