No abstract

We report the successful purification of a cloned lysin encoded by the novel Staphylococcus aureus bacteriophage phi MR11. The lysin, designated MV-L, rapidly and completely lysed cells of a number of S. aureus strains tested, including methicillin-resistant S. aureus (MRSA) and vancomycin-resistant S. aureus and a subset of vancomycin-intermediate S. aureus (VISA) in growing conditions. MV-L-mediated killing is specific to S. aureus and not to other species, except for S. simulans. MV-L exerted its staphylocidal effect synergistically with glycopeptide antibiotics against VISA. MV-L efficiently eliminated MRSA that had been artificially inoculated into the nares of mice. The intraperitoneal administration of MV-L also protected mice against MRSA septic death, without any harmful effects. Although MV-L evoked detectable levels of a humoral response in mice, the antibodies did not abolish the bacteriolytic activity. These results indicate that MV-L might be useful as a powerful therapeutic agent against multidrug-resistant S. aureus infections.

Vancomycin-resistant Enterococcus faecalis (VRE) has become a significant threat in nosocomial settings. Bacteriophage (phage) therapy is frequently proposed as a potential alternative therapy for infections caused by this bacterium. To search for candidate therapeutic phages against Enterococcus faecalis infections, 30 Enterococcus faecalis phages were isolated from the environment. One of these, virulent phage phiEF24C, which has a broad host range, was selected for analysis. The plaque-forming ability of phiEF24C was virtually unaffected by differences in the clinical host strains. Furthermore, the phage had a shorter latent period and a larger burst size than ordinary tailed phages, indicating that phiEF24C has effective lytic activity against many Enterococcus faecalis strains, including VRE. Morphological and genomic analyses revealed that phiEF24C is a large myovirus (classified as family Myoviridae morphotype A1) with a linear double-stranded DNA genome of c. 143 kbp. Analyses of the N-terminal amino acid sequences of the virion proteins, together with the morphology and the genome size, speculated that phiEF24C is closely related to other myoviruses of Gram-positive bacteria that have been used experimentally or practically for therapy or prophylaxis. Considering these results, phiEF24C may be a potential candidate therapeutic phage against Enterococcus faecalis infections.

Along with the increasing threat of nosocomial infections by vancomycin-resistant Enterococcus faecalis, bacteriophage (phage) therapy has been expected as an alternative therapy against infectious disease. Although genome information and proof of applicability are prerequisites for a modern therapeutic phage, E. faecalis phage has not been analyzed in terms of these aspects. Previously, we reported a novel virulent phage, EF24C, and its biology indicated its therapeutic potential against E. faecalis infection. In this study, the EF24C genome was analyzed and the in vivo therapeutic applicability of EF24C was also briefly assessed. Its complete genome (142,072 bp) was predicted to have 221 open reading frames (ORFs) and five tRNA genes. In our functional analysis of the ORFs by use of a public database, no proteins undesirable in phage therapy, such as pathogenic and integration-related proteins, were predicted. The noncompetitive directions of replication and transcription and the host-adapted translation of the phage were deduced bioinformatically. Its genomic features indicated that EF24C is a member of the SPO1-like phage genus and especially that it has a close relationship to the Listeria phage P100, which is authorized for prophylactic use. Thus, these bioinformatics analyses rationalized the therapeutic eligibility of EF24C. Moreover, the in vivo therapeutic potential of EF24C, which was effective at a low concentration and was not affected by host sensitivity to the phage, was proven by use of sepsis BALB/c mouse models. Furthermore, no change in mouse lethality was observed under either single or repeated phage exposures. Although further study is required, EF24C can be a promising therapeutic phage against E. faecalis infections.

The therapeutic effects of bacteriophage (phage) KPP12 in Pseudomonas aeruginosa keratitis were investigated in mice. Morphological analysis showed that phage KPP12 is a member of the family Myoviridae, morphotype A1, and DNA sequence analysis revealed that phage KPP12 is similar to PB1-like viruses. Analysis of the phage KPP12 genome did not identify any genes related to drug resistance, pathogenicity or lysogenicity, and so phage KPP12 may be a good candidate for therapeutic. KPP12 showed a broad host range for P. aeruginosa strains isolated from clinical ophthalmic infections. Inoculation of the scarified cornea with P. aeruginosa caused severe keratitis and eventual corneal perforation. Subsequent single-dose administration of KPP12 eye-drops significantly improved disease outcome, and preserved the structural integrity and transparency of the infected cornea. KPP12 treatment resulted in the suppression of neutrophil infiltration and greatly enhanced bacterial clearance in the infected cornea. These results indicate that bacteriophage eye-drops may be a novel adjunctive or alternative therapeutic agent for the treatment of infectious keratitis secondary to antibiotic-resistant bacteria.