High murine blood persistence of phage T3 and suggested strategy for phage therapy

Serwer, Philip; Wright, Elena T.; Lee, John C.

doi:10.1186/s13104-019-4597-1

Cited by 8 publications

(6 citation statements)

References 37 publications

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“…The data indicate that murine blood persistence varies by orders of magnitude among different phages. Phage T3 has the highest known persistence with detectable blood titer loss not occurring for 3-4 h [54]. Thus, rapid screening for high persistence is possibly going to become rate limiting for the speed at which phage therapy cocktails are assembled.…”

Section: Further Screening In-plaque: Native Gel Electrophoresis (Agementioning

confidence: 99%

In-Gel Isolation and Characterization of Large (and Other) Phages

Serwer

Wright

2020

Viruses

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We review some aspects of the rapid isolation of, screening for and characterization of jumbo phages, i.e., phages that have dsDNA genomes longer than 200 Kb. The first aspect is that, as plaque-supporting gels become more concentrated, jumbo phage plaques become smaller. Dilute agarose gels are better than conventional agar gels for supporting plaques of both jumbo phages and, prospectively, the even larger (>520 Kb genome), not-yet-isolated mega-phages. Second, dilute agarose gels stimulate propagation of at least some jumbo phages. Third, in-plaque techniques exist for screening for both phage aggregation and high-in-magnitude, negative average electrical surface charge density. The latter is possibly correlated with high phage persistence in blood. Fourth, electron microscopy of a thin section of a phage plaque reveals phage type, size and some phage life cycle information. Fifth, in-gel propagation is an effective preparative technique for at least some jumbo phages. Sixth, centrifugation through sucrose density gradients is a relatively non-destructive jumbo phage purification technique. These basics have ramifications in the development of procedures for (1) use of jumbo phages for phage therapy of infectious disease, (2) exploration of genomic diversity and evolution and (3) obtaining accurate metagenomic analyses.Here, we both review and augment data projected to help answer the above questions. We propose that data of this type are critical to efficient isolation of large phages, which include those with genomes longer than 200 Kb (jumbo phages [4][5][6]19]) and even larger phages with genome longer than 520 Kb (mega-phages [20,21]). The importance of these answers is illustrated by the fact that no mega-phage has ever been isolated [20,21], although even larger eukaryotic viruses have been isolated [22]. The existence of mega-phages is surmised from assembly of metagenomic sequences [20,21]. Host Bacteria In-Gel: Values of P E for Agarose Gels FundamentalsThe following data indicate that Gram-negative and Gram-positive phage hosts do not fit in the spaces of the 0.5-0.7% agar gels typically used to form phage plaques. This conclusion is derived, first, from the measurement of P E for agarose gels. This measurement begins by determining vs. sphere radius the minimal agarose gel concentration that excludes a sphere during electrophoresis [23]. These P E values are then used to anchor P E values determined from the sieving (gel-induced retardation) of spheres during gel electrophoresis. The most advanced study of the latter [24] yields the following relationship for underivatized, low-electro-osmosis agarose gels cast in 0.025 M sodium phosphate, pH 7.4, 0.001 M MgCl 2 at 25 • C: P E = 148A −0.87 nm; A is the weight percentage of agarose.A 0.7% agarose gel, if characterized by this relationship, has a P E of 202 nm, not large enough to admit a bacterial cell, such as the host for phage G. Our phage G host is~500 nm in radius and 2000-5000 nm in length, as is Escherichia coli [25], a representative of the ...

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Section: Further Screening In-plaque: Native Gel Electrophoresis (Agementioning

confidence: 99%

In-Gel Isolation and Characterization of Large (and Other) Phages

Serwer

Wright

2020

Viruses

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“…At one extreme in the spectrum of phage persistence, as measured via blood titer, we found persistence surprisingly high for phage T3. The peak blood titer of phage T3, after IP injection into a mouse, did not decrease by more than 2× until 3–4 h after injection [ 55 ]. In this case, even if average phage production was as low as 2.5 phages per 5 min per bacterial cell and the initial bacterial concentration was 10 7 per mL (approaching lethal levels), then phages would, conservatively, gain the upper hand within 3 h, assuming a bacterial doubling time of 1 h and 0.001 for the multiplicity of infection.…”

Section: The Persistence Factormentioning

confidence: 99%

“…More detailed tests should be undertaken regarding the correlation of persistence with the magnitude of negative phage σ. This process can be simplified by the following versions of native gel electrophoretic analysis: (1) phage samples from single plaques, without any phage purification or concentration [ 55 ], and (2) two-dimensional NGE to obtain relative σ values in a single gel [ 81 ].…”

Section: Increasing Screening Efficiency: Moving Toward the Ultimate Objective In Phage Therapymentioning

confidence: 99%

Basics for Improved Use of Phages for Therapy

Serwer

Wright

Chapa³

et al. 2021

Antibiotics

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Blood-borne therapeutic phages and phage capsids increasingly reach therapeutic targets as they acquire more persistence, i.e., become more resistant to non-targeted removal from blood. Pathogenic bacteria are targets during classical phage therapy. Metastatic tumors are potential future targets, during use of drug delivery vehicles (DDVs) that are phage derived. Phage therapy has, to date, only sometimes been successful. One cause of failure is low phage persistence. A three-step strategy for increasing persistence is to increase (1) the speed of lytic phage isolation, (2) the diversity of phages isolated, and (3) the effectiveness and speed of screening phages for high persistence. The importance of high persistence-screening is illustrated by our finding here of persistence dramatically higher for coliphage T3 than for its relative, coliphage T7, in murine blood. Coliphage T4 is more persistent, long-term than T3. Pseudomonas chlororaphis phage 201phi2-1 has relatively low persistence. These data are obtained with phages co-inoculated and separately assayed. In addition, highly persistent phage T3 undergoes dispersal to several murine organs and displays tumor tropism in epithelial tissue (xenografted human oral squamous cell carcinoma). Dispersal is an asset for phage therapy, but a liability for phage-based DDVs. We propose increased focus on phage persistence—and dispersal—screening.

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“…Phage-treated bacteremic mice had a survival rate of almost 100%, and no viable pathogen could be detected at 96 h post inoculation. The authors estimate that, in order for therapy of sepsis to be efficient, phages should persist in the blood for at least 3–5 h. A native agarose gel electrophoresis was applied to assess phage surface associated with high blood persistence [ 11 ].…”

Section: Phage Therapy Of Experimentally Induced Sepsismentioning

confidence: 99%

Sepsis, Phages, and COVID-19

2020

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Phage therapy has emerged as a potential novel treatment of sepsis for which no decisive progress has been achieved thus far. Obviously, phages can help eradicate local bacterial infection and bacteremia that may occur in a syndrome. For example, phages may be helpful in correcting excessive inflammatory responses and aberrant immunity that occur in sepsis. Data from animal studies strongly suggest that phages may indeed be an efficient means of therapy for experimentally induced sepsis. In recent years, a number of reports have appeared describing the successful treatment of patients with sepsis. Moreover, novel data on the anti-viral potential of phages may be interpreted as suggesting that phages could be used as an adjunct therapy in severe COVID-19. Thus, clinical trials assessing the value of phage therapy in sepsis, including viral sepsis, are urgently needed.

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High murine blood persistence of phage T3 and suggested strategy for phage therapy

Cited by 8 publications

References 37 publications

In-Gel Isolation and Characterization of Large (and Other) Phages

In-Gel Isolation and Characterization of Large (and Other) Phages

Basics for Improved Use of Phages for Therapy

Sepsis, Phages, and COVID-19

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