Genomic surveillance as a scalable framework for precision phage therapy against antibiotic-resistant pathogens.

Phage therapy is gaining increasing interest in the fight against critically antibiotic-resistant nosocomial pathogens. However, the narrow host range of bacteriophages hampers the development of broadly effective phage therapeutics and demands precision approaches. Here, we combine large-scale phylogeographic analysis with high-throughput phage typing to guide the development of precision phage cocktails targeting carbapenem-resistant Acinetobacter baumannii, a top-priority pathogen. Our analysis reveals that a few strain types dominate infections in each world region, with their geographical distribution remaining stable within 6 years. As we demonstrate in Eastern Europe, this spatiotemporal distribution enables preemptive preparation of region-specific phage collections that target most local infections. Finally, we showcase the efficacy of phage cocktails against prevalent strain types using in vitro and animal infection models. Ultimately, genomic surveillance identifies patients benefiting from the same phages across geographical scales, thus providing a scalable framework for precision phage therapy.

Phage therapy: From biological mechanisms to future directions.

Increasing antimicrobial resistance rates have revitalized bacteriophage (phage) research, the natural predators of bacteria discovered over 100 years ago. In order to use phages therapeutically, they should (1) preferably be lytic, (2) kill the bacterial host efficiently, and (3) be fully characterized to exclude side effects. Developing therapeutic phages takes a coordinated effort of multiple stakeholders. Herein, we review the state of the art in phage therapy, covering biological mechanisms, clinical applications, remaining challenges, and future directions involving naturally occurring and genetically modified or synthetic phages.

Targeted suppression of human IBD-associated gut microbiota commensals by phage consortia for treatment of intestinal inflammation.

Human gut commensals are increasingly suggested to impact non-communicable diseases, such as inflammatory bowel diseases (IBD), yet their targeted suppression remains a daunting unmet challenge. In four geographically distinct IBD cohorts (n = 537), we identify a clade of Klebsiella pneumoniae (Kp) strains, featuring a unique antibiotics resistance and mobilome signature, to be strongly associated with disease exacerbation and severity. Transfer of clinical IBD-associated Kp strains into colitis-prone, germ-free, and colonized mice enhances intestinal inflammation. Stepwise generation of a lytic five-phage combination, targeting sensitive and resistant IBD-associated Kp clade members through distinct mechanisms, enables effective Kp suppression in colitis-prone mice, driving an attenuated inflammation and disease severity. Proof-of-concept assessment of Kp-targeting phages in an artificial human gut and in healthy volunteers demonstrates gastric acid-dependent phage resilience, safety, and viability in the lower gut. Collectively, we demonstrate the feasibility of orally administered combination phage therapy in avoiding resistance, while effectively inhibiting non-communicable disease-contributing pathobionts.

Predictive phage therapy for Escherichia coli urinary tract infections: Cocktail selection for therapy based on machine learning models.

This study supports the development of predictive bacteriophage (phage) therapy: the concept of phage cocktail selection to treat a bacterial infection based on machine learning (ML) models. For this purpose, ML models were trained on thousands of measured interactions between a panel of phage and sequenced bacterial isolates. The concept was applied to Escherichia coli associated with urinary tract infections. This is an important common infection in humans and companion animals from which multidrug-resistant (MDR) bloodstream infections can originate. The global threat of MDR infection has reinvigorated international efforts into alternatives to antibiotics including phage therapy. E. coli exhibit extensive genome-level variation due to horizontal gene transfer via phage and plasmids. Associated with this, phage selection for E. coli is difficult as individual isolates can exhibit considerable variation in phage susceptibility due to differences in factors important to phage infection including phage receptor profiles and resistance mechanisms. The activity of 31 phage was measured on 314 isolates with growth curves in artificial urine. Random Forest models were built for each phage from bacterial genome features, and the more generalist phage, acting on over 20% of the bacterial population, exhibited F1 scores of >0.6 and could be used to predict phage cocktails effective against previously untested strains. The study demonstrates the potential of predictive ML models which integrate bacterial genomics with phage activity datasets allowing their use on data derived from direct sequencing of clinical samples to inform rapid and effective phage therapy.

