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  • 1.
    Cooper, Callum
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Cardiff University, UK.
    Buyuk, Fatih
    Schelkle, Bettina
    Saglam, Aliye Gulmez
    Celik, Elif
    Celebi, Ozgur
    Sahin, Mitat
    Hawkyard, Tom
    Baillie, Les
    Virulence plasmid stability in environmentally occurring Bacillus anthracis from North East Turkey2017In: Antonie van Leeuwenhoek. International Journal of General and Molecular Microbiology, ISSN 0003-6072, E-ISSN 1572-9699, Vol. 110, no 1, p. 167-170Article in journal (Refereed)
    Abstract [en]

    The Bacillus anthracis virulence plasmid pXO2, which encodes for a polypeptide capsule, can be lost during long term laboratory storage. To determine if pXO2 is lost in nature we screened B. anthracis isolates obtained from B. anthracis spores from contaminated animal burial sites in Turkey for their ability to express a capsule upon primary culture. A total of 672 B. anthracis colonies were examined of which ten produced a mixed mucoid (capsule +ve)/non-mucoid (capsule -ve) phenotype and a further one colony yielded non-mucoid colonies upon repeated culture. Screening by PCR using pXO2 specific primers revealed that seven of these isolates had eliminated the plasmid. Of the four colonies which were positive by PCR, one regained the ability to express a capsule upon repeated culture suggesting that the defect was reversible. This is an important observation as capsule expression is a principal marker of virulence and in the absence of PCR serves as a key diagnostic marker. The results of this preliminary study suggest that pXO2 is lost in nature and that further studies are need to determine the mechanisms by which this occurs.

  • 2.
    Cooper, Callum J.
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Koonjan, Shazeeda
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Nilsson, Anders S.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Enhancing Whole Phage Therapy and Their Derived Antimicrobial Enzymes through Complex Formulation2018In: Pharmaceuticals, ISSN 1424-8247, E-ISSN 1424-8247, Vol. 11, no 2, article id UNSP 34Article, review/survey (Refereed)
    Abstract [en]

    The resurgence of research into phage biology and therapy is, in part, due to the increasing need for novel agents to treat multidrug-resistant infections. Despite a long clinical history in Eastern Europe and initial success within the food industry, commercialized phage products have yet to enter other sectors. This relative lack of success is, in part, due to the inherent biological limitations of whole phages. These include (but are not limited to) reaching target sites at sufficiently high concentrations to establish an infection which produces enough progeny phages to reduce the bacterial population in a clinically meaningful manner and the limited host range of some phages. Conversely, parallels can be drawn between antimicrobial enzymes derived from phages and conventional antibiotics. In the current article the biological limitations of whole phage-based therapeutics and their derived antimicrobial enzymes will be discussed. In addition, the ability of more complex formulations to address these issues, in the context of medical and non-medical applications, will also be included.

  • 3.
    Cooper, Callum J.
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Mirzaei, Mohammadali Khan
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Nilsson, Anders S.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Adapting Drug Approval Pathways for Bacteriophage-Based Therapeutics2016In: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 7, article id 1209Article, review/survey (Refereed)
    Abstract [en]

    The global rise of multi-drug resistant bacteria has resulted in the notion that an antibiotic apocalypse is fast approaching. This has led to a number of well publicized calls for global funding initiatives to develop new antibacterial agents. The long clinical history of phage therapy in Eastern Europe, combined with more recent in vitro and in vivo success, demonstrates the potential for whole phage or phage based antibacterial agents. To date, no whole phage or phage derived products are approved for human therapeutic use in the EU or USA. There are at least three reasons for this: (i) phages possess different biological, physical, and pharmacological properties compared to conventional antibiotics. Phages need to replicate in order to achieve a viable antibacterial effect, resulting in complex pharmacodynamics/pharmacokinetics. (ii) The specificity of individual phages requires multiple phages to treat single species infections, often as part of complex cocktails. (iii) The current approval process for antibacterial agents has evolved with the development of chemically based drugs at its core, and is not suitable for phages. Due to similarities with conventional antibiotics, phage derived products such as endolysins are suitable for approval under current processes as biological therapeutic proteins. These criteria render the approval of phages for clinical use theoretically possible but not economically viable. In this review, pitfalls of the current approval process will be discussed for whole phage and phage derived products, in addition to the utilization of alternative approval pathways including adaptive licensing and Right to try legislation.

