CARB-X has announced its 2019 calls for proposals. Funding Round 1 covers non-traditional alternatives to antibiotics, including phage! Learn how to apply by tuning in May 16 for a webinar. Expressions of interest can be submitted June 3 – June 10, 2019. (Thanks to Wellcome Trust’s Chief Medical Officer John Rex’s newsletter for this info!)
You may recall reading about the discovery of arbitrium, a peptide produced by Bacillus phages to regulate lysogeny decisions. A new paper by Avigail Stokar-Avihail and colleagues in the Sorek lab at the Weizmann Institute has shown that arbitrium-like systems are widespread in phages! What’s more, they’re frequently located on mobile pathogenicity elements, and seem to be controlled by anti-sense RNA.
Ho Bin Jang, Benjamin Bolduc and colleagues from the Sullivan lab at Ohio State University, along with several collaborators from the US, UK, Canada, France, and Belgium, have published a paper this week in Nature Biotechnology describing a new tool, vConTACT v.2.0, that can provide reliable taxonomic assignments for uncultivated viral genomes. Paper | Bitbucket.
The business combination transaction between AmpliPhi and C3J, two phage therapy biotech companies, is expected to close today. The new name of the combined company will be Armata Pharmaceuticals, Inc., and shares will be expected to start trading on the New York Stock Exchange under the symbol “ARMP” tomorrow.
The 26th Biennial Conference on Phage/Virus Assembly is happening July 14-19, 2019 at Cragun’s Resort in Brainerd, Minnesota. Registration is open until May 30.
ICON Government and Public Health Solutions specializes in preclinical through phase IV support of clinical research and clinical trial services for biologics, drugs, and devices. The candidate would perform research on bacteriophages and microbial pathogens of military relevance (e.g. Staphylococcus aureus, Shigella, Pseudomonas, Klebsiella, etc.), including in-depth bacteriophage characterization and animal modeling.
The Leptihn Group offers a Postdoctoral Fellowship: The project involves viruses of microbes (bacteriophages) with aspects on both function and structure of phage proteins, as well as phage therapy. Experience in imaging and protein biochemistry or microbiology are desired.
I’m looking for phage typing collections of Enterobacteriaciae (eg. E. coli, Klebsiella, Salmonella) to pursue capsular variant-phage type correlation studies. Please email me at [email protected] or tweet @mkoeris to get in touch!
This week, we’re going over a new phage therapy case report. This report marks the first time engineered phages have been used to treat a human patient, and the first time a Mycobacterium infection has been treated with phage therapy.
The study was published yesterday in Nature Medicine by Rebekah Dedrick, Carlos Guerrero-Bustamente and colleagues in Graham Hatfull’s phage lab at U Pitt, alongside Helen Spencer, an MD at Great Ormond Street Hospital (GOSH) in London, UK, her colleagues, and Chip Schooley of UCSD’s IPATH.
A 15-year-old girl with cystic fibrosis in the UK, who’d had a double lung transplant, was diagnosed with disseminated Mycobacterium abscessus subspecies massiliense, a form of non-tuberculous Mycobacteria (NTM).
M. abscessus was cultured in her sputum, she had lesions on her liver and skin, and her skin continued to develop new infected nodules, even with antibiotics.
She was sent home 7 months post-transplant with a palliative care plan in place, and given a 1% chance of survival (most patients don’t recover when NTM grows post-transplant).
A personalized phage therapy approach was used to treat the patient. A large phage library was screened against the patient’s isolate, and a three-phage cocktail (two of which were engineered to lyse the strain more efficiently) was given to the patient over at least 32 weeks.
The patient improved substantially, and went on to resume normal activities. Most of her Mycobacterium infection was eliminated. No adverse effects of the phages were observed.
The patient’s strain was isolated 1 month post-transplant and found to be Mycobacterium abscessus, subspecies massiliense. It was resistant to all antibiotics tested: clarithromycin, amikacin, tobramycin, ciprofloxacin, moxifloxacin, cefoxitin, cotrimoxazole, doxycycline and linezolid.
M. abscessus is a species of non-tuberculous Mycobacteria (NTM). NTM frequently colonize CF patients, and is commonly associated with poor prognosis due to antibiotic resistance.
The strain was sent to Graham Hatfull’s Mycobacterium phage genetics lab. A collection of >10,000 phages isolated by undergraduate students in the SEA-PHAGES program was exploited (representatives were screened against the clinical strain). They also screened >100 environmental isolates, and cultured the patient’s strain with a pool of unsequenced phages to enrich for active phages.