Lining the small intestine with mycobacteriophages protects from Mycobacterium avium subsp. paratuberculosis and eliminates fecal shedding.

Johne's disease (JD), a chronic, infectious enteritis caused by Mycobacterium avium subsp. paratuberculosis (MAP), affects wild and domestic ruminants. There is no cure or effective prevention, and current vaccines have substantial limitations, leaving this disease widespread in all substantial dairy industries causing economic, and animal welfare implications. Mycobacteriophages (MPs) have been gaining interest in recent years and are proposed as a promising solution to curtailing MAP infection. Using a well-validated infection model, we have demonstrated the preventative potential of MPs to protect dairy calves against MAP infection. Calves were supplemented daily with a phage cocktail from birth till weaning at 2 m of age and inoculated with MAP at 2 wk of age. Infection status was measured for 4.5 mo through blood, fecal, and postmortem tissue samples. Our findings highlight the remarkable efficacy of orally administered MPs. Notably, fecal shedding of MAP was entirely eliminated within 10 wk, in contrast to the infected control group where shedding continued for the entirety of the trial period. Postmortem tissue culture analysis further supported the effectiveness of MPs, with only 1 out of 6 animals in the phage-treated group testing positive for MAP colonized tissues compared to 6 out of 6 animals in the infected control group. Additionally, plaque assay results demonstrated the ability of phages to persist within the intestinal tract. Collectively, these results underscore the potential of orally administered MP cocktails as a highly effective intervention strategy to combat JD in dairy calves and by extension in the dairy industry.

Advances in phage-host interaction prediction: in silico method enhances the development of phage therapies.

Phages can specifically recognize and kill bacteria, which lead to important application value of bacteriophage in bacterial identification and typing, livestock aquaculture and treatment of human bacterial infection. Considering the variety of human-infected bacteria and the continuous discovery of numerous pathogenic bacteria, screening suitable therapeutic phages that are capable of infecting pathogens from massive phage databases has been a principal step in phage therapy design. Experimental methods to identify phage-host interaction (PHI) are time-consuming and expensive; high-throughput computational method to predict PHI is therefore a potential substitute. Here, we systemically review bioinformatic methods for predicting PHI, introduce reference databases and in silico models applied in these methods and highlight the strengths and challenges of current tools. Finally, we discuss the application scope and future research direction of computational prediction methods, which contribute to the performance improvement of prediction models and the development of personalized phage therapy.

Variable fitness effects of bacteriophage resistance mutations in Escherichia coli: implications for phage therapy.

Bacteria exposed to bactericidal treatment, such as antibiotics or bacteriophages (phages), often develop resistance. While phage therapy is proposed as a solution to the antibiotic resistance crisis, the bacterial resistance emerging during phage therapy remains poorly characterized. In this study, we examined a large population of phage-resistant extra-intestinal pathogenic Escherichia coli 536 clones that emerged from both in vitro (non-limited liquid medium) and in vivo (murine pneumonia) conditions. Genome sequencing uncovered a convergent mutational pattern in phage resistance mechanisms under both conditions, particularly targeting two cell-wall components, the K15 capsule and the lipopolysaccharide (LPS). This suggests that their identification in vivo could be predicted from in vitro assays. Phage-resistant clones exhibited a wide range of fitness according to in vitro tests, growth rate, and resistance to amoeba grazing, which could not distinguish between the K15 capsule and LPS mutants. In contrast, K15 capsule mutants retained virulence comparable to the wild-type strain, whereas LPS mutants showed significant attenuation in the murine pneumonia model. Additionally, we observed that resistance to the therapeutic phage through a nonspecific mechanism, such as capsule overproduction, did not systematically lead to co-resistance to other phages that were initially capable or incapable of infecting the wild-type strain. Our findings highlight the importance of incorporating a diverse range of phages in the design of therapeutic cocktails to target potential future phage-resistant clones effectively. IMPORTANCE: This study isolated more than 50 phage-resistant mutants from both in vitro and in vivo conditions, exposing an extra-intestinal pathogenic Escherichia coli strain to a single virulent phage. The characterization of these clones revealed several key findings: (1) mutations occurring during phage treatment affect the same pathways as those identified in vitro; (2) the resistance mechanisms are associated with the modification of two cell-wall components, with one involving receptor deletion (phage-specific mechanism) and the other, less frequent, involving receptor masking (phage-nonspecific mechanism); (3) an in vivo virulence assay demonstrated that the absence of the receptor abolishes virulence while masking the receptor preserves it; and (4) clones with a resistance mechanism nonspecific to a particular phage can remain susceptible to other phages. This supports the idea of incorporating diverse phages into therapeutic cocktails designed to collectively target both wild-type and phage-resistant strains, including those with resistance mechanisms nonspecific to a phage.