  • 4. Henein, Alexandra E.
    et al.
    Hanlon, Geoffrey W.
    Cooper, Callum J.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Denyer, Stephen P.
    Maillard, Jean-Yves
    A Partially Purified Acinetobacter baumannii Phage Preparation Exhibits no Cytotoxicity in 3T3 Mouse Fibroblast Cells2016In: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 7, article id 1198Article in journal (Refereed)
    Abstract [en]

    A surge in the level and scale of antibiotic resistance has prompted renewed interest in the application of bacteriophages to treat bacterial infections. However, concerns still exist over their efficacy and safety. Acinetobacter baurnannii phage BS46, a member of the family Myoviridae, has previously been shown to be effective in murine models. The cytotoxic effect of this phage was evaluated in mouse fibroblast 3T3 cells using four different assays: trypan blue; staining with Hoechst and propidium iodide; lactate dehydrogenase release; and the MTS assay. The addition of phage concentrations up to 2 x 10(9) pfu/mL showed little to no impact on the viability of 3T3 cells after 24 h exposure using the different assays. This study demonstrates that phage BS46 is non-cytotoxic to 3T3 cells using four different assays and that appropriate quality assurance protocols for phage therapeutics are required.

  • 5.
    Khan Mirzaei, Mohammadali
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Ek Blom, Linnea
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Cooper, Callum J.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Weiss, Howard
    Udekwu, Klas I.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Nilsson, Anders S.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Infection Dynamics within a Two Phage One Bacterium System: 1 Implications for TherapyManuscript (preprint) (Other academic)
  • 6.
    Khan Mirzaei, Mohammadali
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Haileselassie, Yeneneh
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Navis, Marit
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Cooper, Callum
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Sverremark-Ekström, Eva
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Nilsson, Anders S.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Immunogenic profiling of structurally distinct bacteriophages and their interaction with human cellsManuscript (preprint) (Other academic)
    Abstract [en]

    Due to a global increase in the range and number of infections caused by multi-resistant bacteria, 11 phage therapy is currently experiencing a resurgence of interest. However, there are a number of 12 well-known concerns over the use of phages to treat bacterial infections. In order to address concerns 13 over safety and the poorly understood pharmacokinetics of phages and their associated cocktails, 14 immunological characterization is required. In the current investigation, the immunogenicity of four 15 distinct phages and their interaction with donor derived peripheral blood mononuclear cells and 16 immortalized cell lines (HT-29 and Caco-2 intestinal epithelial cells) were investigated using 17 standard immunological techniques. When exposed to high phage concentrations (109 PFU/well), 18 cytokine driven inflammatory responses were induced from all cell types. Although phages appeared 19 to inhibit the growth of intestinal epithelial cell lines, they also appear to be non-cytotoxic. Despite 20 co-incubation with different cell types, phages maintained a high killing efficiency, reducing 21 extended-spectrum beta-lactamase-producing Escherichia coli numbers by 1-4 log10 compared to 22 untreated controls. Phages were also able to actively reproduce in the presence of human cells 23 resulting in an approximately 2 log10 increase in phage titer compared to the initial inoculum. 24 Through an increased understanding of the complex pharmacokinetics of phages, it may be possible 25 to address some of the safety concerns surrounding phage preparations prior to creating new 26 therapeutic strategies.

