Phage therapy

Institute of Molecular Biomedicine

Comenius University, Faculty of Medicine

www.imbm.sk

tothova.lubomira@gmail.com

Outline

• Introduction

• Taxonomy

• Cycles of phages

• Bacterial resistance

• Phage therapy

• Conditions for phage therapy

• Enzybiotics

• Phages in other industries

• Phage typing

• Phage display

• Bacterial sensing

• Future outlook

Microbiome

• Total microorganisms, their genomesand ecosystem in/on the human body

• Together „supraorganism“

• New genes with advantages

• Mouth, skin, gut...

• Unique ecosystems

Human cellsBacterial

cells

We are bacteria

10 13 10 14

• „core microbiome“

• „variable microbiome“

• Personalized medicine

• Host´s physiology

• Health status

HUMAN MICROBIOME

PROJECT 2007

Mammalian virome

• Viruses of eukaryotic cells (e.virome)

• Bacteriophages infecting bacterial cells (b.virome)

• Viruses of archea (a.virome)

• Virus-derived genetic elements in host chromosomes

1031 members of virome1% explored

Bacteriophages

• Most abundant entities in biosphere

• Viruses specific for bacterial cells

• Distribution according to their hosts

Where can we find phages ?

• Everyday life

• Soil

• Sewage

• Food

• Water

....

• Everywhere where bacteria are present

Taxonomy• ICTV (not CVTI ) International Committee for Taxonomy of Viruses

• Up-to-date taxanomic guidelines

Tailed bacteriophages order CAUDOVIRALES

Three families Myoviridae

Siphoviridae

Podoviridae

Different genera

13 families and 31 genera

• Diverse genome size

• ss DNA, ds DNA or ss RNA, ds RNA

• Phage particles (VIRIONS) lipoprotein

or protein coat

• Tails

• ~ 200 nm

Life cycles of phages

• Lytic

• Lysogenic

• Pseudolysogenic

Phage therapy• Therapeutical usage of bacteriophages for treating bacterial infections

• Long history (1915/1917)

• Some approvals

• Georgia, Russia and Poland

Frederick Twort Félix d´Herelle

• Medicine, dentistry, veterinary science and agriculture

Mechanisms of antibiotics

• Cell wall synthesis

• Protein synthesis

• Nucleic acid synthesis

• Cell membrane functions

• Metabolism

• Others

Resistance of bacteria

• Antibiotics vs. multidrug resistant bacterial strains

• Biofilm

Resistance of bacteria

• Absence of a susceptible target

• Multidrug resistance effluxpumps

• Mutations

• Transformation

• Modification of target site

• Direct antibiotic inactivation

Lack of new antibiotics or effective treatment !!!

Phage therapy renaissance

Pros and cons of bacteriophage therapy

• Bacteriophages infect only bacterial cells

• Bacteriophages are capable

of self-replication and self-elimination

• Bacteriophages with narrow host range

• Phage therapy trials without serious side effects or allergic reactions

• Single dose would be sufficient to cure the patient

• Bacteriophages in combination with antibiotic therapy may lower the occurrence of bacterial resistance

• Bacteriophages are highly species-specific, it is necessary to precisely characterize the infectious bacterial agent prior to their application

• Phage defined in vitro environment asstrictly lytic, could change this cycleunder the physiological conditions of the human body

• Possibility of toxic shock after the bacterial lysis and subsequent amplification of neutralizing antibodies during the re-treatment of the patient

Prerequisites for phage therapy

• Host range

• Purity of phage preparations

• Clearance of phage therapeuthics

• Lysogeny in vivo

• Complete phage characterization (genes, receptors ... host-phageinteractions ..., )

• Safety testing !!!

Commercial products

PhagobiodermBiophage-PA

Routes of administration

OralTopicalIntravenous

Phages in agriculture

• Intralytix, Inc. (http://www.intralytix.com)

• EBI Food Safety (http://www.ebifoodsafety.com)

• Omnilytics, Inc. (http://www.omnilytics.com)

• Sevapharma, a.s.(http://www.sevapharma.cz)

Enzybiotics

• Hybrid name lytic enzymes + antimicrobial potential

• Enzymes able to cause microbial cell death

Endolysins (from phages)

Bacteriocins (from bacteria)

Autolysins (from bacteria)

Lysozymes (various organisms)

How endolysins work?

• Enzymatic cleavage of bacterial cell wall peptidoglycans (cell lysis)

• ds DNA phages

• end of the lytic cycle

(1) N-acetylmuramoyl-L-alanine amidases;

(2) endopeptidases;

(3) N-acetyl-β-D-glucosaminidase;

(4) N-acetyl-β-D-muramidases (lysozymes)

(5) lytic transglycosylases

Methods: Vectors in MB

Methods: Phage typing

• Used for identification and differentiating of bacterial pathogens

• Lysis due to viral infection is specific

• Bacteriopage-based diagnostics

Methods : Bacteria sensing

Biosensors advantages

• Sensitive, rapid and selective detection

• Cost-effective and portable

• Lower limit of detection

• Minimum sample preparation

Methods : Phage display

More information by J.B.

Future outlook

• Large range appplication of phages in molecular biology,

biotechnology and medicine ! ! ! ! !

• Safety and efficacy issues

• Unexplored entities

Thanks for Your attention !