Phage engineering and phage‐assisted CRISPR‐Cas delivery to combat multidrug‐resistant pathogens

Abstract

Antibiotic resistance ranks among the top threats to humanity. Due to the frequent use of antibiotics, society is facing a high prevalence of multidrug resistant pathogens, which have managed to evolve mechanisms that help them evade the last line of therapeutics. An alternative to antibiotics could involve the use of bacteriophages (phages), which are the natural predators of bacterial cells. In earlier times, phages were implemented as therapeutic agents for a century but were mainly replaced with antibiotics, and considering the menace of antimicrobial resistance, it might again become of interest due to the increasing threat of antibiotic resistance among pathogens. The current understanding of phage biology and clustered regularly interspaced short palindromic repeats (CRISPR) assisted phage genome engineering techniques have facilitated to generate phage variants with unique therapeutic values. In this review, we briefly explain strategies to engineer bacteriophages. Next, we highlight the literature supporting CRISPR-Cas9-assisted phage engineering for effective and more specific targeting of bacterial pathogens. Lastly, we discuss techniques that either help to increase the fitness, specificity, or lytic ability of bacteriophages to control an infection.

Bacteriophages as nanocarriers for targeted drug delivery and enhanced therapeutic effects

“…80,81 They can contribute to therapeutic actions by the initial lyses of pathogens; subsequently, the direct intracellular delivery of loads, minimising off-target effects and immune responses. 82,83 Because they are chiefly biocompatible with human and bacterial cells, they can significantly reduce the chances of toxicity and adverse effects. [84][85][86] Ineffective antibiotics owing to the development of bacterial resistance can be reprioritised by the initial lysis and compromise of the bacterial pathogen, especially when the bacterial resistance mechanism is inherent in the cell wall.…”

Bacteriophages as a potential substitute for antibiotics: A comprehensive review

“…The utility of phages can be extendable by using rDNA technology which may serve desired results against MDR and XDR strains of host cells with a high competency rate can be used in this method. 19 In phage-mediated gene delivery, a gene coding for an antibacterial activity or elevating the susceptibility of bacteria against phages can be bioengineered. Engineered phages can be used in vaccine design by cloning antigens in the phage genome, and host cell antigenpresenting cells will recognize the antigen and express antibodies against it, triggering adaptive immunity.…”

“…Genome engineering of phage with plasmid recombination, direct cloning, CRISPR-Cas selection, shuttle-phasmid combination, yeast-mediated assembly, and so forth, are feasible for phage therapy 15. In Bacteriophage recombineering of electroporated DNA (BRED), application of coelectroporation of phage genomic and synthetic DNA (desired gene) into bacterial cells results in gene expression, and progeny plaques are recovered and identified by polymerase chain reaction 15,19. BRED increases the frequency of recombination but has the disadvantage that it entirely depends on the electroporation of donor and phage DNA.…”

On the ever‐growing functional versatility of the CRISPR‐Cas13 system

“…In addition, exploiting this antiviral activity for counterselection in bacteria, the genomes of diverse phages were edited without leaving a trace (Adler et al., 2022 ). This development seems relevant because phage engineering represents a promising weapon in the ongoing battle against antibiotic‐resistant bacteria (Khambhati et al., 2023 ). The use of phages allows for the precise targeting of pathogenic bacteria, while minimizing collateral damage to beneficial microorganisms.…”