Bacteriophages are the most abundant entities on earth. These bacterial viruses have genetic material in the form of either DNA or RNA, encapsidated by a protein coat.The capsid is attached to a tail which has fibers, used for attachments to receptors on bacterial cell surface.

1. Bacteriophages

2. ANTIMICROBIAL DRUG DISCOVERY
THROUGH BACTERIOPHAGE GENOMICS

3. Bacteriophages have been viewed not only as important
genetic but also as potential antibacterial therapeutic
Over evolutionary time bacteriophages have developed
unique proteins that arrest critical cellular processes to
commit bacterial host metabolism to phage reproduction.
Introduction

4. One can exploit this concept of phage- mediated
bacterial growth inhibition to antibiotic discovery
So far many phages have been sequenced and
identified several novel polypeptide families that
inhibited growth upon expression in bacteria.

5. There is an urgent need to develop new classes of
antibiotics to tackle the increase in resistance in
many common bacterial pathogens.
Pathogens such as Staphylococcus aureus,
Streptococcus pneumoniae and Enterococcus
faecalis, which are each capable of causing severe
and even fatal infections , have become
increasingly resistant to multiple antibiotics.

6. The cellular targets for some of these polypeptides were
identified and several were shown to be essential
components of the host DNA replication and transcription
machineries
Mimicking the growth–inhibitory effect of phage
polypeptides by a chemical compound , coupled
with the plethora of phages on earth, will yield new
antibiotic to combat infectious diseases.

7. Phages are recently resurfaced as the saviors of
humankind in the best selling novel-Prey by
Michael Crichton(2002) –in which phages are
used to destroy laboratory–escaped “bacterial
nanoparticles” threatening life on earth.This
reflects the potential of bacteriophages to be
used as a powerful tool in dealing with infectious
diseases of bacterial etiology.

8. Bacteriophage

9. SEM of Phage

10. Structure of Bacteriophage
•Phage head: composed of coat
protein and genome in the core
•Genome: DNA codes for enzymes
and proteins necessary to replicate
more viruses
•Tail Sheath: DNA travels from
head to bacteria through sheath
•Tail fiber: helps anchor the phage
on the cell membrane

11. Phage life cycle: Lytic vs Lysogenic
Phage replicates by lytic life cycle
Non-integration of phage genetic material
Phage lyse host bacterium
lytic or virulent phage
Phage replicates by lysogenic life cycle
Integration of phage genetic material
temperate phages (prophages) generally larger than lytic
phages (carry ~40kb genetic material)

12. Adsorption by Lytic Bacteriophage
The bacteriophage binds to specific
receptors on the bacterial cell wall.
Tail conformation changes/contracts
central core penetrates cell wall

13. Penetration
• The bacteriophage injects its genome into
the bacterium's cytoplasm

14. Early Replication
•
-Phage-coded enzymes shut down host’s DNA,RNA,protein
synthesis
-Early function inovolve the takeover of the host cell and the
synthesis of early viral mRNA
-Late functions include the subsequent synthesis of other proteins
and assembly of the nucleocapsid.
-Replication phage DNA protected from host restriction
endonucleaes

15. Phage Release
A bacteriophage-coded enzyme break down the
peptidoglycan in the bacterial cell wall
causing osmotic lysis.

17. Conventional Bacteriophage Therapy in
humans
Biomedical technology today is very different
from what it was in the early days of phage
therapy research
In early days bacteriophage therapy was used by
making bacteriophage preparation and are
effective against P. aeruginosa, E.coli, S.aureus,
Streptococcus and proteus

18. The first reviwed report of the therapeutic efficacy
of PhagoBioDerm (Cock tail of lytic bacteriophages)
was recently published (Markoishvili et al., 2002)
107 patients with ulcers – failed to response to
conventional therapy
With PhagoBioDerm - Ulcers healed completely in
67(70%)

19. S.aureus infection
Treated with phage
impregnated pad Improvement in wound healing

20. “Pio bacteriophagum fluidum”- one of the polyvalent phage
preparartions produced by the EIBMV.The preparation targets
a variety of bacterial pathogens, including P.aeruginosa, E.
coli, S.aureus, Streptococcus and proteus

21. Limitations of phage therapy
1.Emergence of bacterial strains resistant to
particular phages. The emergence of phage –
resistant bacterial mutants was observed and
the phenomenon was suggested to be a
potential problem of phage therapy
(Summers, 1999; d’Herelle,1930)

22. Limitations of phage therapy
2.The development of phage–neutralizing
antibodies-The production of neutralizing
antibodies should not be a significant obstacle
during initial or relatively short-term
therapeutic treatments at least.

