Engineering bacteriophages for enhanced host range and efficacy: insights from bacteriophage-bacteria interactions

Abstract

Bacteriophages, the most abundant organisms on earth, have the potential to address the rise of multidrug-resistant bacteria resulting from the overuse of antibiotics. However, their high specificity and limited host range can hinder their effectiveness. Phage engineering, through the use of gene editing techniques, offers a means to enhance the host range of bacteria, improve phage efficacy, and facilitate efficient cell-free production of phage drugs. To engineer phages effectively, it is necessary to understand the interaction between phages and host bacteria. Understanding the interaction between the receptor recognition protein of bacteriophages and host receptors can serve as a valuable guide for modifying or replacing these proteins, thereby altering the receptor range of the bacteriophage. Research and development focused on the CRISPR-Cas bacterial immune system against bacteriophage nucleic acids can provide the necessary tools to promote recombination and counter-selection in engineered bacteriophage programs. Additionally, studying the transcription and assembly functions of bacteriophages in host bacteria can facilitate the engineered assembly of bacteriophage genomes in non-host environments. This review highlights a comprehensive summary of phage engineering methods, including in-host and out-of-host engineering, and the use of high-throughput methods to understand their role. The main aim of these techniques is to harness the intricate interactions between bacteriophages and hosts to inform and guide the engineering of bacteriophages, particularly in the context of studying and manipulating the host range of bacteriophages. By employing advanced high-throughput methods to identify specific bacteriophage receptor recognition genes, and subsequently introducing modifications or performing gene swapping through in-host recombination or out-of-host synthesis, it becomes possible to strategically alter the host range of bacteriophages. This capability holds immense significance for leveraging bacteriophages as a promising therapeutic approach against antibiotic-resistant bacteria.

Transformation-associated recombination (TAR) cloning and its applications for gene function; genome architecture and evolution; biotechnology and biomedicine

“…They are ubiquitous in the environment and are recognized as the most abundant type of organisms on earth. Recently they have received renewed attention for their potential to address the rise of multidrug-resistant bacteria resulting from the overuse of antibiotics [ 126 , 127 ]. Therefore, modification of phages by homologous recombination in yeast or their assembly by TAR cloning provides a promising approach against antibiotic-resistant bacteria.…”

“…One among multiple examples of TAR engineering of phages is a full phiX174 genome assembly with the yield 44% of the required clones [ 128 ]. More examples of TAR-based assembly of phage DNA fragments into complete genomes in yeast, followed by transformation into the hosts to produce activated phages for drug production are summarized by Jia et al [ 127 ].…”

Bacterial Receptors but Not Anti-Phage Defence Mechanisms Determine Host Range for a Pair ofPseudomonas aeruginosaLytic Phages

“…One primary impediment is the narrow host range exhibited by many phages 18 . Each phage typically targets a specific bacterial host, a characteristic that also contributes to the safety of phage therapy.…”

“…The identification of bacterial receptors, which are important for host range determination, remains a major hurdle, as the majority of these receptors remain unknown. The lack of comprehensive knowledge about bacterial receptors hinders our ability to predict and engineer phages for effective therapies 18 .…”

CRISPR RNA-Guided Transposases Facilitate Dispensable Gene Study in Phage

“…Although efficiency can be improved by integrating CRISPR-Cas systems, this method risks causing cell death through double-strand breaks (DSBs) [ 11 , 12 , 13 ]. Complete genome reconstruction in non-host systems allows for genome design flexibility, but these reconstructed genomes often fail to reboot [ 14 , 15 , 16 , 17 ]. These emphasize the need for new phage genome engineering methods to identify the importance of phage genes.…”