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In silico Footprinting and Genomic Signature Analysis for Encephalic and Hemorrhagic Viruses

In silico Footprinting and Genomic Signature Analysis for Encephalic and Hemorrhagic Viruses. Willy A. Valdivia-Granda Orion Integrated Biosciences, Inc. Willy.Valdivia@orionbiosciences.com. Outline of this talk. Encephalitic and Hemorrhagic Viruses

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In silico Footprinting and Genomic Signature Analysis for Encephalic and Hemorrhagic Viruses

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  1. In silico Footprinting and Genomic Signature Analysis for Encephalic and Hemorrhagic Viruses Willy A. Valdivia-Granda Orion Integrated Biosciences, Inc. Willy.Valdivia@orionbiosciences.com

  2. Outline of this talk Encephalitic and Hemorrhagic Viruses Arenaviridae, Bunyaviridae, Flaviviridae, Filoviridae In silico Genomic Footprinting Genomic Signature Detection Binary Clustering Analysis of Genomic Signatures Examples: Flavivirus and Filoviruses Directions in the Use of Genomic Sigantures Flavi-chip, Arena-chip, Bio-detection, Multimeric Vaccines

  3. Dissemination of Infectious Diseases BioScience, May 2003: One million tons of dust may contain 10 quadrillion microbes (USGS).

  4. Encephalic and Hemorrhagic Viruses: NIAID Cat. A-C BSL4 Flaviviridae Hepatitis, Dengue, Yellow fever, Japanese encephalitis and West Nile viruses Filoviridae Ebola and Marburg viruses Arenaviridae Argentine, Bolivian, and Venezuelan hemorrhagic fevers Lassa fevers Bunyaviridae Hantavirus, the Congo-Crimean, Rift Valley fever virus

  5. West Nile Virus Summer 2003

  6. 100,000X 123,000X 25 Å 4 Å Ecological Genomics and Biocomplexity of Viruses

  7. Flaviviridae Family Hepaciviruses (~9.4 Kb) 5’ UTR CAP NS1 C E1 E2 NS2 NS3 NS4 NS4B NS5A NS5B 3’ UTR Flavivirus (~ 11Kb) 70 Species transmitted by Mosq. Ticks, Non-Vector 5’ UTR IRES NS2B E NS5B C PreM NS1 NS2A NS3 NS4A NS4B 3’ UTR Pestiviruses (~ 12Kb) 5’ UTR IRE NS1 NS4A NS4B NS5A NS5B C PreM E E2 NS2-3 3’ UTR

  8. Molecular Detection Methods for Viruses Sub- Species Family Genus Species Strain DNA Sequencing DNA-DNA reassociation RT-PCR Degenerated-PCR Immunology Microarray

  9. Murine Hepatitis Virus (MHV) 33/37 AA Murine Hepatitis Virus (MHV) 52/157 AA Infectious Bronchitis Virus (IBV) 32/32 nt Viral Sequence Recovery Using DNA Microarrays Viral sequences were physically scraped, amplified, cloned, and sequenced Prototypical Coronavirus Genome Structure Wang and DeRisi et al. 2003 PLoS Biology | Volume 1 | Issue 2 | http://biology.plosjournals.org

  10. Molecular Detection Methods for Viruses Nucleotide substitution Why Aminoacid? Information Content 3:1 Nucleotide substitution 10-3 per site per year

  11. Flavivirus (~ 11Kb) NS2B E NS5B C PreM NS1 NS2A NS3 NS4A NS4B 20 Genomes Target Generation Profile Comparison Ungapped Whole Genomic Footprinting Profile Generation Genomic Footprinting

  12. Ungapped Whole Genomic Footprinting Mosquito Ticks No-vector

  13. Core genome Tick-Borne Adaptation Region Mosquito Borne Cyclic Region Non Vector Ungapped Whole Genomic Footprinting

  14. 59 Species of the Flavivirus Genus Flavivirus Genomic Signature for NS5

  15. 0 0 0 0 1 1 1 1 1 1 2 2 Binary classification of Flaviviruses

  16. >NP_476520.1|Deer tick virus|ctb30|8160|NS3 >NP_476520.1|Deer tick virus|ctb30|9651|NS5

  17. Arising Biological Questions About Genomic Signatures Are genomic signatures relevant for pathogen replication? Silencing of host genes. MHC? Competitive advantage over other viral serotypes Role in virus recombination and the generation of new variability Are some genomic signatures duplications defining host range and are related with vector transmission?

  18. Binary classification of Mosquito Borne Flaviviruses

  19. NS5 Genomic Signature Phylogenic Incongruence Homologous Recombination Regions - Natural selection - Mechanistic/ecological - Genome segment reassortment J. Virol., April 1, 2004; 78(7): 3319 - 3324. J. Virol., February 15, 2004; 78(4): 2114 - 2120.

