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Computational prediction and characterization of genomic islands:  insights into bacterial pathogenicity Morgan G.I. Langille Department of Molecular Biology & Biochemistry Simon Fraser University http://tinyurl.com/genomic-islands
Genomic Island History ,[object Object],[object Object],[object Object],[object Object],[object Object]
Genomic Island Interest ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]
Genomic Island Interest
Methods for Predicting GIs ,[object Object],[object Object],[object Object],[object Object],[object Object]
Methods of Predicting GIs ,[object Object],[object Object],[object Object]
Previous state of GI identification ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]
Outline ,[object Object],[object Object],[object Object],[object Object],[object Object]
Outline ,[object Object],[object Object],[object Object],[object Object],[object Object]
Mauve-whole genome aligner ,[object Object],[object Object],[object Object],[object Object],(Darling, et al., 2004)
IslandPick: Outline Query Genome A Genome B Genome C Genome D Run Mauve Mauve (A & B) Extract unique regions Mauve (A & C) Mauve (A & D) Genome D Putative  Genomic Islands BLAST Identify overlapping unique regions
Selecting Comparative Genomes Run Mauve Mauve (A & B) Extract unique regions Mauve (A & C) Mauve (A & D) Genome D Putative  Genomic Islands BLAST Identify overlapping unique regions Genome B Genome C Genome D Comparative Genome Selection  (using CVTree distances) Query Genome A
What genomes to use?  ,[object Object],[object Object]
CVTree ,[object Object],[object Object],[object Object],[object Object],[object Object],(Qi, et al., 2004)
Example:  Pseudomonas  Tree ,[object Object],[object Object],0.227 0.256 0.397 0.393 0.411 0.428 0.430 0 0.481 P. fluorescens  Pf-5 P. putida  KT2440 P. fluorescens  PfO-1 P. syringae tomato  DC3000 P. syringae phaseolicola  1448A P. syringae syringae  B728a   P. aeruginosa  PAO1 P. aeruginosa  PA14 Acinetobacter  ADP1
Determining Distance Cutoffs ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]
Example:  Pseudomonas  Tree Maximum Distance Cutoff = 0.42 Minimum Number of Genomes = 3 0.227 0.256 0.397 0.393 0.411 0.428 0.430 0 0.481 P. fluorescens  Pf-5 P. putida  KT2440 P. fluorescens  PfO-1 P. syringae tomato  DC3000 P. syringae phaseolicola  1448A P. syringae syringae  B728a   P. aeruginosa  PAO1 P. aeruginosa  PA14 Acinetobacter  ADP1 Minimum Distance Cutoff = 0.10
Predicting Similar Aged GIs GI Insertion Query Genome 1 genome < distance X Query Genome GI Insertion
Outline ,[object Object],[object Object],[object Object],[object Object],[object Object]
Accuracy of GI methods ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]
Developing a Negative Dataset ,[object Object],[object Object],[object Object]
Negative Dataset  Query Genome 1 genome > distance X GI Insertion Query Genome GI Insertion
IslandPick Cutoffs
[object Object],[object Object],[object Object],173 chromosomes 736 chromosomes (Langille, et al., 2008)
GI Prediction Accuracy Positive Dataset Negative Dataset Predicted Dataset Entire Genome TP FP FN Precision = TP / (TP + FP) Recall = TP / (TP + FN) TN
GI Prediction Accuracy (Langille, et al.,2008) Tool Average number of nucleotides in GIs per genome (kb) Precision Recall Overall Accuracy SIGI-HMM 233 92 33.0 86 IslandPath/ Dimob 171 86 36 86 PAI IDA 163 68 32 84 Centroid 171 61 28 82 IslandPath/ Dinuc 444 55 53 82 Alien Hunter 1265 38 77 71 Literature* 639 100 87 96
Outline ,[object Object],[object Object],[object Object],[object Object],[object Object]
IslandViewer  (Langille, et al., 2009) ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]
IslandPick – Manual genome selection
User Genome Submission
Outline ,[object Object],[object Object],[object Object],[object Object],[object Object]
P seudomonas  aeruginosa Liverpool Epidemic Strain (LES) ,[object Object],[object Object],[object Object],[object Object],[object Object]
LES Analysis ,[object Object],[object Object],[object Object],[object Object],(Winstanley, Langille, et al., 2008)
Signature-tagged mutagenesis (STM) ,[object Object],[object Object],[object Object],[object Object],http://www.traill.uiuc.edu/uploads/porknet/papers/LitchtensteigerPaper.pdf
LES Prophage (Winstanley, Langille, et al., 2008)
LES Genomic Islands (Winstanley, Langille, et al., 2008)
LES in-vivo competitive index ,[object Object],[object Object],[object Object],(Winstanley, Langille, 2008)
Outline ,[object Object],[object Object],[object Object],[object Object],[object Object]
