Bacteriophages are a massively diverse group of bacterial viruses and are often cited as the most abundant organisms on the planet. Although the abundance of natural phages means that they...
Genome editing is a revolutionary area of biological research that involves the insertion, deletion, or modification of DNA within the genome of a living organism, with the goal of better understanding or improving the function of specific genes and their protein counterparts. Gene editing consists of several technologies, including CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats and CRISPR-associated protein 9) technology, often referred to as simply CRISPR. Since its inception, scientists across the globe have harnessed the power of CRISPR for several biological challenges, including advanced genome editing strategies. CRISPR-Cas9 can be combined with bacteriophages to provide a powerful tool for precise and targeted genome editing, with potential applications in industrial biotech, medicine, and agriculture, among others. In this article, we will discuss the science behind combining bacteriophages and CRISPR, and what this means for the future of genome editing.
Bacteriophages are highly abundant viruses that specifically target and kill bacteria but are completely harmless to humans. Naturally, they do this by infecting a bacterial host cell with an incredibly high degree of specificity. They are then able to hijack the bacterial host’s replication machinery and replicate within it, producing multiple copies of the phage genome. This eventually causes the bacterium to burst, releasing the newly formed prophages into the surrounding environment where they can infect further host cells. The advantageous natural properties of bacteriophages have been harnessed by scientists for a broad range of applications within human and animal health, agriculture, aquaculture, industry, and environmental sciences.
CRISPR-Cas9 gene editing technology was first reported in 2011 and resulted in a seminal Science paper, published in 2012, which saw the two corresponding authors, Jennifer Doudna and Emmanuelle Charpentier, receive the 2020 Nobel Prize in chemistry for their pioneering work developing this incredibly powerful genome editing tool. The CRISPR-Cas9 system utilises a guide RNA (gRNA) which directs the Cas9 endonuclease to the target DNA sequence, where Cas9 makes double-strand DNA breaks, inducing a DNA repair process during which precise genetic modifications are made.
Bacteriophages have the potential to enhance CRISPR-based gene editing by serving as delivery vehicles: phages can transport the CRISPR-Cas9 system into specific cells for targeted modifications. Concurrently, CRISPR technology can be used as a method of phage genetic engineering, tailoring them for specific uses i.e. to enhance their natural properties, or to introduce new properties, advantageous for a specific application. Therefore, combining bacteriophages and CRISPR technology can create a synergistic enhancement of precision and applicability in genome engineering strategies.
Bacteriophages have several inherent properties that render them advantageous as delivery vehicles for the components of the CRISPR-Cas9 system. Bacteriophages naturally target bacterial cells with an incredibly high degree of selectivity, and they can be genetically modified (including with CRISPR-Cas9-based genetic modification) to target a specific tissue or cell type. Therefore, utilising bacteriophages as delivery vectors for CRISPR-Cas9 facilitates highly specific bacterial genomic modifications, with a low risk of off-target effects since the phages’ selectivity renders them unlikely to infect surrounding cells.
Thus, bacteriophages, with their strain-specific tropism, can efficiently transport the CRISPR-Cas9 system to targeted bacterial cells. Once inside, the gRNA directs Cas9 to introduce a double-strand break at a precise location, which triggers the cell’s endogenous repair machinery, during which time genetic alterations can be implemented. Thus, the combination of bacteriophages’ specificity with CRISPR-Cas9’s precision offers a novel, practical approach for robust, meticulous bacterial genome editing.
The synergy of bacteriophages and the CRISPR-Cas9 system holds significant potential for several biotech applications involving the manipulation of bacterial genomes. Current applications span the industrial, agricultural, and medical fields, though the potential of bacteriophages combined with CRISPR in biotech is vast and the number of applications is likely to grow rapidly in future.
In industrial biotech, the bacteriophage-guided CRISPR system enables targeted modifications of microbial genomes to modify specific genes
and enhance desired phenotypes for a given application. The genetic manipulation of microorganisms using this highly specific, precise system has the potential to drive advancements in biofuel and biomolecule production.
Agriculture can also benefit greatly from this approach. Engineering plant-associated bacterial communities using bacteriophages and CRISPR could introduce beneficial traits to crops, including enhanced nutrient uptake, drought resistance, or disease resilience. These modifications could promote increased crop yield and sustainability in the face of a changing climate.
In the medical field, the use of bacteriophages to carry and precisely target the CRISPR-Cas9 system presents a promising strategy in the fight against antibiotic-resistant bacteria. This technique can be used to selectively target and kill multidrug-resistant pathogens while leaving beneficial bacteria of the microbiome untouched. Furthermore, the system could be tailored to remove antibiotic resistance genes from bacteria, contributing to the mitigation of antibiotic resistance.
The integration of bacteriophages and the CRISPR-Cas9 system demonstrates a powerful biological system with immense potential in the realm of genome editing. This novel combination of technologies offers the advantages of precision, specificity, and efficiency in modifying bacterial genomes, and has significant implications for the biotech and healthcare sectors. In biotechnology, industrial microbe manipulation is set to transform biofuel and biomolecule production. In agriculture, crop enhancement through engineered plant-bacteria interactions offers the potential to enhance food security despite the uncertainty of climate change. The healthcare sector also stands to gain significantly from this symbiosis, specifically by combating the pressing issue of antibiotic resistance. However, challenges remain and further research is required to fully optimise and benefit from the full potential of this strategy. Ongoing work involves enhancing the delivery efficiency of bacteriophages, fine-tuning the specificity of CRISPR-Cas9, and navigating potential off-target effects.