Phage therapy for pulmonary infections: lessons from clinical experiences and key considerations.

Lower respiratory tract infections lead to significant morbidity and mortality. They are increasingly caused by multidrug-resistant pathogens, notably in individuals with cystic fibrosis, hospital-acquired pneumonia and lung transplantation. The use of bacteriophages (phages) to treat bacterial infections is gaining growing attention, with numerous published cases of compassionate treatment over the last few years. Although the use of phages appears safe, the lack of standardisation, the significant heterogeneity of published studies and the paucity of robust efficacy data, alongside regulatory hurdles arising from the existing pharmaceutical legislation, are just some of the challenges phage therapy has to overcome. In this review, we discuss the lessons learned from recent clinical experiences of phage therapy for the treatment of pulmonary infections. We review the key aspects, opportunities and challenges of phage therapy regarding formulations and administration routes, interactions with antibiotics and the immune system, and phage resistance. Building upon the current knowledge base, future pre-clinical studies using emerging technologies and carefully designed clinical trials are expected to enhance our understanding and explore the therapeutic potential of phage therapy.

Identifying Causative Microorganisms in Left Ventricular Assist Device Infections as a Guide for Developing Bacteriophage Therapy.

BACKGROUND: As more left ventricular-assist devices (LVADs) are implanted, multidrug-resistant LVAD infections are becoming increasingly common, partly due to bacterial biofilm production. To aid in developing bacteriophage therapy for LVAD infections, we have identified the most common bacterial pathogens that cause LVAD driveline infections (DLIs) in our heart transplant referral center. MATERIALS AND METHODS: We studied a retrospective cohort of patients who received LVADs from November 2003 to August 2017 to identify the common causative organisms of LVAD infection. We also studied a prospective cohort of patients diagnosed with DLIs from October 2018 to May 2019 to collect bacterial strains from DLIs for developing bacteriophages to lyse causative pathogens. LVAD infections were classified as DLI, bacteremia, and pump/device infections in the retrospective cohort. RESULTS: In the retrospective cohort of 582 patients, 186 (32.0%) developed an LVAD infection, with 372 microbial isolates identified. In the prospective cohort, 96 bacterial strains were isolated from 54 DLIs. The microorganisms causing DLIs were similar in the two cohorts; the most common isolate was Staphylococcus aureus. We identified 6 prospective S. aureus strains capable of biofilm formation. We developed 3 bacteriophages that were able to lyse 5 of 6 of the biofilm-forming S. aureus strains. CONCLUSIONS: Similar pathogens caused LVAD DLIs in our retrospective and prospective cohorts, indicating our bacterial strain bank will be representative of future DLIs. Our banked bacterial strains will be useful in developing phage cocktails that can lyse ≥80% of the bacteria causing LVAD infections at our institution.

The Safety and Toxicity of Phage Therapy: A Review of Animal and Clinical Studies.

Increasing rates of infection by antibiotic resistant bacteria have led to a resurgence of interest in bacteriophage (phage) therapy. Several phage therapy studies in animals and humans have been completed over the last two decades. We conducted a systematic review of safety and toxicity data associated with phage therapy in both animals and humans reported in English language publications from 2008-2021. Overall, 69 publications met our eligibility criteria including 20 animal studies, 35 clinical case reports or case series, and 14 clinical trials. After summarizing safety and toxicity data from these publications, we discuss potential approaches to optimize safety and toxicity monitoring with the therapeutic use of phage moving forward. In our systematic review of the literature, we found some adverse events associated with phage therapy, but serious events were extremely rare. Comprehensive and standardized reporting of potential toxicities associated with phage therapy has generally been lacking in the published literature. Structured safety and tolerability endpoints are necessary when phages are administered as anti-infective therapeutics.

Bacteriophage therapy: recent developments and applications of a renaissant weapon.

Antimicrobial resistance is a global health problem and one of the leading concerns in healthcare sector. Bacteriophages are antibacterial agents ubiquitous in nature. With increase in antibiotic resistance, use of bacteriophages as therapeutics has become resurgent in recent times. This review focuses on the recent developments in phage therapy and its applications with respect to human infections, animal, food and environment. Moreover, use of phage proteins, bioengineered bacteriophages, and phage derived vaccines is also highlighted. Additionally, the limitations and challenges with regard to implementation of phage therapy, host safety and immune responses are also reviewed in this article.

Preclinical Assessment of Bacteriophage Therapy against Experimental Acinetobacter baumannii Lung Infection.