Overview of CRISPRs ,[object Object],[object Object],[object Object]
CRISPRs and HGT ,[object Object],[object Object],[object Object],[object Object],[object Object]
CRISPRs within GIs ,[object Object],[object Object],[object Object],CRISPRs are over-represented in GIs Domain of Life Number of Genomes Number of GIs Proportion of Genome in GIs Total Number of CRISPRs Expected CRISPRs in GIs Observed CRISPRs in GIs Significance (Chi-square Test)* Archaea 49 298 3.7% 206 7.7 14 0.020 Bacteria 306 4874 6.4% 837 53.3 114 8.1x 10 -18 Archaea & Bacteria 355 5172 6.1% 1043 64.0 128 1.6x 10 -16
Phage genes within GIs ,[object Object],[object Object],[object Object],[object Object],Phage genes are over-represented in GIs Genomic Regions Number of ‘phage genes’ Total number of genes in region Chi- Square Test Observed Expected 3 Inside GIs 1 6990 1264.22 165784 ~0 Outside GIs 1 12868 18593.78 2438303
Archaea and CRISPRs Prevalence of CRISPRs in Archaea genomes could result in reduced phage genes Archaea Bacteria Genomes containing a CRISPR 90% 40% Proportion of phage genes 0.10% 0.79% Proportion of GIs with a phage gene 5.1% 17.6%
GIs with CRISPRs and phage genes ,[object Object],GIs containing CRISPR(s) also contain an over-representation of phage genes -> suggesting that some CRISPRs are transferred by phage Genomic Regions Number of ‘phage genes’ Total number of genes in region Chi- Square Test Observed Expected 3 GIs containing CRISPR(s) 2 13 4.5 1500 5.7 x 10 -5 Outside GIs 2 812 820.5 274073
CRISPR conclusions ,[object Object],[object Object],[object Object]
Conclusions ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]
Acknowledgements Supervisor Dr. Fiona Brinkman Supervisor Committee Dr. Baillie Dr. Pio P. aeruginosa  LES Craig Winstanley Roger Levesque Bob Hancock Nick Thomson

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Computational prediction and characterization of genomic islands: insights into bacterial pathogenicity

  • 1. Computational prediction and characterization of genomic islands: insights into bacterial pathogenicity Morgan G.I. Langille Department of Molecular Biology & Biochemistry Simon Fraser University http://tinyurl.com/genomic-islands
  • 2.
  • 3.
  • 4.
  • 6.
  • 7.
  • 8.
  • 9.
  • 10.
  • 11.
  • 12.
  • 13. IslandPick: Outline Query Genome A Genome B Genome C Genome D Run Mauve Mauve (A & B) Extract unique regions Mauve (A & C) Mauve (A & D) Genome D Putative Genomic Islands BLAST Identify overlapping unique regions
  • 14. Selecting Comparative Genomes Run Mauve Mauve (A & B) Extract unique regions Mauve (A & C) Mauve (A & D) Genome D Putative Genomic Islands BLAST Identify overlapping unique regions Genome B Genome C Genome D Comparative Genome Selection (using CVTree distances) Query Genome A
  • 15.
  • 16.
  • 17.
  • 18.
  • 19. Example: Pseudomonas Tree Maximum Distance Cutoff = 0.42 Minimum Number of Genomes = 3 0.227 0.256 0.397 0.393 0.411 0.428 0.430 0 0.481 P. fluorescens Pf-5 P. putida KT2440 P. fluorescens PfO-1 P. syringae tomato DC3000 P. syringae phaseolicola 1448A P. syringae syringae B728a P. aeruginosa PAO1 P. aeruginosa PA14 Acinetobacter ADP1 Minimum Distance Cutoff = 0.10
  • 20. Predicting Similar Aged GIs GI Insertion Query Genome 1 genome < distance X Query Genome GI Insertion
  • 21.
  • 22.
  • 23.
  • 24. Negative Dataset Query Genome 1 genome > distance X GI Insertion Query Genome GI Insertion
  • 26.
  • 27. GI Prediction Accuracy Positive Dataset Negative Dataset Predicted Dataset Entire Genome TP FP FN Precision = TP / (TP + FP) Recall = TP / (TP + FN) TN
  • 28. GI Prediction Accuracy (Langille, et al.,2008) Tool Average number of nucleotides in GIs per genome (kb) Precision Recall Overall Accuracy SIGI-HMM 233 92 33.0 86 IslandPath/ Dimob 171 86 36 86 PAI IDA 163 68 32 84 Centroid 171 61 28 82 IslandPath/ Dinuc 444 55 53 82 Alien Hunter 1265 38 77 71 Literature* 639 100 87 96
  • 29.
  • 30.
  • 31.
  • 32.
  • 33.
  • 34.
  • 35. IslandPick – Manual genome selection
  • 37.
  • 38.
  • 39.
  • 40.
  • 41. LES Prophage (Winstanley, Langille, et al., 2008)
  • 42. LES Genomic Islands (Winstanley, Langille, et al., 2008)
  • 43.
  • 44.
  • 45.
  • 46.
  • 47.
  • 48.
  • 49. Archaea and CRISPRs Prevalence of CRISPRs in Archaea genomes could result in reduced phage genes Archaea Bacteria Genomes containing a CRISPR 90% 40% Proportion of phage genes 0.10% 0.79% Proportion of GIs with a phage gene 5.1% 17.6%
  • 50.
  • 51.
  • 52.
  • 53. Acknowledgements Supervisor Dr. Fiona Brinkman Supervisor Committee Dr. Baillie Dr. Pio P. aeruginosa LES Craig Winstanley Roger Levesque Bob Hancock Nick Thomson