Bacteriophage, Phage Typing and ApplicationBy:- Zebene A.

Out line of the presentation Introduction Composition and Structure of Bacteriophages Infection of Host Cells Phage Multiplication Cycle

A. Lytic or Virulent Phages B. Lysogenic or Temperate Phage

Phage Typing Application of Bacteriophages Reference

Introduction Bacteriophage

Virus that infects bacteria. Obligate intracellular parasites that multiply

inside bacteria by making use of some or all of the host biosynthetic machinery.

Twort and d'Herelle in 1915 and 1917 broth cultures of certain intestinal bacteria could

be dissolved by addition of a bacteria-free filtrate obtained from sewage.

Diversity

There are at least 12 distinct groups of bacteriophages

which are very diverse structurally and genetically;

the best known ones are the common phages of E.coli:T4(lytic phage )&Lambda Phage(Lysogenic phage)

Phage Diversity.docx

Composition and Structure of Bacteriophage

A. Composition They all contain nucleic acid and protein.

Depending upon the phage, the nucleic acid can be either DNA or RNA but not both.

Complexity vary from 3-5 average size gene products code for over 100 gene

The nucleic acids of phages often contain unusual or modified bases. These modified bases protect phage nucleic acid

from nucleases that break down host nucleic acids during phage infection.

B. StructureBacteriophages come in many different sizes and shapes. Size - T4 is among the largest phages; it is

approximately 200 nm long and 80-100 nm wide. Other phages are smaller. Most phages range in size from 24-200 nm in length.

Head or Capsid -All phages contain a head structure which can vary in size and shape. Head acts as the protective covering for the

nucleic acid.

Tail-Many but not all phages have tails attached to the phage head.

Is a hollow tube through which the nucleic acid passes during infection.

size of the tail can vary E.g. Complex phages like T4

The tail is surrounded by a contractile sheath which contracts during infection of the bacterium

At the end the tail have a base plate and one or more tail fibers involved in the binding of the phage to the bacterial cell.

Structure of Enterbacteriacae phage/Bacteriophage T4

Infection of Host CellsA. Adsorption Mediated by the tail fibers or by some analogous

structure on those phages that lack tail fibers and it is reversible.

The tail fibers attach to specific receptors on the bacterial cell and the host specificity of the phage

The nature of the bacterial receptor varies for different bacteria(proteins on the outer surface of the bacterium, LPS, Pili, and lipoprotein)

B. Irreversible attachment

The attachment of the phage to the bacterium via the tail fibers is a weak one and is reversible.

Irreversible binding of phage to a bacterium is mediated by one or more of the components of the base plate.

Phages lacking base plates have other ways of becoming tightly bound to the bacterial cell.

C. Sheath Contraction The irreversible binding of the phage to the

bacterium results in the contraction of the sheath (for those phages which have a sheath) and the hollow tail fiber is pushed through the bacterial envelope.

D. Nucleic Acid Injection The DNA/RNA ejected to the bacterial cell

through the hollow tube The remainder of the phage remains on the

outside of the bacterium

Phage Multiplication Cycle Based on the multiplication cycle /event occur

during replication:-

A. Lytic or Virulent Phages

B. Lysogenic or Temperate Phage

A. Lytic or Virulent Phages

Lytic or virulent phages are phages which can only multiply on bacteria and kill the cell by lysis at the end of the life cycle.

Life cycle 1. Eclipse period  2. Intracellular Accumulation Phase  3. Lysis and Release Phase 

Lytic cycle

B. Lysogenic cycle/Temperate Either multiply via the lytic cycle or enter a

quiescent state in the cell In this quiescent state most of the phage genes are

not transcribed Phage DNA in the repressed state is called a

prophage because it is not a phage but it has the potential to produce phage.

The cell harboring a prophage is termed a lysogen.

Events Leading to Lysogeny:-The Prototype Phage: Lambda

1. Circularization of the phage chromosome

2. Site-specific recombination 

3. Repression of the phage genome A phage coded protein, called a repressor, is made

which binds to a particular site on the phage DNA, called the operator, and shuts off transcription of most phage genes except the repressor gene.

 

What determines whether the virus takes Lytic or Lysogenic cycle?

Determined by the concentration of the repressor and another phage protein called cro in the cell.

If a promoter of repressor gene called C I dominates the phage will progress to lysogenic.

If the an operator gene called cro dominates the phage will progress to lytic cycle.

Events Leading to Termination of Lysogeny Anytime a lysogenic bacterium is exposed to

adverse conditions, the lysogenic state can be terminated. This process is called induction

Adverse conditions lead to the production of proteases (rec A protein) which destroy the repressor protein. This in turn leads to the expression of the phage

genes, reversal of the integration process and lytic multiplication

General cycle In the lysogeny

Used for identification and differentiating of bacterial pathogen.

Specific bacteriophage isolated is mixed with known bacterial pathogen

The lysis due to viral infection of the bacterium is specific and important in identification of the host.

Phage Typing

Plaque assay/Assay for Lytic Phage

Lytic phages are enumerated by a plaque assay

A plaque is a clear area which results from the lysis of bacteria.

Each plaque arises from a single infectious phage.

The infectious particle that gives rise to a plaque is called a pfu (plaque forming unit).

Application of Bacteriophages Model for animal virus transformation 

Lysogeny is a model system for virus transformation of animal cells

Bacteriophage-based Diagnostics and Phage Typing

Phage as vehicles for vaccine delivery Phage display

A molecular technique used in synthesizing polypeptides with novel characteristics.

Cont’d Phage Therapy

Can any phage be used for developing therapeutic phage preparation?

Environmental Applications Bio control for dangerous bacteria

Reference

1. Microbiology byte [Internet].[Last updated 2010, cited 2013] http://www.microbiologybytes.com/virology/Phages.html

2. University of South Carolina School of Medicine: Microbiology and Immunology on line [Internet].[Last updated 2010, cited 2013] Available from http://pathmicro.med.sc.edu/mayer/phage.htm

3. Kenneth Todar.The Microbial WorldLectures in Microbiology by Kenneth Todar PhD    University of Wisconsin-Madison Department of Bacteriology [Internet]. [Last updated 2012, cited 2013] Available from http://textbookofbacteriology.net/themicrobialworld/Phage.html

4. Madigan, Martinko, Stahl, Clark. Brock Biology of Microorganisms 13thEdition: Virus and Virology: Pearson Education .Inc; 2008.

5. Elisabeth Kutter, Alexander Sulakvelidze. Bacteriophage Biology and Application: CRC Press; 2005.

6. Irshad Ul Haq, Waqas Nasir Chaudhry, Maha Nadeem Akhtar, Saadia Andleeb and Ishtiaq Qadri. Bacteriophages and their implications on future biotechnology: a review. Virology Journal 2012; 9(9);1-8

7. Filippov AA, Sergueev KV, Nikolich MP (2013) Bacteriophages against Biothreat Bacteria: Diagnostic, Environmental and Therapeutic Applications. J Bioterr Biodef 2013, S3 : 1-8.