Elucidating the molecular properties and anti-mycobacterial activity of cysteine peptidase domain of D29 mycobacteriophage endolysin.

Emergence of antibiotic resistance in pathogenic Mycobacterium tuberculosis (Mtb) has elevated tuberculosis to a serious global threat, necessitating alternate solutions for its eradication. D29 mycobacteriophage can infect and kill several mycobacterial species including Mtb. It encodes an endolysin LysA to hydrolyze host bacteria peptidoglycan for progeny release. We previously showed that out of the two catalytically active domains of LysA [N-terminal domain (NTD) and lysozyme-like domain], NTD, when ectopically expressed in Mycobacterium smegmatis (Msm), is able to kill the bacterium nearly as efficiently as full-length LysA. Here, we dissected the functioning of NTD to develop it as a phage-derived small molecule anti-mycobacterial therapeutic. We performed a large-scale site-directed mutagenesis of the conserved residues in NTD and examined its structure, stability, and function using molecular dynamic simulations coupled with biophysical and biochemical experiments. Our data show that NTD functions as a putative cysteine peptidase with a catalytic triad composed of Cys41, His112, and Glu137, acting as nucleophile, base, and acid, respectively, and showing characteristics similar to the NlpC/P60 family of cysteine peptidases. Additionally, our peptidoglycan hydrolysis assays suggested that NTD hydrolyzes only mycobacterial peptidoglycan and does not act on Gram-positive and Gram-negative bacterial peptidoglycans. More importantly, the combined activity of exogenously added NTD and sub-lethal doses of anti-mycobacterial drugs kills Msm in vitro and exhibits disruption of pre-formed mycobacterial biofilm. We additionally show that NTD treatment increases the permeability of antibiotics in Msm, which reduces the minimum inhibitory concentration of the antibiotics. Collectively, we present NTD as a promising phage-derived therapeutic against mycobacteria.IMPORTANCEMycobacteriophages are the viruses that use mycobacteria as host for their progeny production and, in the process, kill them. Mycobacteriophages are, therefore, considered as promising alternatives to antibiotics for killing pathogenic Mycobacterium tuberculosis. The endolysin LysA produced by mycobacteriophage D29 plays an important role in host cell lysis and virion release. Our work presented here highlights the functioning of LysA's N-terminal catalytic domain (NTD) in order to develop it as phage-derived small molecule therapeutics. We show that combined treatment of exogenously added NTD and sub-lethal doses of anti-mycobacterial drugs kills M. smegmatis, shows synergism by reducing the minimum inhibitory concentration of these antibiotics, and exhibits disruption of pre-formed mature biofilm. These outcomes and our detailed biochemical and biophysical dissection of the protein further pave the way toward engineering and development of NTD as a promising therapeutic against mycobacterial infections such as tuberculosis.

Characterization of phage evolution and phage resistance in drug-resistant Stenotrophomonas maltophilia.

