Andrzej Górski

Bacteriophage endolysins as a novel class of antibacterial agents.

Endolysins are double-stranded DNA bacteriophage-encoded peptidoglycan hydrolases produced in phage-infected bacterial cells toward the end of the lytic cycle. They reach the peptidoglycan through membrane lesions formed by holins and cleave it, thus, inducing lysis of the bacterial cell and enabling progeny virions to be released. Endolysins are also capable of degrading peptidoglycan when applied externally (as purified recombinant proteins) to the bacterial cell wall, which also results in a rapid lysis of the bacterial cell. The unique ability of endolysins to rapidly cleave peptidoglycan in a generally species-specific manner renders them promising potential antibacterial agents. Originally developed with a view to killing bacteria colonizing mucous membranes (with the first report published in 2001), endolysins also hold promise for the treatment of systemic infections. As potential antibacterials, endolysins possess several important features, for instance, a novel mode of action, a narrow antibacterial spectrum, activity against bacteria regardless of their antibiotic sensitivity, and a low probability of developing resistance. However, there is only one report directly comparing the activity of an endolysin with that of an antibiotic, and no general conclusions can be drawn regarding whether lysins are more effective than traditional antibiotics. The results of the first preclinical studies indicate that the most apparent potential problems associated with endolysin therapy (e.g., their immunogenicity, the release of proinflammatory components during bacteriolysis, or the development of resistance), in fact, may not seriously hinder their use. However, all data regarding the safety and therapeutic effectiveness of endolysins obtained from preclinical studies must be ultimately verified by clinical trials. This review discusses the prophylactic and therapeutic applications of endolysins, especially with respect to their potential use in human medicine. Additionally, we outline current knowledge regarding the structure and natural function of the enzymes in phage biology, including the most recent findings.

Bacteriophage penetration in vertebrates

Bacteriophages are viruses that infect bacteria. They are the most numerous life forms on earth. As antibiotic resistance is becoming an increasingly worldwide challenge, bacteriophages as potential antimicrobial agents are being more intensively explored. Some very important questions involve their ability to penetrate higher organisms, as this determines potential phage activity in antibacterial treatment. Higher organisms are widely exposed to bacteriophages, which penetrate them quite freely. Bacteriophage activity can be influenced by specific antibodies which, together with the nonspecific immune system, can contribute to their rapid clearance from the organism. Bacteriophages can also interact directly with mammalian cells and even play a role in the development of some nonbacterial diseases, although they are not able to multiply in these cells. All aspects of the interaction between phages and higher organism are of interest and importance for further medical and biochemical applications.

The influence of external factors on bacteriophages—review

The ability of bacteriophages to survive under unfavorable conditions is highly diversified. We summarize the influence of different external physical and chemical factors, such as temperature, acidity, and ions, on phage persistence. The relationships between a phage’s morphology and its survival abilities suggested by some authors are also discussed. A better understanding of the complex problem of phage sensitivity to external factors may be useful not only for those interested in pharmaceutical and agricultural applications of bacteriophages, but also for others working with phages.

Clinical aspects of phage therapy.

Phage therapy (PT) is a unique method of treatment of bacterial infections using bacteriophages (phages)-viruses that specifically kill bacteria, including their antibiotic-resistant strains. Over the last decade a marked increase in interest in the therapeutic use of phages has been observed, which has resulted from a substantial rise in the prevalence of antibiotic resistance of bacteria, coupled with an inadequate number of new antibiotics. The first, and so far the only, center of PT in the European Union is the Phage Therapy Unit (PTU) established at the Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Wrocław, Poland in 2005. This center continues the rich tradition of PT in Poland, which dates from the early 1920s. The main objective of this chapter is to present a detailed retrospective analysis of the results of PT of 153 patients with a wide range of infections resistant to antibiotic therapy admitted for treatment at the PTU between January 2008 and December 2010. Analysis includes the evaluation of both the efficacy and the safety of PT. In general, data suggest that PT can provide good clinical results in a significant cohort of patients with otherwise untreatable chronic bacterial infections and is essentially well tolerated. In addition, the whole complex procedure employed to obtain and characterize therapeutic phage preparations, as well as ethical aspects of PT, is discussed.

Transplantation of autologous olfactory ensheathing cells in complete human spinal cord injury.

Numerous studies in animals have shown the unique property of olfactory ensheathing cells to stimulate regeneration of lesioned axons in the spinal cord. In a Phase I clinical trial, we assessed the safety and feasibility of transplantation of autologous mucosal olfactory ensheathing cells and olfactory nerve fibroblasts in patients with complete spinal cord injury. Six patients with chronic thoracic paraplegia (American Spinal Injury Association class A-ASIA A) were enrolled for the study. Three patients were operated, and three served as a control group. The trial protocol consisted of pre- and postoperative neurorehabilitation, olfactory mucosal biopsy, culture of olfactory ensheathing cells, and intraspinal cell grafting. Patients clinical state was evaluated by clinical, neurophysiological, and radiological tests. There were no adverse findings related to olfactory mucosa biopsy or transplantation of olfactory ensheathing cells at 1 year after surgery. There was no evidence of neurological deterioration, neuropathic pain, neuroinfection, or tumorigenesis. In one cell-grafted patient, an asymptomatic syringomyelia was observed. Neurological improvement was observed only in transplant recipients. The first two operated patients improved from ASIA A to ASIA C and ASIA B. Diffusion tensor imaging showed restitution of continuity of some white matter tracts throughout the focus of spinal cord injury in these patients. The third operated patient, although remaining ASIA A, showed improved motor and sensory function of the first spinal cords segments below the level of injury. Neurophysiological examinations showed improvement in spinal cord transmission and activity of lower extremity muscles in surgically treated patients but not in patients receiving only neurorehabilitation. Observations at 1 year indicate that the obtaining, culture, and intraspinal transplantation of autologous olfactory ensheathing cells were safe and feasible. The significance of the neurological improvement in the transplant recipients and the extent to which the cell transplants contributed to it will require larger numbers of patients.

