Engineering of a temperate burkholderia cepacia complex phage to improve efficacy as a potential therapeutic

The National Institute of Allergy and Infectious Diseases awarded J. Craig Venter Institute, Inc. $529,490 on June 10, 2026, under the Allergy and Infectious Diseases Research program (CFDA 93.855) to engineer a temperate Burkholderia cepacia complex bacteriophage for therapeutic use.

The recipient will engineer a prophage designated BCC02, isolated from Burkholderia genomic material, to disable its ability to integrate into bacterial genomes and increase its therapeutic range against bacteria in the Burkholderia cepacia complex that cause difficult-to-treat infections in immunocompromised patients, particularly those with cystic fibrosis. The project employs transformation-associated recombination cloning methods to target and remove the lysogeny control region of the BCC02 genome, converting it to an obligately lytic lifestyle. Specific deliverables include engineering the phage to eliminate lysogenization capability, testing the modified phage against a clinically relevant panel of pathoadapted isolates from cystic fibrosis patients, and characterizing the effects of modifications on bacterial host range.

Performance occurs in Rockville, Maryland. The period of performance runs from the award date through May 31, 2028. The assistance type is Project Grant.

Abstract

Bacteria in the burkholderia cepacia complex (BCC) cause difficult to treat infections in patients with compromised respiratory systems, such as those with cystic fibrosis (CF). Alternative treatment options are needed, since antibiotics often fail these patients. Bacteriophage (phage) therapy is a promising strategy, yet therapeutically ideal phages are difficult to find and narrow in their range of use due to host specificity. In the proposed study, we continue development of a potential phage therapeutic sourced from burkholderia itself. We have isolated a phage, called BCC02, that was present within the genome of a burkholderia bacteria (a prophage) and have shown that IT can kill other bacteria within the same genus. However, this phage still has the potential to integrate into other bacterial genomes, which is an undesirable trait for phage therapy. By engineering changes to the BCC02 genome using synthetic biology techniques, we hypothesize that we can increase its range of therapeutic potential by disabling its ability to integrate into the bacterial genome, and that this change will increase the number of bacteria that IT can lyse. The specific aims of this project are to (1) engineer this phage to lose the ability to lysogenize (integrate into bacterial genomes) then test the effects of these modifications on bacterial host range and (2) test activity of our originally isolated phage, BCC02 as well as our engineered variant on a clinically relevant panel of patho-adapted isolates from patients with CF. We propose to use transformation-associated recombination (TAR) cloning methods to target the lysogeny control region of the BCC02 genome for removal. We hypothesize that loss of integration ability will force this phage into an obligately lytic lifestyle, where IT will lyse all bacteria IT is able to infect. Successful completion of this project will determine the feasibility of engineering obligately lytic burkholderia-targeting phages from burkholderia SPP. Prophages, shed light on the effects of lytic lifestyle on host range, and establish the utility of these phages for tackling particularly problematic clinical infections. In addition, this study may produce a BCC- targeting phage that is primed for development to be used for phage therapy.