Respiratory infections caused by multidrug-resistant Acinetobacter baumannii are difficult to treat and associated with high mortality among critically ill hospitalized patients. Bacteriophages (phages) eliminate pathogens with high host specificity and efficacy. However, the lack of appropriate preclinical experimental models hampers the progress of clinical development of phages as therapeutic agents. Therefore, we tested the efficacy of a purified lytic phage, vB_AbaM_Acibel004, against multidrug-resistant A. baumannii clinical isolate RUH 2037 infection in immunocompetent mice and a human lung tissue model. Sham- and A. baumannii-infected mice received a single-dose of phage or buffer via intratracheal aerosolization. Group-specific differences in bacterial burden, immune and clinical responses were compared. Phage-treated mice not only recovered faster from infection-associated hypothermia but also had lower pulmonary bacterial burden, lower lung permeability, and cytokine release. Histopathological examination revealed less inflammation with unaffected inflammatory cellular recruitment. No phage-specific adverse events were noted. Additionally, the bactericidal effect of the purified phage on A. baumannii was confirmed after single-dose treatment in an ex vivo human lung infection model. Taken together, our data suggest that the investigated phage has significant potential to treat multidrug-resistant A. baumannii infections and further support the development of appropriate methods for preclinical evaluation of antibacterial efficacy of phages.

Therapeutic Perspectives and Mechanistic Insights of Phage Therapy in Allotransplantation.

Bacterio(phages) are bacteria-infecting viruses that employ host translation machinery to replicate, and upon cell lysis, release new particles into the environment. As a result, phages are prey-specific, thus making targeted phage therapy (PT) possible. Indeed, pre- and posttransplant bacterial infections pose a substantial risk to allograft recipients in their clinical course. Moreover, with the increasing threat of antibiotic resistance, the interest in PT as a potential solution to the crisis of multidrug-resistant bacterial pathogens has rapidly grown. Although little is known about the specific characteristics of the phage-directed immune responses, recent studies indicate phages exert anti-inflammatory and immunomodulatory functions, which could be beneficial in allotransplantation (allo-Tx). PT targeting multidrug-resistant Klebsiella pneumoniae, Mycobacterium abscessus, and Pseudomonas aeruginosa have been successfully applied in renal, lung, and liver allo-Tx patients. In parallel, the gastrointestinal microbiota appears to influence allo-Tx immunity by modulating the endoplasmic reticulum stress and autophagy signaling pathways through hepatic EP4/CHOP/LC3B platforms. This review highlights the current relevant immunobiology, clinical developments, and management of PT, and lays the foundation for future potential standard care use of PT in allo-Tx to mitigate early allograft dysfunction and improve outcomes. In conclusion, with novel immunobiology and metabolomics insights, harnessing the potential of PT to modulate microbiota composition/diversity may offer safe and effective refined therapeutic means to reduce risks of infections and immunosuppression in allo-Tx recipients.

Phage Therapy Application to Counteract Pseudomonas aeruginosa Infection in Cystic Fibrosis Zebrafish Embryos.

Antimicrobial resistance, a major consequence of diagnostic uncertainty and antimicrobial overprescription, is an increasingly recognized cause of severe infections, complications, and mortality worldwide with a huge impact on our society and on the health system. In particular, patients with compromised immune systems or pre-existing and chronic pathologies, such as cystic fibrosis (CF), are subjected to frequent antibiotic treatments to control the infections with the appearance and diffusion of multidrug resistant isolates. Therefore, there is an urgent need to address alternative therapies to counteract bacterial infections. Use of bacteriophages, the natural enemies of bacteria, can be a possible solution. The protocol detailed in this work describes the application of phage therapy against Pseudomonas aeruginosa infection in CF zebrafish embryos. Zebrafish embryos were infected with P. aeruginosa to demonstrate that phage therapy is effective against P. aeruginosa infections as it reduces lethality, bacterial burden and pro-inflammatory immune response in CF embryos.

Topical application of bacteriophages for treatment of wound infections.

Wound infections associated with multidrug-resistant (MDR) bacteria are one of the important threats to public health. Bacteriophage (phage) therapy is a promising alternative or supplementary therapeutic approach to conventional antibiotics for combating MDR bacterial infections. In recent years, significant effort has been put into the development of phage formulations and delivery methods for topical applications, along with preclinical and clinical uses of phages for the treatment of acute and chronic wound infections. This paper reviews the application of phages for wound infections, with focus on the current status of phage formulations (including liquid, semi-solid and liposome-encapsulated formulations, phage-immobilized wound dressings), safety and efficacy assessment in clinical settings and major challenges to overcome.

Safety of bacteriophage therapy in severe Staphylococcus aureus infection.