Bacteriophage Therapy of Bacterial Infections: An Update of our Institute’s Experience

1307 patients with suppurative bacterial infections caused by multidrug-resistant bacteria of different species were treated with specific bacteriophages (BP). BP therapy was highly effective; full recovery was noted in 1123 cases (85.9%). In 134 cases (10.9%) transient improvement was observed and only in 50 cases (3.8%) was BP treatment found to be ineffective. The results confirm the high effectiveness of BP therapy in combating bacterial infections which do not respond to treatment with the available antibiotics.

The Phage Therapy Paradigm: Prêt-à-Porter or Sur-mesure?

The present opinion is the result of discussions on the future of phage therapy (personalized or large-scale uniform therapy?) during the first International Congress on Viruses of Microbes, held at the Institut Pasteur in Paris on June 21–25, 2010. Antibiotics are becoming ineffective as important bacterial pathogens evolve to outsmart them. Yet the antibiotic pipeline is running dry with only a few new antibacterial drugs expected to make it to the market in the foreseeable future. Bacteria that are resistant to all available antibacterial drugs, so-called superbugs, are emerging worldwide. Evolutionary ecology might inform practical attempts to bring these pathogens under stronger human control (1). In this context, various laboratories worldwide and a handful of small pharmaceutical companies are turning to (bacterio)phages (2). Phages are natural viruses that specifically infect bacteria. They are (among) the most abundant and ubiquitous lifelike entities on Earth and coevolve with their hosts, the bacteria. Lytic phages bind to receptors on the bacterial cell surface, inject their genetic material, use the bacterium’s reproductive machinery to replicate and subsequently destroy (lyse) the bacterium, irrespective of its resistance to antibiotics, releasing the newly formed phages to seek out new hosts. In 1919, d’Herelle used phages to treat dysentery in Paris, in what was probably the first attempt to use phages therapeutically. d’Herelle eventually developed a commercial laboratory in Paris that produced phage preparations against

The potential role of endogenous bacteriophages in controlling invading pathogens

Abstract.Bacteriophages (phages) are omnipresent in our environment, and recent studies highlight their potential impact on the microbial world. Phages can also be present in mammalian organisms, including man (intestines, oral cavity, urine, sputum and serum). Data are available which suggest that those endogenous phages could play an important role in eliminating bacteria and regulating the body ecosystem. Furthermore, our most recent findings suggest that phages can exert immunosuppressive action in the gut, helping control local inflammatory and autoimmune reactions, and demonstrate anticancer activity. We hypothesize that phages could act in concert with the immune system in immunosurveillance against bacteria, viruses and cancer.

Phage as a modulator of immune responses: practical implications for phage therapy.

Although the natural hosts for bacteriophages are bacteria, a growing body of data shows that phages can also interact with some populations of mammalian cells, especially with cells of the immune system. In general, these interactions include two main aspects. The first is the phage immunogenicity, that is, the capacity of phages to induce specific immune responses, in particular the generation of specific antibodies against phage antigens. The other aspect includes the immunomodulatory activity of phages, that is, the nonspecific effects of phages on different functions of major populations of immune cells involved in both innate and adaptive immune responses. These functions include, among others, phagocytosis and the respiratory burst of phagocytic cells, the production of cytokines, and the generation of antibodies against nonphage antigens. The aim of this chapter is to discuss the interactions between phages and cells of the immune system, along with their implications for phage therapy. These topics are presented based on the results of experimental studies and unique data on immunomodulatory effects found in patients with bacterial infections treated with phage preparations.

Phage as a Modulator of Immune Responses

Although the natural hosts for bacteriophages are bacteria, a growing body of data shows that phages can also interact with some populations of mammalian cells, especially with cells of the immune system. In general, these interactions include two main aspects. The first is the phage immunogenicity, that is, the capacity of phages to induce specific immune responses, in particular the generation of specific antibodies against phage antigens. The other aspect includes the immunomodulatory activity of phages, that is, the nonspecific effects of phages on different functions of major populations of immune cells involved in both innate and adaptive immune responses. These functions include, among others, phagocytosis and the respiratory burst of phagocytic cells, the production of cytokines, and the generation of antibodies against nonphage antigens. The aim of this chapter is to discuss the interactions between phages and cells of the immune system, along with their implications for phage therapy. These topics are presented based on the results of experimental studies and unique data on immunomodulatory effects found in patients with bacterial infections treated with phage preparations.