Improved Adsorption of an Enterococcus faecalis Bacteriophage ΦEF24C with a Spontaneous Point Mutation

Uchiyama, Jumpei; Takemura, Iyo; Satoh, Miho; Kato, Shin‐ichiro; Ujihara, Takako; Akechi, Kazue; Matsuzaki, Shigenobu; Daibata, Masanori

doi:10.1371/journal.pone.0026648

Cited by 47 publications

(52 citation statements)

References 46 publications

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“…(46), phage K (47), Romulus and Remus (48) and other Twort-like phages (45) of S. aureus, the ACT group of Bacillus (49), and phiEF24c of E. faecalis (50). The latter, phiEF24c, which was isolated in Japan, is thus far the most studied E. faecalis Spounavirinae phage (42,(50)(51)(52). This phage, which is close in genome size, GC content, and sequence to EFDG1, was found to protect mice from E. faecalis infection in the abdomen and was not harmful to the mice per se.…”

Section: Discussionmentioning

confidence: 99%

Targeting Enterococcus faecalis Biofilms with Phage Therapy

Khalifa

Brosh

Gelman

et al. 2015

Appl Environ Microbiol

188

158

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Phage therapy has been proven to be more effective, in some cases, than conventional antibiotics, especially regarding multidrug-resistant biofilm infections. The objective here was to isolate an anti-Enterococcus faecalis bacteriophage and to evaluate its efficacy against planktonic and biofilm cultures. E. faecalis is an important pathogen found in many infections, including endocarditis and persistent infections associated with root canal treatment failure. The difficulty in E. faecalis treatment has been attributed to the lack of anti-infective strategies to eradicate its biofilm and to the frequent emergence of multidrug-resistant strains. To this end, an anti-E. faecalis and E. faecium phage, termed EFDG1, was isolated from sewage effluents. The phage was visualized by electron microscopy. EFDG1 coding sequences and phylogeny were determined by whole genome sequencing (GenBank accession number KP339049), revealing it belongs to the Spounavirinae subfamily of the Myoviridae phages, which includes promising candidates for therapy against Gram-positive pathogens. This analysis also showed that the EFDG1 genome does not contain apparent harmful genes. EFDG1 antibacterial efficacy was evaluated in vitro against planktonic and biofilm cultures, showing effective lytic activity against various E. faecalis and E. faecium isolates, regardless of their antibiotic resistance profile. In addition, EFDG1 efficiently prevented ex vivo E. faecalis root canal infection. These findings suggest that phage therapy using EFDG1 might be efficacious to prevent E. faecalis infection after root canal treatment.

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Section: Discussionmentioning

confidence: 99%

Targeting Enterococcus faecalis Biofilms with Phage Therapy

Khalifa

Brosh

Gelman

et al. 2015

Appl Environ Microbiol

188

158

View full text Add to dashboard Cite

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“…The phage proteins were separated electrophoretically on a 12.5% SDS-polyacrylamide gel with an (unstained) XLLadder Broad marker (APRO Life Science Institute, Tokushima, Japan), basically followed by the protocol described elsewhere (49). Phage proteins were subjected to N-terminal protein sequencing and mass spectrometry, as described elsewhere (51).…”

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Characterization of Helicobacter pylori Bacteriophage KHP30

Uchiyama

Takeuchi

Kato

et al. 2013

Appl Environ Microbiol

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Helicobacter pylori inhabits the stomach mucosa and is a causative agent of stomach ulcer and cancer. In general, bacteriophages (phages) are strongly associated with bacterial evolution, including the development of pathogenicity. Several tailed phages have so far been reported in H. pylori. We have isolated an H. pylori phage, KHP30, and reported its genomic sequence. In this study, we examined the biological characteristics of phage KHP30. Phage KHP30 was found to be a spherical lipid-containing phage with a diameter of ca. 69 nm. Interestingly, it was stable from pH 2.5 to pH 10, suggesting that it is adapted to the highly acidic environment of the human stomach. Phage KHP30 multiplied on 63.6% of clinical H. pylori isolates. The latent period was ca. 140 min, shorter than the doubling time of H. pylori (ca. 180 min). The burst size was ca. 13, which was smaller than the burst sizes of other known tailed or spherical phages. Phage KHP30 seemed to be maintained as an episome in H. pylori strain NY43 cells, despite a predicted integrase gene in the KHP30 genomic sequence. Seven possible virion proteins of phage KHP30 were analyzed using N-terminal protein sequencing and mass spectrometry, and their genes were found to be located on its genomic DNA. The genomic organization of phage KHP30 differed from the genomic organizations in the known spherical phage families Corticoviridae and Tectiviridae. This evidence suggests that phage KHP30 is a new type of spherical phage that cannot be classified in any existing virus category. Helicobacter pylori, a Gram-negative spiral bacterium, colonizes the human stomach mucosa (1). It causes chronic inflammation, which may progress to peptic ulceration, atrophic gastritis, and gastric cancer (2). Recent studies of H. pylori have revealed that the pathogenicity of H. pylori strains is related to the specific geographic region from which they derive (3, 4). An East Asian type of H. pylori strain most likely causes gastric cancer and highly likely contains cytotoxin-associated gene A (cagA), a carcinogenic genetic element (1, 3). However, how the pathogenicity of the H. pylori strains has evolved is unknown, although H. pylori inhabits a very restricted niche in the stomach mucosa. Therefore, the factors involved in H. pylori evolution must be investigated.Bacteriophages (phages) are the most diverse and abundant life form on Earth. Phages practice lateral gene transfer and are involved in a coevolutionary arms race with bacteria (5-8), so they may well be factors involved in bacterial evolution. Several H. pylori phages have been reported (9-13), and some of these have been morphologically studied and have been found to belong to the order Caudovirales (i.e., the tailed phages) (9-12). Recent advances in sequencing technologies have allowed intensive genomic analyses of Helicobacter spp. and have also suggested the presence of phages in the bacterial genome (10,11,14,15). However, the biological characteristics of these H. pylori phages are not well-known.We have recently iso...