In this single-arm non-comparative trial, 13 patients in an Australian hospital with severe Staphylococcus aureus infections were intravenously administered a good manufacturing practice-quality preparation of three Myoviridae bacteriophages (AB-SA01) as adjunctive therapy. AB-SA01 was intravenously administered twice daily for 14 d and the clinical, haematological and blood biochemical parameters of the recipients were monitored for 90 d. The primary outcome was the assessment of safety and tolerability (that is, pain and redness at the infusion site and systemic adverse reactions, such as fever, tachycardia, hypotension, diarrhoea or abdominal pain and the development of renal or hepatic dysfunction). No adverse reactions were reported, and our data indicate that AB-SA01 administered in this way is safe in severe S. aureus infections, including infective endocarditis and septic shock. Future controlled trials will be needed to determine the efficacy of AB-SA01 but no phage resistance evolved in vivo and the measurements of bacterial and phage kinetics in blood samples suggest that 12 h dosing of 109 plaque-forming units may be a rational basis for further studies.

Biological challenges of phage therapy and proposed solutions: a literature review.

Introduction: In light of the emergence of antibiotic-resistant bacteria, phage (bacteriophage) therapy has been recognized as a potential alternative or addition to antibiotics in Western medicine for use in humans.Areas covered: This review assessed the scientific literature on phage therapy published between 1 January 2007 and 21 October 2019, with a focus on the successes and challenges of this prospective therapeutic.Expert opinion: Efficacy has been shown in animal models and experimental findings suggest promise for the safety of human phagotherapy. Significant challenges remain to be addressed prior to the standardization of phage therapy in the West, including the development of phage-resistant bacteria; the pharmacokinetic complexities of phage; and any potential human immune response incited by phagotherapy.

Effect of serum anti-phage activity on colibacillosis control by repeated phage therapy in broilers.

Anti-phage activity of serum is of importance in repeated phage therapy. Higher serum anti-phage activity has been associated with greater susceptibility of phages to neutralisation and phage therapy failure. In this study, in vivo and in vitro survivability and immunogenicity of four coliphages (TM1, TM2, TM3 and TM4) were investigated in naive chickens and chickens pre-immunised with phage TM1. Furthermore, two phages that displayed different survivability and immunogenicity (TM1 and TM3) were compared with respect to their efficacy in treating naive or pre-immunised (TM1) chickens suffering from colibacillosis. The efficacy of the treatments was evaluated based on body weight, relative organ weights, mortality, E. coli counts in the lungs as well as severity and frequency of internal organ lesions. At the end of the experiment, both naive and pre-immunised chickens treated with TM3 showed significantly lower mortality and higher body weights than untreated chickens and those treated with TM1. The same trend was observed in incidence and severity of organ lesions as well as relative spleen weight. However, naive chickens treated with TM1 also showed a shortened inflammation period as indicated by spleen weights. E. coli counts in the lungs of chicken treated with TM3 were lower than those of chickens treated with TM1 on days 3 and 10 post challenge. These data indicate that the outcome of phage therapy and the impact of serum anti-phage activity are highly phage-type dependent in broilers.

Phage-Derived Antibacterials: Harnessing the Simplicity, Plasticity, and Diversity of Phages.

Despite the successful use of antibacterials, the emergence of multidrug-resistant bacteria has become a serious threat to global healthcare. In this era of antibacterial crisis, bacteriophages (phages) are being explored as an antibacterial treatment option since they possess a number of advantages over conventional antibacterials, especially in terms of specificity and biosafety; phages specifically lyse target bacteria while not affecting normal and/or beneficial bacteria and display little or no toxicity in that they are mainly composed of proteins and nucleic acids, which consequently significantly reduces the time and cost involved in antibacterial development. However, these benefits also create potential issues regarding antibacterial spectra and host immunity; the antibacterial spectra being very narrow when compared to those of chemicals, with the phage materials making it possible to trigger host immune responses, which ultimately disarm antibacterial efficacy upon successive treatments. In addition, phages play a major role in horizontal gene transfer between bacterial populations, which poses serious concerns for the potential of disastrous consequences regarding antibiotic resistance. Fortunately, however, recent advancements in synthetic biology tools and the speedy development of phage genome resources have allowed for research on methods to circumvent the potentially disadvantageous aspects of phages. These novel developments empower research which goes far beyond traditional phage therapy approaches, opening up a new chapter for phage applications with new antibacterial platforms. Herein, we not only highlight the most recent synthetic phage engineering and phage product engineering studies, but also discuss a new proof-of-concept for phage-inspired antibacterial design based on the studies undertaken by our group.

A Wake-Up Call: We Need Phage Therapy Now.

The rise of multidrug-resistant bacteria has resulted in an increased interest in phage therapy, which historically preceded antibiotic treatment against bacterial infections. To date, there have been no reports of serious adverse events caused by phages. They have been successfully used to cure human diseases in Eastern Europe for many decades. More recently, clinical trials and case reports for a variety of indications have shown promising results. However, major hurdles to the introduction of phage therapy in the Western world are the regulatory and legal frameworks. Present regulations may take a decade or longer to be fulfilled. It is of urgent need to speed up the availability of phage therapy.

