Phagos odonto

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Phagos odonto

  1. 1. La Thérapie par les Bactériophages Approche par la Médecine Dentaire BIF7000 – UQAM 2013 •Patricia Morales •Natalia Samus •Igor Lemos •Deraldo Duran (Lisbon University)
  2. 2. BACKGROUND Problématique Medécine Dentaire
  3. 3. BACKGROUND Problématique Streptococcus mutans • Most important bacteria associated with tooth decay, • Naturally present in the human oral microbiota
  4. 4. BACKGROUND Problématique Healthy teeth
  5. 5. BACKGROUND Problématique Dental caries Periodontal diseases and dental caries are the most prevalent chronic diseases, affecting children, adolescents, adults and the elderly. In fact, dental caries affects 60% to 90% of schoolchildren and the vast majority of adults in most industrialized countries. http://www.phac-aspc.gc.ca/publicat/cdicmcbc/30-4/preface-eng.php#ref_05
  6. 6. BACKGROUND Problématique Dental caries Endodontic therapy • In certain cases, endodontic therapy may be necessary for the restoration of a tooth
  7. 7. BACKGROUND Problématique Dental caries Endodontic therapy Prosthetic teeth Dental surgery
  8. 8. BACKGROUND Problématique Streptococcus mutans 80% - 90%
  9. 9. BACKGROUND Problématique Efficacy of dental floss in addition to a toothbrush ?????????????? X In fact, dental caries affects 60% to 90% of schoolchildren and the vast majority of adults in most industrialized countries.
  10. 10. BACKGROUND Phage therapy Synthetic Biology Bioinformatics + Genetic engineering + Mol Biology
  11. 11. Phage Therapy Phage therapy is the therapeutic use of bacteriophages to treat pathogenic bacterial infections 1:n Lysin Streptococcus mutans
  12. 12. Phage Therapy 1 - Bacteriophage X 1:n n - Bacteries Chain Reaction
  13. 13. Phage Therapy Solution 1 - Bacteriophage 1:1 1 - Bacteria No Chain Reaction
  14. 14. SOLUTION Synthetic Bacteriophage Engineered dna PROMOTER+LYSIN 1:1 Lysin Streptococcus mutans
  15. 15. SOLUTION Synthetic Bacteriophage Toothpaste
  16. 16. Lysin destroys a cell wall of grampositive bacteria (Streptococcus) Lysin Hydrolysis of (1->4)-beta-linkages between N-acetylmuramic acid and N-acetyl-D-glucosamine residues in a peptidoglycan Peptidoglycan
  17. 17. PHAST(PHAge Search Tool) - web server designed to identify, annotate and graphically display prophage sequences within bacterial genomes or plasmids 10947 Genomes Databases for download: vIrus DB (57.902 Mb). Last update: Nov 1 2013 Prophage and virus DB (58.039 Mb). Last update: Nov 1 2013 Bacteria DB (2.626 Gb). Last update: Nov 1 2013 PHAST result DB (tar zip format) (104.167 Mb). Last update: Nov 1 2013
  18. 18. Pathologic Streptococci (compete genom) NC number strain PHAGE # Str. mutans NC_004350.1 UA159 1 Str. mutans AP010655.1 NN2025 0 Str. mutans CP003686.1 GS-5 0 Str. criceti AEUV00000000 HS-6 - Str. ratti - - - Str. sobrinus - - - Str. downei AEKN00000000.1 F0415 1 Str. ferus - - - Str. macacae AEUW00000000 NCTC 11558 - Allan L. Delisle, Ming Guo, Natalia I. Chalmers, et al Appl Environ Microbiol. 2012 Apr;78(7):2264-71
  19. 19. Streptococcus mutans UA159 complete genome Genes de phages
  20. 20. Streptococcus mutans UA159 genome, phage genes