  20. Genomic FootprintingHemorrhagic Viruses Ebola Virus

  21. Ebola-Zaire 94 75 68 59 66 72 95 23 45 28 44 48 8 Ebola-Reston 68 75 23 45 28 59 66 72 95 44 48 8 Marburg 28 23 45 27 94 74 68 40 50 59 66 64 1 75 41 95 Marburg Lake Victoria 27 94 74 68 28 40 50 59 66 64 1 75 41 95 45 23 1 25 50 75 100 125 150 175 200 225 250 275

  22. Phylogenomic Analysis of Flavivirus Genomic Signatures Valdivia-Granda et al. 2002.

  23. 1 Security and Administration Application Firewall Process Flow a. The authorization process begins from the client and is passed to the Security and Administration API . This process select the services API. b. The Administration selects the Data Analysis API and the data request is passed to the Data Abstraction Layer (DAL). c. The data mapping function is invoked, the specified application and URI are referenced and the proper driver. d. The various data drivers implement the Metadatabse layer (MDL) produce common requests and result sets are selectively cached in the Data Abstraction Server. e. Once data is delivered from one of the database, it may be sent to the analysis application. Sequence Analysis Application 2D and 3D Sequence Visualization Microarray Analysis Application Cytogenomic Map Visualization Client Proteomic Analysis Application Transcriptional Network Visualization Global Schema 2 Data Mapping Function The data mapping function maps objects and their attributes to specific databases Data adapters are specific implementations of data drivers for different genomic databases (Metadatabase layer) JDBC 3 Oracle DB2 XML Sybase Flat File Other

  24. 1400 1200 1320 1000 Malaria HIV 800 Viruses 600 Dengue 400 Cancer 158 200 40 16 1 0 Development of New Detection Devices Number of published papers

  25. Collaborators UC Berkeley Centers for Diseases Control University of Zurich, Switzerland MIT Walter Reed Army Institute of Research San Diego Supercomputing Center UMass Med School Pasteur Institute, France Pasteur Institute, Senegal Sandia National Laboratories December 2002 KDD-cup August 2003

  26. (T) (A) (B) (T+A+B) (T:A:B) (2T)

  27. Arising Biological Questions About Genomic Signatures Genomic signatures and the Origen of Life? Lee, D. H.; Granja, J. R.; Martinez, J. A.; Severin, K.; Ghadiri, M. R. "A Self-Replicating Peptide". Nature 1996, 382, 525-28. Hutton, T.J. (2002) Evolvable Self-Replicating Molecules in an Artificial Chemistry. Artificial Life 8(4):341-356. Repeated self-replication and simple evolvability http://www.eastman.ucl.ac.uk/~thutton/Evolution/ Martin A. Nowak, Karl Sigmund Phage-lift for game theory. Nature398, 367 - 368 Paul E. Turner, Lin Chao . Prisoner's dilemma in an RNA virus. Nature398, 441 - 443

  28. Dobrava 17 4 13 6 16 3 4 16 5 Seoul 2 17 16 4 13 6 3 8 4 5 Sin Nombre 8 13 4 4 17 15 1 16 6 16 3 1 25 50 75 100 125 150 175 200 225 250 275 Ungapped Whole Genomic Footprinting

  29. Phylogenomic Analysis of Hemorrhagic Fevers

  30. Viral Extinction threshold Viral Isolation Lethality Dispersion Viral Isolation Viral Extinction threshold Time

  31. Risk for dengue fever (DF) among travelers to Thailand, 2002. http://www.cdc.gov/ncidod/EID/vol10no5/03-0495-G2.htm Christina Frank,* Irene Schöneberg,* Gérard Krause,* Hermann Claus,* Andrea Ammon,* and Klaus Stark* *Robert Koch Institute, Berlin, Germany

  32. Functional Modules • Module directionality • Visualization • Large Scale Organization • Evolutionary perspective • Ecological Level? • Pathways • Directionality • Association accuracy • The Central Dogma • Gene finding algorithms • Mutation • Alternative splicing • Folding dynamics

  33. Viral Adaptation Viral Life Cycle Viral Structural Changes Evolutionary Dinamics Sequence Space Selection

  34. 90% of the Zairian cases and 50% of the Sudanese cases resulted in death. Marburg hemorrhagic fever is between 23-25%.

  35. Genomic Signature Count

  36. A comprehensive genomic analysis of the genus Flavivirus genus suggest the existence of a core viral genome composed by 47 elements each with a length of 12 aminoacids. For 7 of the viral genomes there is at least one copy of each element. However, several genomic signatures are duplicated up to three times. But it remains unclear if the generation of genomic signatures are cyclic events. Our analysis shown that duplication of genomic signatures and the mutation still a relevant process in viral genome evolution, and could be is involved in viral recombination and self interaction. As mutation pressures selected fitted individuals, new species with novel characteristics emerged. However, there is a tradeoff between viral pathogenesis and dispersal. Conclusions

  37. 4.5 microns Each of these particles is about 4.5 microns long--about one-twentieth the diameter of a human hair, which is about 100 microns. Alternating gold and silver stripes create the "barcode" pattern on these tiny particles. When viewed in blue light under a microscope, silver is much more reflective than gold, making different-patterned particles easy to identify.

  38. Aminoacid Usage in the Flavivirus and Filovivirus` Filovirus Flavivirus %

  39. Genomic Footprinting and Biological Complexity

  40. Microarray Detection Analysis

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