Phage Therapy

1. Phage Therapy
Presented by: Usman Shareef
CMS #: 14539
Presented to: Dr. Umair Ilyas

2. Contents
• Phages-An Introduction
• Phage structure
• Fates of phages
• Antibiotic resistance-growing problem
• Phage therapy
• History
• Why phage therapy??
• Initial problems
• Solutions
• Prerequisites for phage therapy
• Administration and examples
• Future implications
• Advantages and disadvantages
• Challenges
• Conclusions

3. Bacteriophage (Phage)
• Definition - Obligate intracellular parasites that multiply inside bacteria by making use
of some or all of the host biosynthetic machinery.
• Significance
– Phage therapy
– Gene transfer in bacteria
– Phage display
– Medical applications
• Identification of bacteria - phage typing
• Treatment and prophylaxsis???

4. Structure of bacteriophage
• A bacteriophage particle consists of a single stranded DNA or RNA molecule,
encapsulated inside a protein coat or lipoprotein coat.
• Tail morphologies: long , flexible tails, double layered, contractile tails.
• The size of phage head is correlated to the size of genome being packaged and varies in
diameter between 45 and 100nm.

6. Fates of bacteriophages

7. Time course of lytic infection cycle

8. Antibiotic resistance-growing
problem
• Main reason is the abusive use of antibiotics over past twenty years.
• The resistance phenomenon represents not only an important healthcare issue but also an
economic problem.
• Penicillin fails to completely eradicate Streptococci in 35% of the pateints.
• Infections caused by Streptomyces agalactiae in pregnant women cannot be treated with
antibiotics because of the risk of abortion.

9. Phage therapy
• Phage therapy is the therapeutic use of bacteriophages for the treatment of pathogenic
bacterial infections.
• This method of therapy is still being tested for treatment of a variety of bacterial
infections.
• Has not yet been approved in countries other than Georgia.
• Phage therapy has many potential applications in human medicines as well as dentistry ,
veterinary science and agriculture.
• Bacteriophages are much more specific (host specific) and have a high therapeutic
index.

10. History
 Edward Twort (1915) and Felix d'Herelle (1917) independently reported isolating
filterable entities capable of destroying bacterial cultures and of producing small cleared
areas on bacterial lawns.
 It was F d'Herelle, a Canadian working at the Pasteur Institute in Paris, who gave them
the name "bacteriophages"-- using the suffix phage (1922).
Frederick Twort
1915 Félix d'Hérelle 1917

12. WHY PHAGE THERAPY???

13. Bacteriophages Antibiotics Comments
Very specific (i.e., usually affect only the targeted
bacterial species); therefore, chances of
developing secondary infections are avoided
Antibiotics target both pathogenic microorganisms
and normal microflora. This affects the microbial
balance in the patient, which may lead to serious
secondary infections.
High specificity may be considered to be a
disadvantage of phages because the disease-
causing bacterium must be identified before phage
therapy can be successfully initiated. Antibiotics
have a higher probability of being effective than
phages when the identity of the etiologic agent has
not been determined.
Replicate at the site of infection and are thus
available where they are most needed
They are metabolized and eliminated from the
body and do not necessarily concentrate at the site
of infection.
The "exponential growth" of phages at the site of
infection may require less frequent phage
administration in order to achieve the optimal
therapeutic effect.
No serious side effects have been described. Multiple side effects, including intestinal
disorders, allergies, and secondary infections (e.g.,
yeast infections) have been reported
A few minor side effects reported for therapeutic
phages may have been due to the liberation of
endotoxins from bacteria lysed in vivo by the
phages. Such effects also may be observed when
antibiotics are used
Phage-resistant bacteria remain susceptible to
other phages having a similar target range.
Resistance to antibiotics is not limited to targeted
bacteria.
Because of their more broad-spectrum activity,
antibiotics select for many resistant bacterial
species, not just for resistant mutants of the
targeted bacteria
Selecting new phages (e.g., against phage-resistant
bacteria) is a relatively rapid process that can
frequently be accomplished in days or weeks.
Developing a new antibiotic (e.g., against
antibiotic-resistant bacteria) is a time-consuming
process and may take several years
Evolutionary arguments support the idea that
active phages can be selected against every
antibiotic-resistant or phage-resistant bacterium
by the ever-ongoing process of natural selection.

14. Initial problems
• Problem # 1: Specific host range
• Problem # 2: Bacterial debris present in phage preparations.
• Problem # 3: Attempts to remove host bacteria from therapeutic preparations
• Problem # 4: Rapid clearance of phages
• Problem # 5: Lysogeny
• Problem # 6: Lack of knowledge

15. WHAT CAN BE THE
SOLUTIONS????

16. Solutions
• Use of phage mixtures (cocktails)
• Application in chronic infections: time to select appropriate phages
• Broad spectrum phages (e.g. all S. aureus) exist.
• Add phages to antibiotics
• Study of genome of phage

17. Prerequisites for phage therapy
Various prerequisites that should be met:
1. Phage therapy should not be attempted before the biology of the therapeutic phage is
well understood.
2. Phage preparations should meet all safety requirements.
3. Phage preparations should contain infective phage particles.
4. The phage receptor should be known.
5. The efficacy of phage should be tested in an animal model.

