Layperson summary
Bacteriophages (phages) are viruses that are able to specifically infect and kill bacteria. In certain exceptional cases, they are used for phage therapy against infections caused by pathogenic bacteria when antibiotic therapy fails. Bacteriophages are highly specific to their host bacterium so that a match between the patient’s bacterium and a therapeutic phage must be identified before phage therapy can be initiated. For this purpose, extensive collections of characterized and purified phages are screened for a suitable match. Sometimes, due to high specificity, a match cannot be found even in large phage collections with >500 phages against one pathogenic bacterial species.
In that case, local sources can be searched to isolate new phages matching the patient’s bacterium. This search is not complicated in principle, as phages are everywhere, but the characterization and purification of the phage may take some time.
There are now hundreds of case reports on phage therapy, for example here and here and here.
Academic summary:
The spread of antibiotic resistant bacteria is a global phenomenon and requires new therapeutic options. An alternative to antibiotics are lytic bacteriophages (phages), which specifically lyse their host bacterium. In Eastern Europe and increasingly in the US, the UK and in the EU, bacteriophage based therapeutics are used by physicians if antibiotic therapy fails.
An overview of a phage therapy pipeline can be found here.
Phage isolation and characterization
Phages for therapy are stable and able to withstand some measure of shearing and breakdown during filtration and freezing. Phages are mostly stored at 4°C or lyophilized. Phage isolation and characterization described below is designed for unenveloped, short tail, DNA bacteriophages mostly represented in the family Caudovirales.
Isolation, enrichment and host range studies Environmental surface and waste water are centrifuged at 7.000 x g for 10 min at 4°C to remove crude sediment, followed by filtration through 0.2 μm filters. The sterile filtrate contains bacteriophages and small particles including other viruses.
Phages are enriched on the host of choice and plaque assays are used to isolate and select a specific phage from sterile filtrates. 100 μl filtrat and 300 μl of a target bacterial strain at OD600 0.4-0.6 are added to 2.5 ml of (40°C) LB-Agar (0.6%) with 1mM CaCl₂ and poured onto a 2% LB Agar plate. Plates are incubated inverted at 37°C for 24h. Further analysis of phage lysis as well as phage purification steps follow. Phage host range is determined by testing on a panel of bacterial target strains and establishing the extent of phage replication using growth curves.
Genome extraction and sequencing DNA is extracted from phage particles using commercially available kits or phenol chloroform DNA extraction protocols. Phage stocks of >1 x 108 PFU (Plaque forming units)/mL are required. Phage genomes are sequenced using, for example, minION ® – and Illumina®/MiSeq® – based systems.
Electron microscopy Purified phage solution is added to carbon-coated Cu400 TEM grids, followed by a negative stain. This is followed by vacuum drying and imaging with a transmission electron microscope at 100 kV and a magnification between × 15,500 and × 21,000.
Phage production Several phage production methods exists, adapted to the patient’s time constraints and type of infection. High expertise in phage production is required to offer safe and effective phage therapy.
Treatment requirements If a patient is eligible for phage therapy and the treating physician approves an experimental therapeutic approach, and suitable and high-quality phages can be found for the bacteria of interest in the required time frame, a phage-based personalized approach might be considered upon request of the patient. The Phage4-1Health Consortium advises patients and physicians on potential approaches and necessary measures before, during, and after phage therapy on a case-by-case basis. We advise the treating physicians to contact the Phage4-1Health Consortium for information prior to phage therapy.
The form of delivery The form of delivery depends on the infection site, and is discussed with phage experts, physicians, and pharmaceutical professionals. In published studies, phages have been applied in various routes including oral, i.v., rectal, topic, and inhalative application. The schedule of treatment, monitoring, and follow-up is developed in an interdisciplinary way for each individual case.