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Isolation and characterization of fMGyn-Pae01, a phiKZ-like jumbo phage infecting Pseudomonas aeruginosa
Kira Ranta1,2, Mikael Skurnik2, Saija Kiljunen3
1HUS Diagnostic Center, Clinical Microbiology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland.
Background:
Pseudomonas aeruginosa is an opportunistic pathogen that causes a wide variety of infections, and belongs to the group of ESKAPE pathogens that are the leading cause of healthcare-associated infections and have high level of antibiotic resistance. The treatment of infections caused by antibiotic-resistant P. aeruginosa is challenging, which makes it a common target for phage therapy. The successful utilization of phage therapy requires a collection of well characterized phages.
Methods:
Phage fMGyn-Pae01 was isolated from a commercial phage therapy cocktail. The phage morphology was studied by transmission electron microscopy and the host range was analyzed with a liquid culture method. The phage genome was sequenced and characterized, and the genome was compared to closest phage genomes. Phage resistant bacterial mutants were isolated and whole genome sequencing and motility, phage adsorption and biofilm formation assays were performed to the mutants and host bacterium.
Results:
The genomic analysis revealed that fMGyn-Pae01 is a lytic, phiKZ-like jumbo phage with genome size of 277.8 kb. No genes associated with lysogeny, bacterial virulence, or antibiotic resistance were identified. Phage fMGyn-Pae01 did not reduce biofilm formation of P. aeruginosa, suggesting that it may not be an optimal phage to be used in monophage therapy in conditions where biofilm formation is expected. Host range screening revealed that fMGyn-Pae01 has a wide host range among P. aeruginosa strains and its infection was not dependent on O-serotype. Whole genome sequencing of the host bacterium and phage resistant mutants revealed that the mutations had inactivated either a flagellar or rpoN gene, thereby preventing the biosynthesis of a functional flagellum. The lack of functional flagella was confirmed in motility assays. Additionally, fMGyn-Pae01 failed to adsorb on non-motile mutants indicating that the bacterial flagellum is the phage-binding receptor.
Conclusion:
fMGyn-Pae01 is a phiKZ-like jumbo phage infecting P. aeruginosa. fMGyn-Pae01 uses the flagellum as its phage-binding receptor, supporting earlier suggestions that flagellum might be utilized by phiKZ but differs from some other previous findings showing that phiKZ-like phages use the type-IV pili as the phage-binding receptor.
Insights
Phage fMGyn-Pae01, a lytic jumbo phage, infects Pseudomonas aeruginosa by binding to its flagellum. This phage shows broad host range but does not inhibit biofilm formation, impacting its use in phage therapy.
Area of Science:
- Microbiology
- Virology
- Bacteriology
Background:
- Pseudomonas aeruginosa is a leading cause of healthcare-associated infections.
- This opportunistic pathogen exhibits high antibiotic resistance, complicating treatment.
- Phage therapy is a promising alternative for combating resistant bacterial infections.
Purpose of the Study:
- To characterize phage fMGyn-Pae01 for potential therapeutic applications.
- To investigate the interaction between phage fMGyn-Pae01 and Pseudomonas aeruginosa.
- To identify the bacterial receptor used by fMGyn-Pae01 for infection.
Main Methods:
- Isolation and characterization of phage fMGyn-Pae01 from a commercial cocktail.
- Transmission electron microscopy for morphology analysis.
- Genome sequencing and comparative analysis.
- Host range determination and biofilm formation assays.
- Isolation and whole genome sequencing of phage-resistant mutants.
- Motility assays to confirm flagellar function.
Main Results:
- fMGyn-Pae01 is a lytic, phiKZ-like jumbo phage with a 277.8 kb genome.
- No virulence or antibiotic resistance genes were detected in the phage genome.
- The phage did not inhibit Pseudomonas aeruginosa biofilm formation.
- fMGyn-Pae01 demonstrated a wide host range, independent of O-serotype.
- Phage resistance in mutants was linked to inactivated flagellar or rpoN genes.
- The bacterial flagellum was identified as the phage-binding receptor.
Conclusions:
- Phage fMGyn-Pae01 is a phiKZ-like jumbo phage with potential for Pseudomonas aeruginosa infection treatment.
- The bacterial flagellum serves as the primary receptor for fMGyn-Pae01.
- The phage's inability to inhibit biofilm formation may limit its monotherapy application.
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