Characterization of bacteriophage HN48 and its protective effects in Nile tilapia Oreochromis niloticus against Streptococcus agalactiae infections.

Streptococcus agalactiae is a causative agent responsible for massive mortalities of tilapia that has led to catastrophic losses to tilapia culture globally. Bacteriophages represent a new class of antimicrobials against bacteria. In this study, we characterized the bacteriophage HN48, which formed small and round-transparent plaques on a double-layer plate. With a hexagonal head and a long tail, this phage may belong to the Caudovirales according to the International Committee on Taxonomy of Viruses. HN48 was found to have a relatively wide and highly specific host range, to be sensitive to high temperature (60-80°C) and low pH (3-5), and to be relatively stable at alkaline pH (8-10). Intraperitoneal injection with HN48 had no adverse effects on tilapia and effectively inactivated the bacteria in the kidney. Fish that received phage therapy had 60% ± 3.3% survival rates and a delayed mean death time of about 3 days when compared to the control group. To the best of knowledge, this is the first study of tilapia streptococcal phage. Overall, the results indicated that phage HN48 could prevent tilapia from experimental S. agalactiae infection, suggesting it has the potential to control this disease.

Combined bacteriophages and antibiotics as an efficient therapy against VRE Enterococcus faecalis in a mouse model.

Clinical applications of bacteriophage therapy have been recently gathering significant attention worldwide, used mostly as rescue therapy in cases of near-fatal antibiotic failure. Thus, clinically relevant in-vivo models presenting both short- and long-term implications of phage therapy given as rescue treatment for fulminant infections are of highest importance. In this study, a cocktail consisting of two lytic bacteriophages was used to evaluate the therapeutic efficacy of phage therapy as a rescue treatment for severe septic peritonitis in a mouse model. We established that a single injection of the bacteriophage cocktail was sufficient to completely reverse a 100% mortality trend caused by Vancomycin-Resistant Enterococcus faecalis, with significant improvement in both the clinical state and laboratory test results, and without harmful effects on the microbiome. The combination of bacteriophages with a suboptimal antibiotic regimen imparts an additional beneficial effect on the treatment success.

[Bacteriophages in the battle against multidrug resistant bacteria]. / Bacteriofagen in de strijd tegen multiresistente bacteriën.

Bacteriophages are viruses that infect bacteria. They are highly specific for a bacterial species. The so-called 'lytic phages' can lyse bacteria when they infect them; these phages can be used to treat bacterial infections. Despite a century of experience with phage therapy, the evidence for clinical efficacy is limited. Side effects are generally considered to be mild. The selection, preparation and administration of phages for therapy is laborious, and investigations into the clinical benefits are not easy. More research is needed, also in the face of the increasing antimicrobial resistance.

Refractory Pseudomonas Bacteremia in a 2-Year-Old Sterilized by Bacteriophage Therapy.

Here, we report a complex case that involved a pediatric patient who experienced recalcitrant multidrug-resistant Pseudomonas aeruginosa infection complicated by bacteremia/sepsis; our antibacterial options were limited because of resistance, allergies, and suboptimal source control. A cocktail of 2 bacteriophages targeting the infectious organism introduced on 2 separate occasions sterilized the bacteremia.

In Vivo Studies on the Influence of Bacteriophage Preparations on the Autoimmune Inflammatory Process.

Phage preparations used for phage therapy may have not only direct antibacterial action but also immunomodulating effects mediated by phages themselves as well as by bacterial antigens. Therefore phage application in patients with immune disorders, and especially with autoimmune diseases, requires special attention. The aim of this study was to investigate the effect of phage lysates (staphylococcal phages A3/R, phi200, and MS-1 cocktail, enterococcal phage 15/P, Pseudomonas phage 119x, and E. coli T4 phage) as well as purified T4 phage on the course of murine collagen-induced arthritis (CIA), commonly used as an animal model of rheumatoid arthritis. Intraperitoneal application of phage lysates or purified T4 phage did not aggravate the course of autoimmune joint disease. Moreover, although endotoxins are known to potentiate CIA, the systemic administration of phage lysate of Pseudomonas aeruginosa, which contains debris of this Gram-negative bacillus, did not significantly influence CIA although the sonicate of the corresponding bacterial strain did. Interestingly, a purified T4 phage revealed some anti-inflammatory activity when applied under the therapeutic scheme. Our preliminary results do not suggest that phages may aggravate the symptoms of rheumatoid arthritis. In contrast T4 phage may even exert an immunosuppressive effect.