18. Culture-commercial preparations
• D’Herelle’s commercial laboratory in Paris produced at least 5 different phage
preparation against various bacterial infection.
• The preparations were called as:
• Bacte-coli-phage
• Bacte-rhino-phage
• Bacte-intesti-phage
• Bacte-pyo-phage
• Bacte-staphy-phage.
• Therapeutic phages were also produced in United States.
• In the 1940’s Eli Lilly Company produced seven phage products for human use.

19. Administration
• Orally
• Topically on infected wounds
• Application in liquid form is possible and stored preferably in refrigerated vials
• Injections are rarely used

20. Examples
• Killing of Mycobacterium avium and Mycobacterium tuberculosis by a
Mycobacteriophage delivered by a non virulent Mycobacterium
• Tuberculosis is a serious health problem that results in millions of deaths around the
world each year
• Mycobacterium smegmatis , an avirulent mycobacterium is used to deliver the lytic
phage TM4 where both M.avium and M.tuberculosis reside within macrophages
• These results showed that treatment of M.avium infected as well as M.tuberculosis
infected with M.smegmatis infected with TM4 resulted in significant reduction in
number of viable intracellular bacillli.

21. Example # 2
• Killing of S.aureus by using bacteriophage that kills S.aureus .
• By treating the infection with the use of phage impregnated pad
• Successful results were reported

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

23. Future implications
• As a vaccine delivery vehicle
• Prophylaxsis???
• Phage display
• Phage typing
• Genetically manipulated lysogenic phages for in situ gene delivery:
--> in situ delivery to bacterial cells of
* killing genes (doc)
* antisense RNA to block translation
Westwater et al. 2003. Use of a genetically engineered phage to deliver antimicrobial agents
to bacteria: an alternative therapy for treatment of bacterial infections.
Phages as bio-control and bacteriophage bioprocessing

24. As vaccine delivery vehicle
• The stability of whole bacteriophage lambda particles, used as a DNA vaccine delivery
system has been examined.
• When phage lambda was diluted into water, a marginal loss in titre was observed over a
2-week period.
• Over a 24 h period, liquid phage suspensions were stable within the pH range pH 3-11,
therefore oral administration of bacteriophage DNA vaccines via drinking water may be
possible.

25. Phage typing
• Phage typing is also known as the use of sensitivity patterns to specific phages for
precisely identifying microbial strains.
• The sensitivity of detection would be increased if the phages bound to bacteria are
detected by specific antibodies
• The technique has most extensively been used for the detection of Mycobacterium
tuberculosis , E.coli , Pseudomonas , Salmonella , Listeria and Campylobacter species

26. Advantages and disadvantages
Advantages
• Phages are very specific and do not
harm the useful bacteria that live in and
in the body
• They replicate at the site of infection
• They are active against antibiotic
resistant bacteria
• Once administered it will not need more
dosages
Disadvantages
• The great specificity is also a
disadvantage when the exact species of
bacteria is unknown and also in case of
multiple infections
• Infections whose agents are hidden in
the interior of the cells may be
inaccessible to phages

27. Cont’d
Advantages
• High expectations of safety
• Can carry accessory genes for additional
therapeutic benefits
Disadvantages
• Resistance can arise (may use
cocktails)
• Immune response of host may limit
or prevent the re-use
• 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.

28. Challenges
• Specificity of phages
• Novelty
• Efficacy and other technical challenges
• Regulatory approvals
• Market acceptance
• Patient safety

29. Conclusions
• Multidrug resistance bacteria have opened a second window for phage therapy
• Modern innovations combined with careful scientific methodology can enhance
mankind’s ability to make it work this time around
• Phage therapy can then serve as a stand alone therapy for infections that are fully
resistant
• It will then be serve as a co-therapeutic agent for infections that are still susceptible to
antibiotics by helping to prevent the emergence of bacterial mutants against either agent

32. References
• Hugos and Russel’s Microbiology
• Carlton R M (1999) Phage therapy: Past history and Future prospects
• Broxmeyer et al. 2002. J. Infect. Dis. 186:1155-1160.
• (Dabrowska et al. 2005. Bacteriophage penetration in vertebrates. J. Appl.
Microbiol. 98: 7-13.)
• Antimicrob. Agents Chemother. 47: 1301-1307.
• Nature Reviews/Drug Discovery 2: 489-497.
• www.phagetherapycenter.com/ - Phage Therapy Center of Tbilsi, Georgia.
“…effective treatment solution for patients who have bacterial infections that do
not respond to conventional antibiotics”
• http://www.researchgate.net/publication/8514842_Bacteriophage_lambda_is_a_h
ighly_stable_DNA_vaccine_delivery_vehicle