Mapping the Tail Fiber as the Receptor Binding Protein Responsible for Differential Host Specificity of Pseudomonas aeruginosa Bacteriophages PaP1 and JG004

Le, Shuai; He, Xuesong; Tan, Yinling; Huang, Guangtao; Zhang, Lin; Lux, Renate; Shi, Wenyuan; Hu, Fuquan

doi:10.1371/journal.pone.0068562

Cited by 132 publications

(114 citation statements)

References 45 publications

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“…Among these ORFs, ORF060 of phage KPP22 had the lowest protein sequence identity to the homologous ORF of phage KPP12, ORF043, and a dissimilarity was observed particularly in the C-terminal region of the protein. Mutations in the distal portion of tail fibers have been shown to be involved in improvements in adsorption in other phages (33,39), which supports that ORF060 can be a tail fiber protein.…”

Section: Resultsmentioning

confidence: 87%

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Analyses of Short-Term Antagonistic Evolution of Pseudomonas aeruginosa Strain PAO1 and Phage KPP22 (Myoviridae Family, PB1-Like Virus Genus)

Uchiyama

Suzuki

Nishifuji

et al. 2016

Appl Environ Microbiol

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Pseudomonas aeruginosa causes serious intractable infections in humans and animals. Bacteriophage (phage) therapy has been applied to treat P. aeruginosa infections, and phages belonging to the PB1-like virus genus in the Myoviridae family have been used as therapeutic phages. To achieve safer and more effective phage therapy, the use of preadapted phages is proposed. To understand in detail such phage preadaptation, the short-term antagonistic evolution of bacteria and phages should be studied. In this study, the short-term antagonistic evolution of bacteria and PB1-like phage was examined by studying phage-resistant clones of P. aeruginosa strain PAO1 and mutant PB1-like phages that had recovered their infectivity. First, phage KPP22 was isolated and characterized; it was classified as belonging to the PB1-like virus genus in the Myoviridae family. Subsequently, three KPP22-resistant PAO1 clones and three KPP22 mutant phages capable of infecting these clones were isolated in three sets of in vitro experiments. It was shown that the bacterial resistance to phage KPP22 was caused by significant decreases in phage adsorption and that the improved infectivity of KPP22 mutant phages was caused by significant increases in phage adsorption. The KPP22-resistant PAO1 clones and the KPP22 mutant phages were then analyzed genetically. All three KPP22-resistant PAO1 clones, which were deficient for the O5 antigen, had a common nonsense mutation in the wzy gene. All the KPP22 mutant phage genomes showed the same four missense mutations in the open reading frames orf060, orf065, and orf086. The information obtained in this study should be useful for further development of safe and efficient phage therapy. IMPORTANCEPseudomonas aeruginosa causes serious intractable infections in humans and animals; bacteriophage (phage) therapy has been utilized to treat P. aeruginosa infections, and phages that belong to the PB1-like virus genus in the family Myoviridae have been used as therapeutic phages. The preadapted phage is trained in advance through the antagonistic evolution of bacteria and phage and is proposed to be used to achieve safer and more effective phage therapy. In this study, to understand the phage preadaptation, the in vitro short-term antagonistic evolution was studied using P. aeruginosa strain PAO1 and the newly isolated PB1-like phage KPP22. Phage KPP22 was characterized, and the molecular framework regarding the phage preadaptation of KPP22 was elucidated. The importance of study of antagonistic evolution of bacteria and phage in phage therapy is discussed.

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Section: Resultsmentioning

confidence: 87%

“…Some mutant phages increase their infectivity by improving their adsorption efficiency (18,(32)(33)(34). The adsorption efficiencies of the KPP22 mutant phages were compared with that of phage KPP22 (Fig.…”

Section: Resultsmentioning

confidence: 99%

Analyses of Short-Term Antagonistic Evolution of Pseudomonas aeruginosa Strain PAO1 and Phage KPP22 (Myoviridae Family, PB1-Like Virus Genus)

Uchiyama

Suzuki

Nishifuji

et al. 2016

Appl Environ Microbiol

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“…The bacterial host harboring the donor plasmid is then infected by the phage to be engineered. Homologous recombination occurs between the plasmid and the phage genome, allowing the heterologous gene to be integrated into the phage genome and eventually packaged within the phage particle (59,60). However, often only a small fraction of the progeny phage will be recombinant.…”

Section: Techniques For Engineering Synthetic Phages Homologous Recommentioning

confidence: 99%

“…The T3/7 recombinant phage exhibited a broader host range and a better adsorption efficiency than those of either of the wildtype phages, i.e., T3 and T7 (132). Le et al demonstrated that host specificity in P. aeruginosa phages is also tail fiber dependent (60). They first isolated a spontaneous mutant phage that exhibited a broader host range than that of the parental phage, JG004.…”