  21. 21. Streptococcus mutans UA159, genes of phages # CDS_POSITION BLAST_HIT E-VALUE SEQUENCE 1 complement(1777567..17 78640) PHAGE_Entero_phiV10: O-acetyltransferase; SMU_1884c; phage(gi89152472) 4e-16 Click 2 complement(1781127..17 81393) PHAGE_Human__2: EBNA-1; SMU_1889c; phage(gi139424506) 5e-06 Click 3 complement(1781999..17 82835) PROPHAGE_Escher_MG1655: IS150 transposase B; SMU_1893c; phage(gi16131429) 8e-74 Click 4 complement(1782829..17 83347) PROPHAGE_Ralsto_GMI1000: ISRSO11-transposase ORFA protein; SMU_1894c; phage(gi17546156) 2e-07 Click 5 complement(1783662..17 83964) PHAGE_Staphy_PT1028: ORF011; SMU_1896c; phage(gi66395175) 4e-07 Click 6 1784304..1784786 PHAGE_Bacill_SPBc2: ABC transporter; SMU_1897; phage(gi9630145) 3e-07 Click 7 1784881..1786392 PHAGE_Bacill_SPBc2: ABC transporter; SMU_1898; phage(gi9630145) 5e-62 Click 8 1786502..1787515 PHAGE_Megavi_lba: hypothetical protein; SMU_1900; phage(gi448825457) 1e-12 Click 9 complement(1788212..17 89234) PHAGE_Equid__9: envelope glycoprotein J; SMU_1904c; phage(gi216905924) 5e-05 Click
  22. 22. Streptococcus downei F0415 genome, phage genes
  23. 23. Endolysin 70%
  24. 24. Putative endolysin, phage M-102 Definition: Streptococcus phage M102 Accession: NC_012884 Length: 31147 bp LOCUS_TAG M102_gp20 ORF START 20879 ORF STOP 21394 STRAND Forward REGION 1 REGION intact or incomplete 39.25 PROTEIN SEQUENCE MLKKTLAILGLSASLLFVSAHANAHTSRLTLDQTNELYTRLAAEGRGVDT DQVYGMQCVD IDTDLTNNYVGIPISGNAIDLLDSARRAGYEIVPANKPPRAGDLFVMDTN ALYGHPFGHT GYVYKVNPDGSFETVEQNVGDDSNFYTGTVAKFMHRTRDYMLGYIRLA YRK HOMOLOG/ORTHOLOG SPECIES PHAGE_Strept_M102 HOMOLOG/ORTHOLOG PROTEIN putative endolysin; phage(gi242345591) HOMOLOG/ORTHOLOG EVALUE 1e-96
  25. 25. Result BLAST endolysin phage M-102 •lysin [Streptococcus mutans] •NCBI Reference Sequence: WP_002287207.1 •LOCUS WP_002287207 171 aa linear BCT 12-MAY-2013 •DEFINITION lysin [Streptococcus mutans]. •ACCESSION WP_002287207 VERSION WP_002287207.1 GI:488215999 •SOURCE Streptococcus mutans •ORGANISM Streptococcus mutans Bacteria; Firmicutes; Bacilli; Lactobacillales; Streptococcaceae; Streptococcus. •COMMENT REFSEQ: This record represents a single, non-redundant, protein sequence which may be annotated on many different RefSeq genomes from the same, or different, species. •FEATURES Location/Qualifiers source 1..171 /organism="Streptococcus mutans" /db_xref="taxon:1309" Protein 1..171 /product="lysin" /calculated_mol_wt=18727 Region 54..165 /region_name="CHAP" /note="CHAP domain; pfam05257" /db_xref="CDD:218523" •ORIGIN 1 •mlkktltflg lsvgllivsp hanahtsrlt ldqtnelytr laaegrgvdt dqqygmqcvd 61 idtdltnnyv gvpisgnaid lldsaraagy eivpaskppr agdlfvmdts tvyghpfght 121 gyiyrvnpdg sfetveqnvg ddsnlytgtv akfmhrtrdy mlgyirlayr k
  26. 26. Phage PH-10 complete genome
  27. 27. Lysin A, phage PH-10 Definition: Streptococcus phage PH10 Accession: NC_012756 Length: 31276 bp LOCUS_TAG PH10_gp51 ORF START 29525 ORF STOP 30529 STRAND Forward REGION 1 REGION intact or incomplete 39.53 PROTEIN SEQUENCE MKKNDLFIDVSSHNGYDITGILEQMGTTNTIIKISESTSYMNPCLSAQVEQSNPIGFYHFAWFG GDIEEAEREARYFLDNVPQKVKYLCLDYEDHASGDKQANTDACIRFMEILKENGYEPIYYSYKP FTLNNIYYEQILEKFPNSLWIAGYGLNDGTADFEYFPSMDGIRWWQYSSNPYDKNIVLLDDEE AKPKWKRNDTGWWYEYPDGSYPKEEWEKIDGTWYYFNERGYSIASRWLKDDGKWYYLKE NGAMAVGWVLVNGKWYYLDASGAMVTGWVQYKDKLYHFKEENGEMSSKEL VKVEGGWYYVNEDGSRSDKPALDVLPDGLIVTTK HOMOLOG/ORTHOLOG SPECIES PHAGE_Strept_PH10 HOMOLOG/ORTHOLOG PROTEIN lysA; phage (gi238821368)
  28. 28. Str. downei Str. mutants Phages M-102, PH-10 Phage M-102 Protein LYSIN
  29. 29. Résultats •En utilisant la base de données de PHAST nous avons trouvé rapidement deux Streptococcus pathogènes (Str. mutants et Str. downei) les quels contiennent gènes de phages dans ses génomes. •On a trouvé parmi ces deux bacteriophages specifiques de streptococcus mutants (M-102 and PH-10), le gène qui contiennent la protéine lysin, la plus active des protéines antimicrobiennes. •Ainsi les protéines de ces phages pourraient être concernés comme des agents potentiels contre les streptocoques pathogènes pour d'autres études.

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