Phage therapy has become a viable antimicrobial treatment as an alternative to antibiotic treatment, with an increase in antibiotic resistance. Phage resistance is a major limitation in the therapeutic application of phages, and the lack of understanding of the dynamic changes between bacteria and phages constrains our response strategies to phage resistance. In this study, we investigated the changing trends of mutual resistance between Stenotrophomonas maltophilia (S. maltophilia) and its lytic phage, BUCT603. Our results revealed that S. maltophilia resisted phage infection through mutations in the cell membrane proteins, while the evolved phage re-infected the resistant strain primarily through mutations in structure-related proteins. Compared with the wild-type strain (SMA118), the evolved phage-resistant strain (R118-2) showed reduced virulence, weakened biofilm formation ability, and reduced resistance to aminoglycosides. In addition, the evolved phage BUCT603B1 in combination with kanamycin could inhibit the development of phage-resistant S. maltophilia in vitro and significantly improve the survival rate of S. maltophilia-infected mice. Altogether, these results suggest that in vitro characterization of bacteria-phage co-evolutionary relationships is a useful research tool to optimize phages for the treatment of drug-resistant bacterial infections.IMPORTANCEPhage therapy is a promising approach to treat infections caused by drug-resistant Stenotrophomonas maltophilia (S. maltophilia). However, the rapid development of phage resistance has hindered the therapeutic application of phages. In vitro evolutionary studies of bacteria-phage co-cultures can elucidate the mechanism of resistance development between phage and its host. In this study, we investigated the resistance trends between S. maltophilia and its phage and found that inhibition of phage adsorption is the primary strategy by which bacteria resist phage infection in vitro, while phages can re-infect bacterial cells by identifying other adsorption receptors. Although the final bacterial mutants were no longer infected by phages, they incurred a fitness cost that resulted in a significant reduction in virulence. In addition, the combination treatment with phage and aminoglycoside antibiotics could prevent the development of phage resistance in S. maltophilia in vitro. These findings contribute to increasing the understanding of the co-evolutionary relationships between phages and S. maltophilia.

Advances in the field of phage-based therapy with special emphasis on computational resources.

In the current era, one of the major challenges is to manage the treatment of drug/antibiotic-resistant strains of bacteria. Phage therapy, a century-old technique, may serve as an alternative to antibiotics in treating bacterial infections caused by drug-resistant strains of bacteria. In this review, a systematic attempt has been made to summarize phage-based therapy in depth. This review has been divided into the following two sections: general information and computer-aided phage therapy (CAPT). In the case of general information, we cover the history of phage therapy, the mechanism of action, the status of phage-based products (approved and clinical trials) and the challenges. This review emphasizes CAPT, where we have covered primary phage-associated resources, phage prediction methods and pipelines. This review covers a wide range of databases and resources, including viral genomes and proteins, phage receptors, host genomes of phages, phage-host interactions and lytic proteins. In the post-genomic era, identifying the most suitable phage for lysing a drug-resistant strain of bacterium is crucial for developing alternate treatments for drug-resistant bacteria and this remains a challenging problem. Thus, we compile all phage-associated prediction methods that include the prediction of phages for a bacterial strain, the host for a phage and the identification of interacting phage-host pairs. Most of these methods have been developed using machine learning and deep learning techniques. This review also discussed recent advances in the field of CAPT, where we briefly describe computational tools available for predicting phage virions, the life cycle of phages and prophage identification. Finally, we describe phage-based therapy's advantages, challenges and opportunities.

Revolutionising infection control: building the next generation of phage banks.

The escalating global burden of antimicrobial resistance (AMR) represents a critical public health challenge. This rise in antibiotic resistance is concomitant with heightened antibiotic consumption, with an estimated annual usage of 100,000 to 200,000 tons. A recent systematic review, which analysed data from 204 countries, reported that AMR was responsible for 4.95 million deaths in 2019 (Murray et al., 2022). The growing threat of AMR is imposing a significant financial burden on the global economy, with the CDC reporting an additional annual cost of $20 billion in the U.S. and €9 billion in Europe. The emerging field of bacteriophage therapy offers promising potential as a game-changer in the era of AMR. However, existing literature reveals numerous research gaps and technological challenges, including insufficient information on phage pharmacology, genomics, and a lack of preclinical and clinical data. In addition to conducting further research to address existing knowledge gaps, establishing phage banks in clinical facilities could be a transformative advancement in the fight against AMR.

