Phage Therapy.pptx

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.

5. Fates of bacteriophages

6. Time course of lytic infection cycle

7. 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.

8. 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.

9. 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

10. Why phage therapy???

11. 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.

12. 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

13. What can be the solutions????

14. 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

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

16. 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.

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

18. 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 .

19. 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

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

21. 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

22. 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.

23. 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

24. 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

25. 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.

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

27. 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

28. 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_highly_stable_DNA_vaccine_delivery_vehicle