Phage phiKZ-The First of Giants.

The paper covers the history of the discovery and description of phiKZ, the first known giant bacteriophage active on Pseudomonas aeruginosa. It also describes its unique features, especially the characteristic manner of DNA packing in the head around a cylinder-shaped structure ("inner body"), which probably governs an ordered and tight packaging of the phage genome. Important properties of phiKZ-like phages include a wide range of lytic activity and the blue opalescence of their negative colonies, and provide a background for the search and discovery of new P. aeruginosa giant phages. The importance of the phiKZ species and of other giant phage species in practical phage therapy is noted given their broad use in commercial phage preparations.

Transposition Behavior Revealed by High-Resolution Description of Pseudomonas Aeruginosa Saltovirus Integration Sites.

Transposable phages, also called saltoviruses, of which the Escherichia coli phage Mu is the reference, are temperate phages that multiply their genome through replicative transposition at multiple sites in their host chromosome. The viral genome is packaged together with host DNA at both ends. In the present work, genome sequencing of three Pseudomonas aeruginosa transposable phages, HW12, 2P1, and Ab30, incidentally gave us access to the location of thousands of replicative integration sites and revealed the existence of a variable number of hotspots. Taking advantage of deep sequencing, we then designed an experiment to study 13,000,000 transposon integration sites of bacteriophage Ab30. The investigation revealed the presence of 42 transposition hotspots adjacent to bacterial interspersed mosaic elements (BIME) accounting for 5% of all transposition sites. The rest of the sites appeared widely distributed with the exception of coldspots associated with low G-C content segments, including the putative O-antigen biosynthesis cluster. Surprisingly, 0.4% of the transposition events occurred in a copy of the phage genome itself, indicating that the previously described immunity against such events is slightly leaky. This observation allowed drawing an image of the phage chromosome supercoiling into four loops.

Modular Approach to Select Bacteriophages Targeting Pseudomonas aeruginosa for Their Application to Children Suffering With Cystic Fibrosis.

This review discusses the potential application of bacterial viruses (phage therapy) toward the eradication of antibiotic resistant Pseudomonas aeruginosa in children with cystic fibrosis (CF). In this regard, several potential relationships between bacteria and their bacteriophages are considered. The most important aspect that must be addressed with respect to phage therapy of bacterial infections in the lungs of CF patients is in ensuring the continuity of treatment in light of the continual occurrence of resistant bacteria. This depends on the ability to rapidly select phages exhibiting an enhanced spectrum of lytic activity among several well-studied phage groups of proven safety. We propose a modular based approach, utilizing both mono-species and hetero-species phage mixtures. With an approach involving the visual recognition of characteristics exhibited by phages of well-studied phage groups on lawns of the standard P. aeruginosa PAO1 strain, the simple and rapid enhancement of the lytic spectrum of cocktails is permitted, allowing the development of tailored preparations for patients capable of circumventing problems associated with phage resistant bacterial mutants.

Complete Genome Sequence of PM105, a New Pseudomonas aeruginosa B3-Like Transposable Phage.

The complete genome of the Pseudomonas aeruginosa bacteriophage PM105 is 39,593 bp long. The phage belongs to the B3 family of transposable Mu-like phages, as confirmed by the presence of bacterial DNA joined to the phage genome ends. PM105, together with other B3-like phages, form a newly arising species.

Myoviridae bacteriophages of Pseudomonas aeruginosa: a long and complex evolutionary pathway.

Recently we have accomplished the entire DNA sequence of bacteriophage phiKZ, a giant virus infecting Pseudomonas aeruginosa. The 280334-bp of phiKZ genome is a linear, circularly permutated and terminally redundant, AT-rich dsDNA molecule that contains no sites for NotI, PstI, SacI, SmaI, XhoI and XmaIII endonucleases. Limited homology to other bacteriophages on the DNA and protein levels indicated that phiKZ represents a distinct branch of the Myoviridae family. In this work, we analyzed a group of six P. aeruginosa phages (Lin68, Lin21, PTB80, NN, EL, and RU), which are morphologically similar to phiKZ, have similar genome size and low G+C content. All phages have a broad host range among P. aeruginosa strains, and they are resistant to the inhibitory action of many P. aeruginosa plasmids. The analysis of the genomic DNA by restriction enzymes and DNA-DNA hybridization shows that phages are representative of three phiKZ-like species: phiKZ-type (phiKZ, Lin21, NN and PTB80), EL-type (EL and RU) and Lin68 which has a shorter tail than other phages. Except for related phages EL and RU, all phiKZ-like phages have identical N-terminal amino acid sequences of the major capsid protein. Random genome sequencing shows that the EL and RU phages have no homology to the phiKZ-like phages on DNA level. We propose that the phiKZ, Lin21, NN, PTB80 and Lin68 phages can be included in a new phiKZ genus, and that the EL and RU phages belong to a separate genus within the Myoviridae family. Based on the resistance to many restriction enzymes and the transduction ability, there are indications that over the long pathway of evolution, the phiKZ-like phages probably inherited the capacity to infect different bacterial species.

The genome of bacteriophage phiKZ of Pseudomonas aeruginosa.

Bacteriophage phiKZ is a giant virus that efficiently infects Pseudomonas aeruginosa strains pathogenic to human and, therefore, it is attractive for phage therapy. We present here the complete phiKZ genome sequence and a preliminary analysis of its genome structure. The 280,334 bp genome is a linear, circularly permutated and terminally redundant, A+T-rich double-stranded DNA molecule. The phiKZ DNA has no detectable sequence homology to other viruses and microorganisms, and it does not contain NotI, PstI, SacI, SmaI, XhoI, and XmaIII endonuclease restriction sites. The genome has 306 open reading frames (ORFs) varying in size from 50 to 2237 amino acid residues. According to the orientation of transcription, ORFs are apparently organized into clusters and most have a clockwise direction. The phiKZ genome also encodes six tRNAs specific for Met (AUG), Asn (AAC), Asp (GAC), Leu (TTA), Thr (ACA), and Pro (CCA). A putative promoter sequence containing a TATATTAC block was identified. Most potential stem-loop transcription terminators contain the tetranucleotide UUCG loops. Some genes may be assigned as phage-encoded RNA polymerase subunits. Only 59 phiKZ gene products exhibit similarity to proteins of known function from a diversity of organisms. Most of these conserved gene products, such as dihydrofolate reductase, ribonucleoside diphosphate reductase, thymidylate synthase, thymidylate kinase, and deoxycytidine triphosphate deaminase are involved in nucleotide metabolism. However, no virus-encoded DNA polymerase, DNA replication-associated proteins, or single-stranded DNA-binding protein were found based on amino acid homology, and they may therefore be strongly divergent from known homologous proteins. Fifteen phiKZ gene products show homology to proteins of pathogenic organisms, including Mycobacterium tuberculosis, Haemophilus influenzae, Listeria sp., Rickettsia prowazakeri, and Vibrio cholerae that must be considered before using this phage as a therapeutic agent. The phiKZ coat contains at least 40 polypeptides, and several proteins are cleaved during virus assembly in a way similar to phage T4. Eleven phiKZ-encoded polypeptides are related to proteins of other bacteriphages that infect a variety of hosts. Among these are four gene products that contain a putative intron-encoded endonuclease harboring the H-N-H motif common to many double-stranded DNA phages. These observations provide evidence that phages infecting diverse hosts have had access to a common genetic pool. However, limited homology on the DNA and protein levels indicates that bacteriophage phiKZ represents an evolutionary distinctive branch of the Myoviridae family.