23. Combating the limitations
Modernization of phage therapy
1. Sequencing of whole genome
2.Rapid and high –throughput, sequence –based
Screening methodologies(e.g., microarrays)

24. Contd……
High–throughput bacteriophage genomics
strategy is the improvised form of conventional
phage therapy.
Exploitation of the Concept of phage –mediated
inhibition of bacterial growth to systematically
identify antimicrobial phage –encoded
polypeptides.

25. To tackle the increase in resistance in many common
bacterial pathogens.
Methicillin resistance s.aureus
Vancomycin resistant enterococci.
Genomic is providing a new strategy by revealing new
molecular targets and peptides that are giving rise to
novel antimicrobialdrug.

26. Key steps in the genomics driven antibiotic drug
discovery process

27. Key criteria to be considered in target
selection
• The target should be present in a required
spectrum of organism.
• It should be absent in humans.
• It should be essential for bacterial growth.
• It should be expressed and relevant to be
infection process.
• Some thing about the function of target
should be known.

28. Peptides and their targets
• Product of bacteriophage T7 gene2(gp2) binds E.coli RNA
polymerase.
• The AsiA protein of phage T4 the bacterial RNA polymerase
σ70 transcription factors.
• Protein P of phage λ and B of phage P2 each bind to and
redirect the host DnaB helicase to there respective phage
origin of replication.

29. Structure of the T4 bacteriophage

30. • Bacteriophages are even more elaborate than the poxviruses.
The T2, T4, and T6 phages that infect E. coli have been
intensely studied.
• Their head resembles an icosahedrons elongated by one or
two rows of hexamers in the middle and contains the DNA
genome.
• The tail is composed of a collar joining it to the head, a central
hollow tube, a sheath surrounding the tube, and a complex
baseplate.

31. TMV
• The tobacco mosaic virus (TMV) capsid contains a
single type of small subunit possessing 158 amino
acids.
• Only about 474 nucleotides out of 6,000 in the virus
RNA are required to code for coat protein amino
acids.

33. A virus consists of a nucleic acid
surrounded by a protein coat
• Viruses were detected indirectly long before they were
actually seen

34. The Discovery of Viruses: Scientific Inquiry
• Tobacco mosaic disease stunts growth of tobacco
plants and gives their leaves a mosaic coloration
• In the late 1800s, researchers hypothesized that a
particle smaller than bacteria caused the disease
• In 1935, Wendell Stanley confirmed this hypothesis by
crystallizing the infectious particle, now known as
tobacco mosaic virus (TMV)

35. Extracted sap
from tobacco
plant with
tobacco mosaic
disease
Passed sap
through a
porcelain filter
known to trap
bacteria
Healthy plants
became infected
Rubbed filtered
sap on healthy
tobacco plants
1 2 3
4

36. Tobacco mosaic virus
Tobacco mosaic virus (TMV) of plants provides a good model to
learn about the relationship between a virus and its host.
Purpose: to learn about (1) properties of TMV and (2) symptoms
induced by the virus in susceptible and resistant plant hosts
Background information:
Viruses cause a variety of diseases in plants and animals. Virus
diseases that affect humans include measles, mumps, and polio. At
one time, these diseases posed major health concerns, but with
the widespread use of vaccinations in children, these diseases
occur rarely.

37. 1. View healthy and infected tobacco and tomato plats
1. Observe crushing of healthy and mosaic diseased tomato leaves in buffer using
mortar and pestle to produce virus infected plant sap.
2. Mark tobacco leaves with your initials using Sharpie.
3. Use sponge to rub TMV sap from crushed leaves onto your marked healthy
tobacco leaf.
Tobacco Mosaic Virus Infection Lesson:
Transmission of Virus and Mosaic Disease

38. Tobacco mosaic virus

39. Tobacco mosaic virus structure

40. Tobacco mosaic virus structure

41. Tobacco mosaic virus structure

42. Tobacco mosaic virus symptoms

43. Tobacco mosaic virus transmission

44. Application of phages
 Model system of
molecular biology
 Cloning and
expression
 Phage display system
 Phage typing
 Phage therapy:
 phage as natural, self-
replicating, self-
limiting antibiotics.

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