Synthetic engineering and biological containment of bacteriophages.

The serious threats posed by drug-resistant bacterial infections and recent developments in synthetic biology have fueled a growing interest in genetically engineered phages with therapeutic potential. To date, many investigations on engineered phages have been limited to proof of concept or fundamental studies using phages with relatively small genomes or commercially available "phage display kits". Moreover, safeguards supporting efficient translation for practical use have not been implemented. Here, we developed a cell-free phage engineering and rebooting platform. We successfully assembled natural, designer, and chemically synthesized genomes and rebooted functional phages infecting gram-negative bacteria and acid-fast mycobacteria. Furthermore, we demonstrated the creation of biologically contained phages for the treatment of bacterial infections. These synthetic biocontained phages exhibited similar properties to those of a parent phage against lethal sepsis in vivo. This efficient, flexible, and rational approach will serve to accelerate phage biology studies and can be used for many practical applications, including phage therapy.

Zebrafish as an effective model for evaluating phage therapy in bacterial infections: a promising strategy against human pathogens.

The escalating prevalence of antibiotic-resistant bacterial infections necessitates urgent alternative therapeutic strategies. Phage therapy, which employs bacteriophages to specifically target pathogenic bacteria, emerges as a promising solution. This review examines the efficacy of phage therapy in zebrafish models, both embryos and adults, which are proven and reliable for simulating human infectious diseases. We synthesize findings from recent studies that utilized these models to assess phage treatments against various bacterial pathogens, including Enterococcus faecalis, Pseudomonas aeruginosa, Mycobacterium abscessus, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, and Escherichia coli. Methods of phage administration, such as circulation injection and bath immersion, are detailed alongside evaluations of survival rates and bacterial load reductions. Notably, combination therapies of phages with antibiotics show enhanced efficacy, as evidenced by improved survival rates and synergistic effects in reducing bacterial loads. We also discuss the transition from zebrafish embryos to adult models, emphasizing the increased complexity of immune responses. This review highlights the valuable contribution of the zebrafish model to advancing phage therapy research, particularly in the face of rising antibiotic resistance and the urgent need for alternative treatments.

Pseudomonas aeruginosa ventricular assist device infections: findings from ineffective phage therapies in five cases.

Left ventricular assist devices (LVAD) are increasingly used for management of heart failure; infection remains a frequent complication. Phage therapy has been successful in a variety of antibiotic refractory infections and is of interest in treating LVAD infections. We performed a retrospective review of four patients that underwent five separate courses of intravenous (IV) phage therapy with concomitant antibiotic for treatment of endovascular Pseudomonas aeruginosa LVAD infection. We assessed phage susceptibility, bacterial strain sequencing, serum neutralization, biofilm activity, and shelf-life of phage preparations. Five treatments of one to four wild-type virulent phage(s) were administered for 14-51 days after informed consent and regulatory approval. There was no successful outcome. Breakthrough bacteremia occurred in four of five treatments. Two patients died from the underlying infection. We noted a variable decline in phage susceptibility following three of five treatments, four of four tested developed serum neutralization, and prophage presence was confirmed in isolates of two tested patients. Two phage preparations showed an initial titer drop. Phage biofilm activity was confirmed in two. Phage susceptibility alone was not predictive of clinical efficacy in P. aeruginosa endovascular LVAD infection. IV phage was associated with serum neutralization in most cases though lack of clinical effect may be multifactorial including presence of multiple bacterial isolates with varying phage susceptibility, presence of prophages, decline in phage titers, and possible lack of biofilm activity. Breakthrough bacteremia occurred frequently (while the organism remained susceptible to administered phage) and is an important safety consideration.

