Ruiz Martinez, Carlota

Universitat Autònoma de Barcelona. Facultat de Biociències

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My enemy’s enemy is my friend: Bacteriophage therapy  Carlota Ruiz Martinez,  Universitat  Autònoma de Barcelona, Grau en Biologia 2012 - 2013 Introduction  Results Conclusions Bacteriophages (phage) are viruses that only infect bacteria and typically attack only a single bacterial strain (Fig.1). This specificity, together with the killing capacity makes them the “natural enemies” of bacteria. [1]  Phage therapy involves using phages or their products as bioagents for the treatment or prophylaxis of bacterial infectious diseases. There are different alternatives:   1. Therapy using living phages: is the “classical” version in which the phage infect the bacteria near the surface and kill them by internal replication and continues multiplying for as long as there are bacteria to infect. The aim of this project is to study in detail bacteriophage therapy and its particularities, breaking this into different types of therapy and analyze the problems to be overcome and the potential solutions that are possible.  Figure 2. Bacteriolysis. (1) Adsorption and DNA injection; (2) DNA replication; (3) production of head and tail; (4) synthesis of lysin; (5) DNA packaging; (6) completion of phage; (7) disruption of the cell wall and release the progeny; (8) circularization of DNA; (9) integration of the phage DNA into the host genome.  From [1] 3. Utilization of phage lysin: most tailed phages produce peptidoglycan hydrolase (lysin) to release their  progeny at the final stage of multiplication.       Lysin (Fig.4) destroys the peptidoglycan directly exerting a bacteriolytic effect. It can also destroy the cell walls of non-growing  bacteria.  [1] Figure 3. Schematic representations of the genetic organization of the modified phage Pf3R.    From [2] Figure 4. Attack points of phage encoded lysins on the peptidoglycan of gram-positive bacteria. Amidase, endopeptidase, muramidase or glucosaminidase may be released, depending on the cutting site.   From [1] Figure 5. Overview of potential solutions (a) The complexity of phage cocktails is limited by regulation and manufacturing. (b) Evolution can produce phages with enhanced properties such as decreased clearance. (c) Drug-delivery technologies can enhance systemic phage delivery and reduce phage inactivation and clearance. (d) Enzymatic bacteriophages can be engineered to degrade barriers to phage adsorption (left) and disrupt the structure of bacterial biofilms (right). (e) Non-lytic and/or non-replicative phages can reduce the release of bacterial endotoxins. (f) Bacteriophages can be engineered to target intracellular defense pathways, which protect bacteria against antibiotics. From  [4]   2. Problems to overcome 4. Comparing phage and antibiotics 1. Types of bacteriophage therapy: Figure 7. Antibiotics have direct and indirect effects on the microbiota.   From [6] Figure 6. Color enhanced Transmission Electron Micrograph (TEM) of T4 enterobacteria phages attacking an E. coli cell.   From © Eye of Science/Science Source   References 1. Matsuzaki, S. et  al. (2005) J infect Chemother 11:211-219. 2. Hagens, S. et al. (2004) Antimicrob Agents Chemother 48:3817-3822. 3. Fenton, K. et  al. (2010) Bioengineered Bugs 1:1,9-16. 4. Lu, KT. et  al. (2011) Current Opinion in Microbiology 14:524-531.  5. Kutateladze, M. and Adamia, R. (2012) Trends in Biotechnology 28:12 591-595. 6. P. Willing B. et al. (2011) Nature Rewiev in Microbiology  9: 233-243. 2. Phage therapy using non-replicating genetically modified phages: Hagens et al. constructed a recombinant phage Pf3R, in which the export protein gene of the genome was replaced with a restriction endonuclease gene (Fig. 3). The endonuclease digested the host genomic DNA and consequentially killed the bacteria with minimal release of endotoxin in vitro.  [2] 4. Protein antibiotics: some small phages do not have the genes for holin or lysin proteins. Instead, they produce a protein that inhibits a step in murein monomer synthesis. Their inhibitory gene products are known as “protein antibiotics”.  [1, 3]  Currently, many pathogenic bacteria have acquired multiple drug resistance. In this context, one intriguing approach is to use bacteriophages therapy.  Phages could play an important role in treating bacterial infections in humans, animals and crops, as well as in decontaminating food supplies and communal environments.  However, it is also quite clear that the safe  and controlled use of phage therapy will required detailed information on the properties and behavior of the specific phage-bacterium system. 3. Potential solutions: Figure 1. Phage T4.    From  Science Source   1. Biology of bacteriophages: 2. Life cycle of phages: Phages can be divided into two groups  3. Objective:   Bacteriophages Antibiotics    Environment  Are the most abundant living entities on the planet and natural enemies of bacteria   Normally synthetics or semisynthetic   Host range   Narrower Destroy all bacterial cells disturbing the normal microflora  Side effects  Have not been described  Have severe side effects   Concentration  Decrease as soon as bacterial cells are eliminated   Decreases with time Figure 2 illustrates the general mechanism of baceteriolysis by tailed phages.   Accessibility  Restriction / Modification         Immunogenicity  Resistance  Phages won’t be successful where they cannot make contact with the target bacteria, or do so only in a very limited way (Ex. Intracellular bacteria) In order to protect themselves many types of bacteria have developed a method to chop up any foreign DNA  endonuclease that restrict the infection of phages. In prolonged treatments has been observed a development of antibodies against phages.   In continued incubation of a culture lysed by phage, resistant bacteria can appear        Lytic phage • Repeats a cycle in which self-proliferation is synchronous with destruction of bacteria (lytic cycle) Lysogenic phage  • Has a lysogenic cycle (the phage genome multiplies cooperatively with the host without destroying it) in addition to a lytic cycle 

My enemy's enemy is my friend : bacteriophage therapy

http://ddd.uab.cat/pub/tfg/2013/113524/TFG_carlotaruizmartinez.pdf

My enemy's enemy is my friend : bacteriophage therapy

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