similar to traditional phage therapy, the distribution of phages into target tissues or organs is crucial. The use of phage-delivered CRISPR-Cas as an antimicrobial agent against a target bacterium also requires sufficient titre and time. Encapsulation is a method that improves phage adsorption and distribution, increasing the circulation time by evasion of the immune response [50], stomach acidity [51], [52] and enzyme-rich tissue fluids or free radical

Bacteria can develop resistance through mutations and other mechanisms. However, acquisition of resistance against phages may be accompanied by a decrease in virulence or a reduction in bacterial fitness, based on the ‘trade-offs’ evolutionary rationale

The use of phage-based vectors as phage therapy to deliver the CRISPR-Cas system into target bacteria currently has several limitations, including a narrow host range, bacterial resistance, safety issues and phage clearance

In phage genome delivery, engineered temperate bacteriophages are generally used to deliver the CRISPR-Cas system

CRISPR-Cas delivery into target cells has multiple approaches. However, its delivery as an antimicrobial targeting antimicrobial-resistant bacterial cells is limited.

Specific target genes in bacteria can be knocked out using genome editing using the CRISPR-Cas system. These cleavage targets include bacterial virulence factors [32], antibiotic resistance genes [28], [31], [32], [35], [36] and genes unique to bacterial species. Following cleavage of target genes, several mechanisms resulting in cell death may occur: (i) irreparable genome damage; (ii) cell lysis during phage lytic replication; and (iii) re-sensitisation to accompanying antimicrobials

Non-viral delivery includes the use of nanoparticles [28], electroporation [29], hydrodynamic delivery [30] and conjugative delivery [31], [32]. Viral delivery encompasses the use of retroviruses, lentiviruses, adeno-associated viruses, adenoviruses, herpes viruses and, recently, bacteriophage delivery

There are four principal material combinations that can be delivered into the target cell by the CRISPR-Cas system: (A) plasmid or phagemid encoding Cas9 protein and sgRNA [23]; (B) Cas9 mRNA and sgRNA [24]; (C) Cas9 protein and sgRNA [25]; and (D) sgRNA alone

The mechanism of the CRISPR-Cas system response to invading phages or plasmids can be summarised in three stages: adaptation; expression; and interference

This system is separated into two classes (class 1 and class 2), with three major types in each class (class 1 including types I, III and IV, and class 2 including types II, V and VI) according to phylogeny, sequence, locus organisation and content [6], [19], [20]. Currently, the type II system (CRISPR-Cas9) is the most studied and most frequently used for genome editing

Phage therapy through intravesical administration has been used to treat urinary tract infections in patients scheduled to undergo transurethral resection of the prostate

However, the design and execution of the delivery of the CRISPR-Cas system for targeting micro-organisms remains a significant challenge.

abbreviated as the CRISPR-Cas system, which is an adaptive immune system of prokaryotes

However, a lack of randomised controlled trials (RCTs) demonstrating the safety and efficacy of phage therapy, as well as several regulatory issues such as production and marketing authorisation, pose obstacles to its practical use

However, with the emergence of antimicrobial resistance, phage therapy has become a promising therapeutic option for combating these ‘superbugs’.

Bacteriophages (phages) are a category of viruses capable of infecting bacteria.

Phage therapy is a promising approach to combat MDR bacteria. An increasing number of reports have been published on phage therapy and the successful application of antibacterials derived using this method. Additionally, the CRISPR-Cas system has been used to develop antimicrobials with bactericidal effects in vivo

Multidrug-resistant (MDR) bacterial infections in humans are increasing worldwide

It is vitally important that we learn from past mistakes and explore the possible applications of phages in order to appropriately exploit their very potential in human medicine.

There are great advantages with phage therapy to treat resistant strains but theoretical treatment methods and successful clinical trials are essential before their implementation

Antibiotics are treasures and many times lifesaving agents. They can retain this stature only if they are handled correctly and prescribed appropriately.

Bacteria can inhibit phage genome injection by the super injection exclusion (Sie) system (protein-mediated event) encoded by prophage which render bacteria resistant to further phage infections

In phage DNA vaccines, eukaryotic gene expression cassette clone the sequence of antigen inserted into the phage genome

Phages are receiving huge attention as optimal vaccine delivery vehicle, as they are highly stable and simple.

Despite antimicrobials being the sole lifesaving tool in defending against microbial infections, pathogens quickly acquire phenotype resistance within few years of their introduction.

ioengineered phages are designed, created or modified form of existing biological entities to accomplish desired task as they would not do naturally

Anti-phage antibodies may be produced due to the rapid release of endotoxins which might cause major life-threatening immune reactions like anaphylaxis.

Bacteriophage is known to induce both humoral and cellular immune system. These two immune systems sometimes eliminate antibacterial activity of phages by neutralizing and clearing them from the body

Phage resistant bacteria may develop mutated receptors that are not always disadvantageous because the bacteria may completely lose their virulence [60] and become more sensitive to other phages.