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“…The genomic DNA of the phages was isolated from phage particles that had been purified by CsCl density gradient ultracentrifugation, essentially as described elsewhere (11). The genomic sequences were determined with a primer walking method using an ABI Prism 3100-Avant genetic analyzer (Applied Biosystems, Foster City, CA), and both strands were sequenced.…”

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Complete Genome Sequences of Two Helicobacter pylori Bacteriophages Isolated from Japanese Patients

Uchiyama

Takeuchi

Kato

et al. 2012

J Virol

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Helicobacter pylori causes peptic ulcers and gastric cancer, which lead to significantly higher morbidity in Japan than elsewhere in the world. As bacteriophage (phage) and host bacteria coevolve, the study of H. pylori phages is important to extend understanding of the evolution and pathogenesis of H. pylori. Here we report two complete genome sequences of H. pylori phages KHP30 and KHP40, which were released spontaneously from the most pathogenic East Asian-type isolates from Japanese patients. Helicobacter pylori, a Gram-negative spiral bacterium, colonizes the human stomach and infects approximately 50% of the global population (2, 10). Infection can cause chronic inflammation, which may progress to peptic ulceration, atrophic gastritis, or gastric cancer (8). There is a significant correlation between the H. pylori strain type and the incidence of gastric cancer. In particular, the East Asian-type strain of H. pylori is the individual strain most likely to cause gastric cancer (10).Several bacteriophages (phages) of H. pylori have been reported (4, 5, 9, 12). In general, phages are considered to contribute to bacterial evolution and may affect host features, such as biological behavior, pathogenesis, or adaptation, via their possible roles in horizontal gene transfer and bacteria-phage antagonistic coevolution (4, 5, 7). In this study, to extend our understanding not only of the H. pylori phages themselves but also of the process of coevolution of H. pylori and its phages, phages KHP30 and KHP40 were isolated from the culture supernatants of East Asian-type isolates from Japanese patients living in distinct geographic regions, and their complete genomic sequences were determined.The genomic DNA of the phages was isolated from phage particles that had been purified by CsCl density gradient ultracentrifugation, essentially as described elsewhere (11). The genomic sequences were determined with a primer walking method using an ABI Prism 3100-Avant genetic analyzer (Applied Biosystems, Foster City, CA), and both strands were sequenced. The genome sequences of phages KHP30 and KHP40 were circularly connected. Whole-genome PCR scanning also validated our sequencing results. A BLASTN analysis was conducted between the genome sequences of phages KHP30 and KHP40. Open reading frames (ORFs) were predicted using Prodigal and GeneMark.hmm with heuristic models (1, 3) and were then manually confirmed with reference to the ribosomal binding site sequences. The conserved protein domains were analyzed using the NCBI Conserved Domain Database (6).The complete DNA sequencing of phages KHP30 and KHP40 revealed that their genomes consist of 26,215 bp (GϩC, 35.8%) and 26,449 bp (GϩC, 35.8%), respectively. A comparison of the genomic sequences of KHP30 and KHP40 with BLASTN indicated a high level of sequence similarity (total score, 3.540e Ϫ 04; query coverage, 96%; E value, 0.0). Moreover, 30 and 32 ORFs were inferred in phages KHP30 and KHP40, respectively. A genomic analysis of KHP30 and KHP40 predicted an integrase (ORF2 in ...

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Improved Adsorption of an Enterococcus faecalis Bacteriophage ΦEF24C with a Spontaneous Point Mutation

Cited by 47 publications

References 46 publications

Targeting Enterococcus faecalis Biofilms with Phage Therapy

Targeting Enterococcus faecalis Biofilms with Phage Therapy

Characterization of Helicobacter pylori Bacteriophage KHP30

Complete Genome Sequences of Two Helicobacter pylori Bacteriophages Isolated from Japanese Patients

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