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Characterization of JG024, a pseudomonas aeruginosa PB1-like broad host range phage under simulated infection conditions.

Garbe J, Wesche A, Bunk B, Kazmierczak M, Selezska K, Rohde C, Sikorski J, Rohde M, Jahn D, Schobert M - BMC Microbiol. (2010)

Bottom Line: The receptor of phage JG024 was determined as lipopolysaccharide.Alginate production was identified as a factor reducing phage infectivity.Phage JG024 is a suitable broad host range phage which could be used in phage therapy.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Microbiology, Technische Universit├Ąt Braunschweig, Spielmannstr, 7, 38106 Braunschweig, Germany.

ABSTRACT

Background: Pseudomonas aeruginosa causes lung infections in patients suffering from the genetic disorder Cystic Fibrosis (CF). Once a chronic lung infection is established, P. aeruginosa cannot be eradicated by antibiotic treatment. Phage therapy is an alternative to treat these chronic P. aeruginosa infections. However, little is known about the factors which influence phage infection of P. aeruginosa under infection conditions and suitable broad host range phages.

Results: We isolated and characterized a phage, named JG024, which infects a broad range of clinical and environmental P. aeruginosa strains. Sequencing of the phage genome revealed that the phage JG024 is highly related to the ubiquitous and conserved PB1-like phages. The receptor of phage JG024 was determined as lipopolysaccharide. We used an artificial sputum medium to study phage infection under conditions similar to a chronic lung infection. Alginate production was identified as a factor reducing phage infectivity.

Conclusions: Phage JG024 is a suitable broad host range phage which could be used in phage therapy. Phage infection experiments under simulated chronic lung infection conditions showed that alginate production reduces phage infection efficiency.

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Related in: MedlinePlus

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.
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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.

Mentions: Since phage JG024 is able to infect 84% of the tested clinical isolates in vitro we were interested if this phage is able to infect P. aeruginosa under simulated CF lung conditions. An artificial sputum medium (ASM) was used to mimic the CF lung environment. Growth in ASM leads to formation of typical biofilm-like microcolonies of P. aeruginosa and supports other phenotypic changes observed under chronic infection conditions [12]. At first, we tested the ability of phage JG024 to lyse the non-mucoid wild type strain P. aeruginosa PAO1 in ASM compared to LB medium. As described in Methods, we monitored phage particles and noted an increase of phage particles by a factor of nearly 500 000 in LB and in ASM by a factor of 10 000 (Figure 4). This indicates cell lysis by phage JG024 under these non ideal conditions (note that PAO1 was in stationary phase prior to infection to allow comparison to ASM). When we monitored infection of P. 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 phage lysis, the efficiency does not decrease dramatically under ASM growth conditions.


Characterization of JG024, a pseudomonas aeruginosa PB1-like broad host range phage under simulated infection conditions.

Garbe J, Wesche A, Bunk B, Kazmierczak M, Selezska K, Rohde C, Sikorski J, Rohde M, Jahn D, Schobert M - BMC Microbiol. (2010)

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.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3008698&req=5

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.
Mentions: Since phage JG024 is able to infect 84% of the tested clinical isolates in vitro we were interested if this phage is able to infect P. aeruginosa under simulated CF lung conditions. An artificial sputum medium (ASM) was used to mimic the CF lung environment. Growth in ASM leads to formation of typical biofilm-like microcolonies of P. aeruginosa and supports other phenotypic changes observed under chronic infection conditions [12]. At first, we tested the ability of phage JG024 to lyse the non-mucoid wild type strain P. aeruginosa PAO1 in ASM compared to LB medium. As described in Methods, we monitored phage particles and noted an increase of phage particles by a factor of nearly 500 000 in LB and in ASM by a factor of 10 000 (Figure 4). This indicates cell lysis by phage JG024 under these non ideal conditions (note that PAO1 was in stationary phase prior to infection to allow comparison to ASM). When we monitored infection of P. 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 phage lysis, the efficiency does not decrease dramatically under ASM growth conditions.

Bottom Line: The receptor of phage JG024 was determined as lipopolysaccharide.Alginate production was identified as a factor reducing phage infectivity.Phage JG024 is a suitable broad host range phage which could be used in phage therapy.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Microbiology, Technische Universit├Ąt Braunschweig, Spielmannstr, 7, 38106 Braunschweig, Germany.

ABSTRACT

Background: Pseudomonas aeruginosa causes lung infections in patients suffering from the genetic disorder Cystic Fibrosis (CF). Once a chronic lung infection is established, P. aeruginosa cannot be eradicated by antibiotic treatment. Phage therapy is an alternative to treat these chronic P. aeruginosa infections. However, little is known about the factors which influence phage infection of P. aeruginosa under infection conditions and suitable broad host range phages.

Results: We isolated and characterized a phage, named JG024, which infects a broad range of clinical and environmental P. aeruginosa strains. Sequencing of the phage genome revealed that the phage JG024 is highly related to the ubiquitous and conserved PB1-like phages. The receptor of phage JG024 was determined as lipopolysaccharide. We used an artificial sputum medium to study phage infection under conditions similar to a chronic lung infection. Alginate production was identified as a factor reducing phage infectivity.

Conclusions: Phage JG024 is a suitable broad host range phage which could be used in phage therapy. Phage infection experiments under simulated chronic lung infection conditions showed that alginate production reduces phage infection efficiency.

Show MeSH
Related in: MedlinePlus