The zebrafish embryo model: unveiling its potential for investigating phage therapy against methicillin-resistant Staphylococcus aureus infection.

Staphylococcus aureus is a pathogenic bacterium responsible for a broad spectrum of infections, including cutaneous, respiratory, osteoarticular, and systemic infections. It poses a significant clinical challenge due to its ability to develop antibiotic resistance. This resistance limits therapeutic options, increases the risk of severe complications, and underscores the urgent need for new strategies to address this threat, including the investigation of treatments complementary to antibiotics. The evaluation of novel antimicrobial agents often employs animal models, with the zebrafish embryo model being particularly interesting for studying host-pathogen interactions, establishing itself as a crucial tool in this field. For the first time, this study presents a zebrafish embryo model for the in vivo assessment of bacteriophage efficacy against S. aureus infection. A localized infection was induced by microinjecting either methicillin-resistant S. aureus (MRSA) or methicillin-susceptible S. aureus (MSSA). Subsequent treatments involved administering either bacteriophage, vancomycin (the reference antibiotic for MRSA), or a combination of both via the same route to explore potential synergistic effects. Our findings indicate that the bacteriophage was as effective as vancomycin in enhancing survival rates, whether used alone or in combination. Moreover, bacteriophage treatment appears to be even more effective in reducing the bacterial load in S. aureus-infected embryos post-treatment than the antibiotic. Our study validates the use of the zebrafish embryo model and highlights its potential as a valuable tool in assessing bacteriophage efficacy treatments in vivo.

Host immunity involvement in the outcome of phage therapy against hypervirulent Klebsiella pneumoniae infections.

Highly encapsulated hypervirulent Klebsiella pneumoniae (hvKp) causes severe infections. Bacteriophage therapy, an antibiotic alternative, effectively treats bacterial infections. Phage φFK1979 encoding polysaccharide depolymerases can target and disarm the capsule of hvKp FK1979, showing promise against FK1979 infection. Resistant strains induced by φFK1979 are possibly eliminated by host immunity and new phage phiR3 targeting them. We constructed varied immunocompromised FK1979 infection mouse models to assess the therapy efficacy of φFK1979 alone or in combination with phiR3. Survival rates, bacterial loads, histopathology, inflammation, and immune cell distribution of mice were studied. Prompt and adequate administration of φFK1979, rather than phiR3, significantly improved survival rates in mice with different immune statuses. However, immunocompromised mice showed lower efficacy due to reduced tolerance to low-virulence φFK1979-resistant bacteria compared to immunocompetent mice. Adding phiR3 sequentially greatly enhanced therapy efficacy for them, leading to increased survival rates and notable improvements in pathology and inflammation. Immunocompetent mice exhibited the most favorable response to φFK1979 monotherapy, as their immune system cleared φFK1979-resistant bacteria while avoiding a robust response to phiR3 combating φFK1979-resistant bacteria. This study revealed host immunity involvement in the outcome of phage therapy against infections and introduced, for the first time, personalized phage therapy strategies for hvKp-infected mice with varying immune statuses.IMPORTANCEHypervirulent Klebsiella pneumoniae (hvKp), with high capsular polysaccharide production, can cause severe invasive infections. Capsule-targeting phage poses the potential to fight against hvKp. We previously elucidated that the capsule-targeting phage induces resistance in hvKp, while phage-resistant strains exhibit sensitivity to host innate immunity and new phages targeting them. This indicated that phage-resistant strains can be eliminated by the immune system in immunocompetent patients, whereas they may require treatment with phages targeting resistant bacteria in immunocompromised patients. HvKp can infect individuals with varying immune statuses, including both immunocompetent and immunocompromised/deficient patients. This study, for the first time, developed personalized phage therapy strategies for hvKp-infected mice with different immune statuses, optimizing phage therapy against hvKp infections. This research is expected to provide a theoretical foundation and novel insights for clinical phage therapy against hvKp infections, offering significant societal benefits and clinical value.

