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Updated: Jul 8, 2025
Phage Phenomics: Physiological Approaches to Characterize Novel Viral Proteins
Published on: June 11, 2015
Genome sequence and characterization of a novel Pseudomonas putida phage, MiCath
James Jaryenneh1, Joseph S Schoeniger1, Catherine M Mageeney2
1Sandia National Laboratories, Livermore, CA, 94550, USA.
Abstract:
Pseudomonads are ubiquitous bacteria with importance in medicine, soil, agriculture, and biomanufacturing. We report a novel Pseudomonas putida phage, MiCath, which is the first known phage infecting P. putida S12, a strain increasingly used as a synthetic biology chassis. MiCath was isolated from garden soil under a tomato plant using P. putida S12 as a host and was also found to infect four other P. putida strains. MiCath has a ~ 61 kbp double-stranded DNA genome which encodes 97 predicted open reading frames (ORFs); functions could only be predicted for 48 ORFs using comparative genomics. Functions include structural phage proteins, other common phage proteins (e.g., terminase), a queuosine gene cassette, a cas4 exonuclease, and an endosialidase. Restriction digestion analysis suggests the queuosine gene cassette encodes a pathway capable of modification of guanine residues. When compared to other phage genomes, MiCath shares at most 74% nucleotide identity over 2% of the genome with any sequenced phage. Overall, MiCath is a novel phage with no close relatives, encoding many unique gene products.
Insights
Researchers discovered MiCath, a novel bacteriophage targeting Pseudomonas putida S12, a key bacterium in synthetic biology. This unique phage, isolated from soil, has a large genome with novel gene functions.
Area of Science:
- Microbiology
- Virology
- Synthetic Biology
Background:
- Pseudomonads are widespread bacteria vital in various fields, including medicine, agriculture, and biomanufacturing.
- Pseudomonas putida S12 is increasingly utilized as a chassis in synthetic biology applications.
- Bacteriophages (phages) are viruses that infect bacteria and play crucial roles in microbial ecosystems.
Purpose of the Study:
- To report the isolation and characterization of a novel bacteriophage, MiCath, infecting Pseudomonas putida S12.
- To investigate the genomic features and potential functions encoded by the MiCath phage.
- To assess the novelty of MiCath by comparing its genome to existing phage databases.
Main Methods:
- Isolation of bacteriophage MiCath from garden soil using P. putida S12 as the host.
- Genomic DNA extraction and sequencing of the isolated phage.
- Bioinformatic analysis of the phage genome, including open reading frame (ORF) prediction and comparative genomics.
- Restriction digestion analysis to investigate specific gene functions.
Main Results:
- MiCath was successfully isolated and identified as a novel phage infecting P. putida S12 and four other P. putida strains.
- The phage possesses a double-stranded DNA genome of approximately 61 kbp, encoding 97 predicted ORFs.
- Comparative genomics revealed that MiCath shares minimal nucleotide identity with other sequenced phages, indicating its novelty.
- Unique gene products identified include a queuosine gene cassette, a cas4 exonuclease, and an endosialidase, with evidence suggesting a guanine modification pathway.
Conclusions:
- MiCath represents a novel bacteriophage with a unique genomic makeup and no close relatives among known phages.
- The identified unique gene products, such as the queuosine pathway and endosialidase, offer new avenues for research in phage biology and P. putida engineering.
- This discovery expands the phage toolkit for potential applications in synthetic biology and microbial control involving P. putida strains.
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