It has elevated the risk from soil to humans as it enters the food chain, threatens antimicrobial efficacy to combat pathogenic infections

Phage cocktails found to be effective than individual phage in the treatment of UTIs caused by UPECs. Phage cocktail (T4 and KEP10 phage) were obtained from wastewater, first introduced into the mouse peritoneal cavity and then to its internal organs through blood for the successful elimination of uro-pathogenic strains in UTIs

In 2015, phage therapy to treat wound infections in animal model was demonstrated by the US Navy research group. In their study, cocktails of bacteriophages against A. baumannii demonstrated therapeutic effect in immunosuppressed full-thickened dorsally infected wound mice model. The cocktail reduced the biofilm production in wound, averted the spread of infection and necrosis to surrounding tissues as well as infection–associated morbidity

The phage not only effectively reduced the biofilm formation but also inhibited growth of phage resistant bacterial mutants

Studies have shown promising results with phage isolated from hospital sewage against Enteropathogenic E. coli (EPEC) in mice [44] and coliphages effective against planktonic and biofilm related infections

Several reports have demonstrated phage applications to evade bacterial infections from various crops such as onions [34] and citrus [35]. US FDA has also approved phage cocktails in food processing industries. Phage preparations are accepted as antibacterial food additives for poultry products and ready-to-eat meat

Currently bacteriophage therapy as a novel approach is gaining major interest, as antimicrobial resistant pathogens have emerged. They are successfully used in agriculture worldwide. Food and Drug Administration (FDA) has approved the use of certain phages on crops in order to reduce crop diseases.

Indeed, bacteriophages are the potential agent to be utilized in diverse biotechnological applications, varying from food – animal agriculture, human and veterinary medicine to environmental science

Phage cocktails could also be used to ensure the adequate coverage of common strains and to reduce the probability of phage resistant bacterial mutants [14]. To circumvent immune response issues, one should carefully study each case and choose the appropriate administration route, phage exposure time, dose and buffers.

Several studies have been done to investigate the role of phages in combination with antibacterial drugs to restore antibacterial activities through synergy

Amid the looming crisis of antimicrobial resistant pathogens, dual therapy is considered as a newer strategy to combat global threat

Phage encoded enzymes serve as an alternative to the whole phage application. Phage genome encodes numeral enzymes and proteins required to rupture bacterial cells during infection.

Bioengineered phages are used for the targeted delivery of genes or lethal substances to the infection site

The major goal of genetic engineering is to generate bacteriophage with broader host range, lack of toxin genes and other traits to perform desired task.

Thus, phage therapy demonstrates propitious evidence and safety profile.

Enteric pathogens form principal part of the medical burden around the globe. The most common intestinal infections causing bacteria are E. coli causing gastroenteritis, Helicobacter pylori causing chronic gastritis, Salmonella enterica causing salmonellosis and Clostridium difficile causing diarrhea.

phage therapy is the direct application of naturally isolated virulent phage to the patient with an aim of lysing pathogenic bacteria responsible for causing infectious diseases

Bacteriophage therapy effectively reduced bacterial load and enhanced wound healing which was infected with P. aeruginosa and S. aureus, in a study demonstrated in both swine and rodent models by Mendes

everal studies have evaluated phage therapy potentials for the treatment of gastrointestinal infections caused by V. cholera, C. difficile, E. coli and S. enterica. Nale et al. [17] demonstrated that phage therapy significantly reduced C. difficile biofilm and prevented colonization in a G. mellonella model when used solely or in combination with antibiotics (vancomycin).

Myoviridae, Siphoviridae and Podoviridae are the three abundant virus families in the human gut

Patients in intensive care unit (ICU) settings often suffer from ventilator associated pneumonia (VAP) infections caused by MDR Pseudomonas aeruginosa, Streptococcus pneumoniae, Staphylococcus aureus, Acinetobacter baumanni and Haemophilus influenzae.

(Pneumonia, and cystic fibrosis), gastrointestinal infections, and topical and wound infections.

In later stages due to the external stimuli lysogenic cycle is switched to active lytic cycle.

phages can be divided into two types, namely lytic (virulent) phages and lysogenic (temperate) phage

The classifications are based upon the evaluation of diverse phage properties such as genome composition, morphology, host range, sequence similarity and pathogenicity

Typically, bacteriophage morphology exhibit well defined three-dimensional structure. The genetic material is enclosed in an icosahedral protein capsid head, a tail (spiral contractile sheath surrounding a core pipe and a baseplate with tail fibers) and surface receptor proteins responsible to recognize specific surface molecules on the host bacterium

Bacteriophages are antibacterial agents ubiquitous in nature. With increase in antibiotic resistance, use of bacteriophages as therapeutics has become resurgent in recent times.

Bacteriophages are viruses, the most abundant organisms and the natural predators of bacteria. They are self-replicating, obligatory intracellular parasites and inert biochemically in extracellular environment. They control the biosynthetic machinery of bacterial host and behest them to produce different viral proteins. They are considered as particles outside the host cell containing nucleic acid (DNA or RNA) which encode necessary information required for their replication. They are primordial ubiquitous organisms found in diverse environment such as soil, water, feces etc

phages can be used for different applications ranging from human antibiotherapy to environment disinfection.

Phage therapy is an age old practice of pre-antibiotic era in which bacteriophages are harnessed as bio-agents against bacterial infections.

Indiscriminate use of antibiotics practiced in clinical setup, industry, animal husbandry and agriculture has resulted in the surge of resistance in microorganisms

Development of drugs has revolutionized the treatment methods of bacterial infectious diseases that have had killed millions of humans in pre-antibiotic era.

Development of drugs has revolutionized the treatment methods of bacterial infectious diseases that have had killed millions of humans in pre-antibiotic era.