rologie i - European Congress of Virology

5 th European Congress of Virologypneumonia during follow up and both patients had extremely low B cellscounts. The low B cell count might be caused by the infection with TTMV,or it could be that an infection with the TTMV is poorly cleared due to thelow B cell immunity.REF 410Powerful sequence similarity searches identify homologs in manyapparently “orphan” viral proteins: a guide to genome annotatorsDavid KARLIN 1,2 , Durga KUCHIBHATLA 3 , Westley SHERMAN 3 ,Betty CHUNG 4 , Shelley COOK 5 , Georg SCHNEIDER 3 , BirgitEISENHABER 31 Department of Zoology, Oxford, UNITED KINGDOM; 2 Departmentof Structural Biology (STRUBI), Oxford, UNITED KINGDOM;3 Bioinformatics Institute (A*Biomedical Sciences Institutes), Singapore,SINGAPORE; 4 Department of Plant Sciences, Cambridge, UNITEDKINGDOM; 5 Life Sciences Parasite and Vectors Division, London, UNI-TED KINGDOMGenome sequences of new viruses often report many “orphans” or “taxonspecific” proteins apparently lacking homologs. However, we show thatcommonly used sequence similarity detection methods such as Blast orPsi Blast overlook homologs (predictably, since viral proteins evolve veryfast). We analysed a dataset of proteins characterized as “genus specific” byBlast, using powerful methods developed recently, such as profile profilecomparison (HHblits, HHpred). Homologs were detected in other generafor a quarter of these proteins. In depth manual analyses on a subset of theremaining sequences, guided by contextual information such as taxonomy,gene order or domain order, identified distant homologs for a third ofthem. Extrapolating these results, a combination of powerful automatedmethods and manual analyses may uncover distant homologs for overhalf of proteins found to be “orphans” by Blast. As a case in point, were analyzed the genome of a bee pathogen, chronic bee paralysis virus(CBPV), and identified homologs, and probable functions, for almost allits “orphan” proteins. CBPV encodes a previously unrecognized enzyme,a new type of virion membrane protein homologous to that of insect andplant viruses with a different morphology, and a virion glycoprotein alsofound in these viruses. We used only software available through web based,user friendly interfaces, and will present recommendations for virologistson how to use these methods to annotate (or re annotate) genomes.REF 411Unbiased detection of infectious agents in respiratory syndromes ofpoultry using a metagenomic approachEtienne LIAIS 1,2 , Guillaume CROVILLE 1,2 , Jérome MARIETTE 3 ,Christopher KLOPP 3 , Cécile DONNADIEU 4 , Jérome LLUCH 4 ,Mariette DUCATEZ 1,2 , Jean Luc GUÉRIN 1,21 Université de Toulouse, INP, ENVT, Toulouse, FRANCE; 2 INRA, UMR1225, Toulouse, FRANCE; 3 Plateforme bioinformatique Toulouse MidiPyrénées, UBIA, INRA, Castanet Tolosan, FRANCE; 4 GeT PlaGe, Genotoul,INRA, Castanet Tolosan, FRANCELittle is known so far on respiratory viruses in poultry. In order to identifyall the viral pathogens in duck respiratory tract, tracheal swabs were collectedon birds showing respiratory clinical signs and/or egg drop syndromesand were submitted to a metagenomic analysis. Using a high throughputsequencing approach (Miseq, Illumina), we aimed at describing therespiratory viral and bacterial flora of ducks showing respiratory clinicalsigns and/or egg drop syndromes. Viral nucleic acids were concentratedby centrifugation and DNase/RNase treatment. DNA and RNA wereextracted and PCR amplified using random primers (Victoria & al., 2009).Around 2.8 million reads (of about 150 nucleotides) were generated.The assembled contigs and singlet sequences were compared to Genbankvirus database using GAAS software. Sequences of bacteriophages(Siphoviridae, Myoviridae, Podoviridae, Microviridae) and animal viruses(Retroviridae, Reoviridae, Adenoviridae„Picornaviridae, and Paramyxoviridae)were mainly identified. These sequences were submitted to aphylogenetic analysis and their putative association with respiratory pathologywas also subsequently assessed. Altogether, we described an avianrespiratory virome. Deep sequencing is a powerful approach to detectmost infectious agents in a clinical case. It also allows for the detectionof the opportunistic pathogen flora, unrelated to clinical signs. This studyincreases our understanding of the viral diversity in ducks and highlightsthe complexity of co infections in poultry respiratory tract.REF 412A novel virus discovery approach to identify unrecognizable virusesBas OUDE MUNNINK 1 , Seyed Mohammad JAZAERI FARSANI 1 ,Martin DEIJS 1 , Marta CANUTI 1 , Jiri JONKERS 1 , JoostVERHOEVEN 1 , Greet IEVEN 2 , Herman GOOSSENS 2 , MatthewCOTTEN 3 , Lia VAN DER HOEK 11 Amsterdam Medical Center, Amsterdam, THE NETHERLANDS;2 University of Antwerp, Antwerpen, BELGIUM; 3 Sanger Institute, Cambridge,UNITED KINGDOMViral infections remain a major cause of human diseases. Nevertheless,it is assumed that there are pathogenic viruses that have escaped identification,while the threat of new viruses adapting to human hosts remainsunabated. Discovery of new viruses has in the last decade been boostedby viral metagenomics: second generation virus discovery whereby tenthousands of sequences from a clinical sample are generated. With thedevelopment of the VIDISCA 454 technology, we were one of the firstto exploit these new possibilities, and identified several novel viruses.However, this experience also showed the limitations of next generationsequencing based approaches. In clinical samples positive identificationis entirely based on similarity with known virus families. Unknown virusfamilies still escape detection. We have developed a novel VIDISCA basedapproach which will allow identification of viruses that lack similarity toknown viruses. Key thereby is that infected patients show an adaptiveimmune response resulting in virus specific antibodies in blood. The IgGin convalescent serum can be used to capture and enrich viruses. Whenthe enriched fraction is used as input in VIDISCA 454, unrecognizablenew viruses can be detected. This new method was evaluated with severalvirus infections: 13 respiratory and 7 intestinal infections. Identificationvia Enrichment VIDISCA 454 allowed identification of all 13 respiratoryviruses, and 6 of 7 intestinal viral infections.REF 413Bufavirus, a Novel Human Virus in the Family ParvoviridaeElina VÄISÄNEN 1 , Tung G. PHAN 2,3 , Eric DELWART 2,3 , EveliinaTARKKA 4 , Klaus HEDMAN 1,4 , Maria SÖDERLUND VENERMO 11 Department of Virology, Haartman institute, University of Helsinki, Helsinki,FINLAND; 2 Blood Systems Research Institute, San Francisco, USA;3 University of California, San Francisco, USA; 4 Helsinki University CentralHospital Laboratory Division, Helsinki, FINLANDParvoviruses are small, non enveloped viruses with 4 6 kb ssDNAgenomes. They infect diverse animals from insects to humans, and causediseases with mild to severe symptoms. Since 2005, nine new humanparvoviruses have been found and last year, metagenomic analysis offeces of children with acute diarrhea in Burkina Faso revealed a novelhighly divergent parvovirus provisionally named Bufavirus. The initialbufavirus sequence showed in the NS1 region