All three phages were found to be siphoviridae, but all were from unrelated families and showed little genetic similarity.
The two temperate phages were each genetically engineered to lose their lytic repressor gene using the BRED technique, developed in the Hatfull lab by Laura Marinelli and colleagues in 2008.
BRED engineering successfully made one phage, ZoeJ, an efficient killer, but the other, BPs, was still producing turbid plaques. For this phage, host range mutants were isolated that could kill more efficiently (genetic analysis showed single base changes in a portal gene of the phage). The three phages were combined in a cocktail.
The three-phage cocktail was assessed for phage resistance by culturing with the patient’s strain in vitro. Survivor colonies, which grew at a low frequency, were tested individually for phage resistance, and all retained sensitivity to at least one phage in the cocktail.
First, a topical test-dose of the phage was administered to the sternal wound. After 24 h observation, and no ill-effects, the three-phage cocktail (10^9 PFU per phage) was administered intravenously every 12 h for at least 32 weeks. After noticing that the sternal wound, which had received a topical dose of phages, began to close first, phages were also administered topically to the other skin lesions.
Endotoxin levels were not a concern in this case, as Mycobacteria do not produce lipopolysaccharide (LPS).
Yes: the patient remained on the same antibiotics regimen throughout phage therapy (amikacin, imipenem/cilastatin, tigecycline, bedaquiline, clofazimine).
After 9 days, the patient was discharged, but continued taking IV phage therapy at home. Over the next 6 months, her sternal wound (the lung transplant incision site) healed, her infected skin nodules substantially resolved (though some skin nodules remained), and her lung and liver function improved.
M. abscessus could not be cultured from her blood or sputum at any time post-phage threatment, although it could still be cultured from the remaining skin nodules.
The patient was able to go back to school and resume her normal life, although according to a BBC news article from this week, her phage treatment continues to this day. In addition, the Hatfull lab is working to prepare a fourth phage to add to the cocktail, to help eradicate the remaining M. abscessus.
No adverse effects, other than feeling sweaty and flushed at first, were observed. Immune reactions were minimal (low cytokine response), and although a weak anti-phage protein antibody response was detected, it was not enough to neutralize the phages.
The authors tracked phage concentrations (using both plaque assays and digital PCR) at various body sites, and found substantial replication of phages (10 PFU/mL to 10^9 PFU/mL) in the blood within about 4 days.
They tracked phages in serum, feces, wounds and sputum over several months. By plaque assay, phages were first detected in the serum (1 day post-initial phage treatment), and then in the feces (days 4-6) and in wounds (days 3-5), followed by a spike of phages in the sputum (day 9), then about a month of undetectable phages, then two days where phages were again detected in serum. dPCR showed similar results, albeit resulting in generally lower numbers compared with plaque assays.
Although some phage resistance was observed in vitro, phage resistance was not detected in vivo. Not all the skin nodules were resolved, but wound swab isolates showed no phage resistance.
(For detailed info, see the Supplementary Material for the paper)
This study shows the safe treatment of a multi-drug resistant, disseminated, non-tuberculous Mycobacterium infection in a post-transplant patient with CF using a three-phage cocktail. Phages against the patient’s strain were uncommon, but could be identified with extensive searches. The treatment substantially reduced the presence of M. abscessus, phage replication was clearly demonstrated, and the condition of a patient who was not expected to live was substantially improved.
The phage cocktail was not effective on other M. abscessus strains the lab tested, suggesting against its general use in other patients. However, the Hatfull lab did show that they could find other phages in their collection that were effective against these strains. This suggests that a personalized approach could be promising for this species, given a large enough phage library. However, the paper mentions another patient with a similar infection, who unfortunately died before they could find appropriate phages, emphasizing the inherent difficulty of using personalized phage therapy for critically-ill patients infected with this species.
Rebekah M. Dedrick, Carlos A. Guerrero-Bustamante, Rebecca A. Garlena, Daniel A. Russell, Katrina Ford, Kathryn Harris, Kimberly C. Gilmour, James Soothill, Deborah Jacobs-Sera, Robert T. Schooley, Graham F. Hatfull & Helen Spencer. Engineered bacteriophages for treatment of a patient with a disseminated drug-resistant Mycobacterium abscessus (2019). Nature Medicine 25, 730–733.
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