  • 7.
    Khan Mirzaei, Mohammadali
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Haileselassie, Yeneneh
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Navis, Marit
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Cooper, Callum
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Sverremark-Ekström, Eva
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Nilsson, Anders S.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Morphologically Distinct Escherichia coli Bacteriophages Differ in Their Efficacy and Ability to Stimulate Cytokine Release In Vitro2016In: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 7, article id 437Article in journal (Refereed)
    Abstract [en]

    Due to a global increase in the range and number of infections caused by multi resistant bacteria, phage therapy is currently experiencing a resurgence of interest. However, there are a number of well-known concerns over the use of phages to treat bacterial infections. In order to address concerns over safety and the poorly understood pharmacokinetics of phages and their associated cocktails, immunological characterization is required. In the current investigation, the immunogenicity of four distinct phages (taken from the main families that comprise the Caudovirales order) and their interaction with donor derived peripheral blood mononuclear cells and immortalized cell lines (HT-29 and Caco-2 intestinal epithelial cells) were investigated using standard immunological techniques. When exposed to high phage concentrations (10(9) PFU/well), cytokine driven inflammatory responses were induced from all cell types. Although phages appeared to inhibit the growth of intestinal epithelial cell lines, they also appear to be non-cytotoxic. Despite co-incubation with different cell types, phages maintained a high killing efficiency, reducing extended-spectrum betalactamase-producing Escherichia colinumbers by 1-4 log(10) compared to untreated controls. When provided with a suitable bacterial host, phages were also able to actively reproduce in the presence of human cells resulting in an approximately 2 log10 increase in phage titer compared to the initial inoculum. Through an increased understanding of the complex pharmacokinetics of phages, it may be possible to address some of the safety concerns surrounding phage preparations prior to creating new therapeutic strategies.

  • 8. Nakonieczna, A.
    et al.
    Cooper, Callum J.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Gryko, R.
    Bacteriophages and bacteriophage-derived endolysins as potential therapeutics to combat Gram-positive spore forming bacteria2015In: Journal of Applied Microbiology, ISSN 1364-5072, E-ISSN 1365-2672, Vol. 119, no 3, p. 620-631Article, review/survey (Refereed)
    Abstract [en]

    Since their discovery in 1915, bacteriophages have been routinely used within Eastern Europe to treat a variety of bacterial infections. Although initially ignored by the West due to the success of antibiotics, increasing levels and diversity of antibiotic resistance is driving a renaissance for bacteriophage-derived therapy, which is in part due to the highly specific nature of bacteriophages as well as their relative abundance. This review focuses on the bacteriophages and derived lysins of relevant Gram-positive spore formers within the Bacillus cereus group and Clostridium genus that could have applications within the medical, food and environmental sectors.

  • 9. Pimchan, T.
    et al.
    Cooper, Callum J.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Eumkeb, G.
    Nilsson, Anders S.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    In vitro activity of a combination of bacteriophages and antimicrobial plant extracts2018In: Letters in Applied Microbiology, ISSN 0266-8254, E-ISSN 1472-765X, Vol. 66, no 3, p. 182-187Article in journal (Refereed)
    Abstract [en]

    The continuing threat of antimicrobial resistance presents a considerable challenge to researchers to develop novel strategies ensuring that bacterial infections remain treatable. Many plant extracts have been shown to have antibacterial properties and could potentially be combined with other antibacterial agents to create more effective formulations. In this study, the antibacterial activity of three plant extracts and virulent bacteriophages have been assessed as individual components and in combination. When assessed with a modified suspension test, these plant extracts also exhibit antiviral activity at bacterial inhibitory concentrations. Hence, to investigate any potential additive effects between the extracts and virulent phages, the extracts were tested at subantiviral concentrations. Phages alone and in combination with plant extracts significantly reduced (< 0·05) the bacterial concentration compared to untreated and extract treated controls up to 6 h (2–3log10), but this reduction did not extend to 24 h. In most cases, the phage and extract combinations did not significantly reduce bacterial content compared to phages alone. Additionally, there was little impact on the ability of the phages to reproduce within their bacterial hosts. To our knowledge, this study represents the first of its kind, in which antimicrobial plant extracts have been combined with virulent phages and has highlighted the necessity for plant extracts to be functionally characterized prior to the design of combinatorial therapies.

    Significance and Impact of Study

    This preliminary study provides insights into the potential combination of bacteriophages and antimicrobial plant bulk extracts to target bacterial pathogens. It is to our knowledge the first time in which virulent bacteriophages have been combined with antimicrobial plant extracts.

1 - 9 of 9
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