Pseudomonas aeruginosa Keratitis in Mice: Effects of Topical Bacteriophage KPP12 Administration

Fukuda, Ken; Ishida, Waka; Uchiyama, Jumpei; Rashel, Mohammad; Kato, Shin‐ichiro; Morita, Tatsuhito; Muraoka, Asako; Sumi, Tamaki; Matsuzaki, Shigenobu; Daibata, Masanori; Fukushima, Atsuki

doi:10.1371/journal.pone.0047742

Cited by 82 publications

(70 citation statements)

References 27 publications

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“…Today, we are globally threatened by antibiotic-resistant bacteria, and phage therapy is being reassessed as a new strategy for treatment of infectious disease (13,17). Indeed, researchers have reported the efficacy of phage therapy for experimental bacterial disease (18)(19)(20)(21)(22). Examples include applications for use of the phages ListShield (Intralytix, Inc., Baltimore, MD) for Listeria monocytogenes and EcoShield (Intralytix) for Escherichia coli O157:H7 to protect against foodborne disease (23,24).…”

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confidence: 99%

Phage Therapy Is Effective in a Mouse Model of Bacterial Equine Keratitis

Furusawa

Iwano

Hiyashimizu

et al. 2016

Appl Environ Microbiol

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Bacterial keratitis of the horse is mainly caused by staphylococci, streptococci, and pseudomonads. Of these bacteria, Pseudomonas aeruginosa sometimes causes rapid corneal corruption and, in some cases, blindness. Antimicrobial resistance can make treatment very difficult. Therefore, new strategies to control bacterial infection are required. A bacteriophage (phage) is a virus that specifically infects and kills bacteria. Since phage often can lyse antibiotic-resistant bacteria because the killing mechanism is different, we examined the use of phage to treat horse bacterial keratitis. We isolated Myoviridae or Podoviridae phages, which together have a broad host range. They adsorb efficiently to host bacteria; more than 80% of the ⌽R18 phage were adsorbed to host cells after 30 s. In our keratitis mouse model, the administration of phage within 3 h also could kill bacteria and suppress keratitis. A phage multiplicity of infection of 100 times the host bacterial number could kill host bacteria effectively. A cocktail of two phages suppressed bacteria in the keratitis model mouse. These data demonstrated that the phages in this study could completely prevent the keratitis caused by P. aeruginosa in a keratitis mouse model. Furthermore, these results suggest that phage may be a more effective prophylaxis for horse keratitis than the current preventive use of antibiotics. Such treatment may reduce the use of antibiotics and therefore antibiotic resistance. Further studies are required to assess phage therapy as a candidate for treatment of horse keratitis. IMPORTANCEAntibiotic-resistant bacteria are emerging all over the world. Bacteriophages have great potential for resolution of this problem. A bacteriophage, or phage, is a virus that infects bacteria specifically. As a novel therapeutic strategy against racehorse keratitis caused by Pseudomonas aeruginosa, we propose the application of phages for treatment. Phages isolated in this work had in vitro effectiveness for a broad range of P. aeruginosa strains. Indeed, a great reduction of bacterial proliferation was shown in phage therapy for mouse models of P. aeruginosa keratitis. Therefore, to reduce antibiotic usage, phage therapy should be investigated and developed further.

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mentioning

confidence: 99%

Phage Therapy Is Effective in a Mouse Model of Bacterial Equine Keratitis

Furusawa

Iwano

Hiyashimizu

et al. 2016

Appl Environ Microbiol

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“…The P. aeruginosa strains used in this study, which had been isolated from the patients in Kochi, Japan, and have been described previously, are listed in Table 1 (14,21). Phage KPP22 was isolated using strain PA33 as the host by a method described elsewhere (22).…”

Section: Methodsmentioning

confidence: 99%

“…After centrifugation (10,000 ϫ g, 40 min, 4°C), the phage pellet was dissolved in TM buffer (10 mM Tris-HCl [pH 7.2], 5 mM MgCl 2 ) containing 100 g ml Ϫ1 of both DNase I and RNase A. After incubation at 37°C for 30 min, the phage suspension was purified by cesium chloride density gradient ultracentrifugation (100,000 ϫ g, 1 h, 4°C), as described elsewhere (14). The phage band was collected and dialyzed against AAS (0.1 M ammonium acetate, 10 mM NaCl, 1 mM CaCl 2 , 1 mM MgCl 2 , pH 7.2) for 1 h at 4°C.…”

Section: Methodsmentioning

confidence: 99%

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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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“…The potential of natural lytic phages to act as antimicrobial agents against targets that include multidrug-resistant bacteria has been studied extensively both in vitro (88)(89)(90)(91) and in vivo (92)(93)(94)(95)(96)(97)(98). Natural phages have been used therapeutically since 1919, when d'Hérelle successfully treated children suffering from severe dysentery with phages (9).…”

Section: Phage Engineering For Pathogen Control Natural Phage-based Amentioning

confidence: 99%

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

Pires

Cleto

Sillankorva

et al. 2016

Microbiol Mol Biol Rev

325

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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Pseudomonas aeruginosa Keratitis in Mice: Effects of Topical Bacteriophage KPP12 Administration

Cited by 82 publications

References 27 publications

Phage Therapy Is Effective in a Mouse Model of Bacterial Equine Keratitis

Phage Therapy Is Effective in a Mouse Model of Bacterial Equine Keratitis

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

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