Phage K exposure generates phage-neutralizing activity in a rabbit model-humoral receptiveness may impact efficacy of phage therapy.

Phage therapy has not been established in the clinical routine, in part due to uncertainties concerning efficacy and immunogenicity. Here, three rabbits were immunized against staphylococcal phage K to assess viral potency in the presence of immunized serum. Three rabbits received weekly intramuscular injections of ~1010±1 pfu/mL phage K. Phage K-specific IgG formation was measured by an enzyme-linked immunosorbent assay (ELISA); phage inactivation was assessed by calculating K-rates. Using transmission electron microscopy (TEM) and immunogold labeling, antibody binding to phage K was visualized. This was numerically assessed by objective imaging analysis comparing the relative distances of each gold particle to the nearest phage head and tail structure. Immunization led to a strong IgG response, plateauing 7 days after the last phage injection. There was no significant correlation between K-rate and antibody titer over time. TEM showed IgG binding to the head structure of phage K. Image analysis showed a significant reduction in relative distances between antibodies and phage head structures when comparing samples from day 0 and day 28 (P < 0.0001). These results suggest that while individual serum analysis for antibodies against therapeutic phage bears consideration prior to and with prolonged therapy, during phage application, the formation of specific antibodies against phage may only partially explain decreased phage potency in the presence of immunized serum. Instead, other factors may contribute to an individual's "humoral receptiveness" to phage therapy. Future investigations should be directed toward the identification of the humoral factors that have the most significant predictive value on phage potency in vivo.

Phage Therapy for Antibiotic-Resistant Bacterial Infections.

Antibiotic resistance in bacterial pathogens presents a substantial threat to the control of infectious diseases. Development of new classes of antibiotics has slowed in recent years due to pressures of cost and market profitability, and there is a strong need for new antimicrobial therapies. The therapeutic use of bacteriophages has long been considered, with numerous anecdotal reports of success. Interest in phage therapy has been renewed by recent clinical successes in case studies with personalized phage cocktails, and several clinical trials are in progress. We discuss recent progress in the therapeutic use of phages and contemplate the key factors influencing the opportunities and challenges. With strong safety profiles, the main challenges of phage therapeutics involve strain variation among clinical isolates of many pathogens, battling phage resistance, and the potential limitations of host immune responses. However, the opportunities are considerable, with the potential to enhance current antibiotic efficacy, protect newly developed antibiotics, and provide a last resort in response to complete antibiotic failure.

The phage fought the cells, and the phage won: a satellite symposium at the ASV 2023 annual meeting.

This satellite symposium was focused on the molecular arms race between bacteria and their predators, the bacteriophages: who's the friend and who's the foe? This Gem recounts highlights of the talks and presents food for thought and additional reflections on the current state of the field.

Making the leap from technique to treatment - genetic engineering is paving the way for more efficient phage therapy.

Bacteriophages (phages) are viruses specific to bacteria that target them with great efficiency and specificity. Phages were first studied for their antibacterial potential in the early twentieth century; however, their use was largely eclipsed by the popularity of antibiotics. Given the surge of antimicrobial-resistant strains worldwide, there has been a renaissance in harnessing phages as therapeutics once more. One of the key advantages of phages is their amenability to modification, allowing the generation of numerous derivatives optimised for specific functions depending on the modification. These enhanced derivatives could display higher infectivity, expanded host range or greater affinity to human tissues, where some bacterial species exert their pathogenesis. Despite this, there has been a noticeable discrepancy between the generation of derivatives in vitro and their clinical application in vivo. In most instances, phage therapy is only used on a compassionate-use basis, where all other treatment options have been exhausted. A lack of clinical trials and numerous regulatory hurdles hamper the progress of phage therapy and in turn, the engineered variants, in becoming widely used in the clinic. In this review, we outline the various types of modifications enacted upon phages and how these modifications contribute to their enhanced bactericidal function compared with wild-type phages. We also discuss the nascent progress of genetically modified phages in clinical trials along with the current issues these are confronted with, to validate it as a therapy in the clinic.