Section: Engineered Phages With Shifted or Broadened Host Rangesmentioning

confidence: 99%

Genetically Engineered Phages: a Review of Advances over the Last Decade

Pires

Cleto

Sillankorva

et al. 2016

Microbiol Mol Biol Rev

335

275

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SUMMARYSoon after their discovery in the early 20th century, bacteriophages were recognized to have great potential as antimicrobial agents, a potential that has yet to be fully realized. The nascent field of phage therapy was adversely affected by inadequately controlled trials and the discovery of antibiotics. Although the study of phages as anti-infective agents slowed, phages played an important role in the development of molecular biology. In recent years, the increase in multidrug-resistant bacteria has renewed interest in the use of phages as antimicrobial agents. With the wide array of possibilities offered by genetic engineering, these bacterial viruses are being modified to precisely control and detect bacteria and to serve as new sources of antibacterials. In applications that go beyond their antimicrobial activity, phages are also being developed as vehicles for drug delivery and vaccines, as well as for the assembly of new materials. This review highlights advances in techniques used to engineer phages for all of these purposes and discusses existing challenges and opportunities for future work.

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“…The introduction or exchange of tail-associated genes enabled E. coli viruses T2 and T7 to infect a broader range of host bacteria (12)(13)(14). Likewise, the tail fiber gene of Pseudomonas virus PaP1 was replaced by the one of phage JG004, which caused a change in host specificity (15). For detection purposes, reporter phages induce production of an easily traceable enzyme or protein.…”

mentioning

confidence: 99%

Engineering of Bacteriophages Y2:: dpoL1-C and Y2:: luxAB for Efficient Control and Rapid Detection of the Fire Blight Pathogen, Erwinia amylovora

Born

Fieseler

Thöny

et al. 2017

Appl Environ Microbiol

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Erwinia amylovora is the causative agent of fire blight, a devastating plant disease affecting members of the Rosaceae. Alternatives to antibiotics for control of fire blight symptoms and outbreaks are highly desirable, due to increasing drug resistance and tight regulatory restrictions. Moreover, the available diagnostic methods either lack sensitivity, lack speed, or are unable to discriminate between live and dead bacteria. Owing to their extreme biological specificity, bacteriophages are promising alternatives for both aims. In this study, the virulent broad-host-range E. amylovora virus Y2 was engineered to enhance its killing activity and for use as a luciferase reporter phage, respectively. Toward these aims, a depolymerase gene of E. amylovora virus L1 (dpoL1-C) or a bacterial luxAB fusion was introduced into the genome of Y2 by homologous recombination. The genes were placed downstream of the major capsid protein orf68, under the control of the native promoter. The modifications did not affect viability of infectivity of the recombinant viruses. Phage Y2::dpoL1-C demonstrated synergistic activity between the depolymerase degrading the exopolysaccharide capsule and phage infection, which greatly enhanced bacterial killing. It also significantly reduced the ability of E. amylovora to colonize the surface of detached flowers. The reporter phage Y2::luxAB transduced bacterial luciferase into host cells and induced synthesis of large amounts of a LuxAB luciferase fusion. After the addition of aldehyde substrate, bioluminescence could be readily monitored, and this enabled rapid and specific detection of low numbers of viable bacteria, without enrichment, both in vitro and in plant material.IMPORTANCE Fire blight, caused by Erwinia amylovora, is the major threat to global pome fruit production, with high economic losses every year. Bacteriophages represent promising alternatives to not only control the disease, but also for rapid diagnostics. To enhance biocontrol efficacy, we combined the desired properties of two phages, Y2 (broad host range) and L1 (depolymerase for capsule degradation) in a single recombinant phage. This phage showed enhanced biocontrol and could reduce E. amylovora on flowers. Phage Y2 was also genetically engineered into a luciferase reporter phage, which transduces bacterial bioluminescence into infected cells and allows detection of low numbers of viable target bacteria. The combination of speed, sensitivity, and specificity is superior to previously used diagnostic methods. In conclusion, genetic engineering could improve the properties of phage Y2 toward better killing efficacy and sensitive detection of E. amylovora cells.KEYWORDS recombinant phage, reporter, depolymerase, luciferase, bacteriophage, fire blight, reporter phage

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Mapping the Tail Fiber as the Receptor Binding Protein Responsible for Differential Host Specificity of Pseudomonas aeruginosa Bacteriophages PaP1 and JG004

Cited by 132 publications

References 45 publications

Analyses of Short-Term Antagonistic Evolution of Pseudomonas aeruginosa Strain PAO1 and Phage KPP22 (Myoviridae Family, PB1-Like Virus Genus)

Analyses of Short-Term Antagonistic Evolution of Pseudomonas aeruginosa Strain PAO1 and Phage KPP22 (Myoviridae Family, PB1-Like Virus Genus)

Genetically Engineered Phages: a Review of Advances over the Last Decade

Engineering of Bacteriophages Y2:: dpoL1-C and Y2:: luxAB for Efficient Control and Rapid Detection of the Fire Blight Pathogen, Erwinia amylovora

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