Sequencing

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11th Annual Sequencing, Finishing, and Analysis in the Future Meeting USAGE OF SEQUENCE INDEPENDENT SINGLE PRIMER AMPLIFICATION NEXT GENERATION SEQUENCING (SISPA NGS) FOR WHOLE GENOME SEQUENCING OF HANTAAN VIRUS Wednesday, 1st June 20:00 La Fonda Mezzanine (2nd Floor) Poster (PS‐2b.07 Daesang Lee 1 , Dong Hyun Song 1 , Won‐keun Kim 2 , Se Hun Gu 1 , Jeong‐Ah Kim 2 , Seung‐Ho Lee 2 , Jin Sun No 2 , Prof. Jin‐won Song 2 , Seong Tae Jeong 1 1 Agency for Defense Development, 2 Korea University Hantavirus, a family of bunyaviridae, is a single‐stranded, negative sense RNA virus containing tripartite genomes. Hantavirus infection causes hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS) with the mortality rate of 1‐36%. The outbreak or endemic infection of hantavirus are a critical threat for world public health because of the lack of effective prophylactic and therapeutic strategies. In addition, according to the Center for Disease Control and Prevention (CDC), hantavirus belongs to Category C of the bioterrorism agent categories, suggesting it is a possible biological agent to threat the national security because of easy production, rapid transmission and less understandable pathogenesis. Recently, the usage of NGS provides a potential tool to acquire whole viral genome sequence for the advance of diagnostics and vaccines. Here, based of Sequence Independent Single Primer Amplification Next Generation Sequencing (SISPA NGS), we have attempted whole genome sequencing of Hantaan virus (HTNV), the prototype of hantavirus and a etiology of HFRS, from endemic areas in the Republic of Korea (ROK). In this study, SISPA NGS applied to the whole genome sequencing and the detection of HTNV tripartite genomes from the isolates. The results showed that whole genome sequences of seven HTNV strains was recovered by the NGS and validated by phylogenetic analysis with hantavirus. Thus, SISPA NGS will be a robust tool for the accurate genomic‐based diagnosis and whole genome sequencing of hantaviruses in natural reservoirs as well as HFRS and HPS patients. 95
11th Annual Sequencing, Finishing, and Analysis in the Future Meeting GENOME SEQUENCE OF RIFT VALLEY FEVER VIRUS ISOLATES FROM THE 2008-2010 DISEASE OUTBREAK IN SOUTH AFRICA Wednesday, 1st June 20:00 La Fonda Mezzanine (2nd Floor) Poster (PS‐2b.08) Prof. Phelix Majiwa 1 , Rachel Maluleke 2 , Alison Lubisi 2 , George Michuki 3 , Maanda Phosiwa 4 , Phemelo Kegakilwe 5 , Steve Kemp 3 1 ARC Onderstepoort Veterinary Institute and University of Pretoria, Faculty of Veterinary Science, 2 ARC‐OVI/University of Pretoria, 3 ILRI, 4 ARC‐OVI, 5 Veterinary Services, Northern Cape Changing climate and increasing pressure to produce food sufficient for the increasing human population lead to higher frequency of contacts between humans and animals particularly in the developing countries. This leads to greater likelihood of disease spill‐over from animals to human, for it is known that a majority of infectious disease of man in the developing countries originate in animals (Foresight, Infectious Diseases: Preparing for the Future (Office of Science and Innovation, London, 2006)). One such disease is Rift Valley fever (RVF), transmitted from infected animals to man by mosquitoes. Humans become infected by Rift Valley fever virus also through contaminative contact with tissues of infected animals. The disease causes major loses in livestock and has significant negative impact on the livelihoods of livestock keepers. The virus responsible for this disease has a potential for dual use for inhumane purposes. Rift Valley fever is a re‐emerging zoonotic disease. Recent outbreaks (2008, 2009 and 2010) in South Africa occurred unexpectedly and with increased frequency. As of August 2010, there were 232 human cases, with 26 confirmed deaths. In order to obtain comprehensive information on the genetic composition of the RVF viruses (RVFVs) circulating in South Africa, genome sequence analyses was undertaken on RVFVs isolated from samples collected over time from animals at discrete foci of the outbreaks, with emphasis on isolates from 2008‐ 2010 outbreaks. The genome sequences of these viruses were compared with those of the viruses from earlier outbreaks in South Africa and elsewhere. Representative virus isolates from the 2008‐2010 RVF outbreak cluster into two distinct clades, one similar to clade C and the other to clade H previously described by Grobbelaar et al (Grobbelaar AA, Weyer J, Leman PA, Kemp A, Paweska JT, Swanepoel R. Molecular epidemiology of Rift Valley fever virus. Emerg Infect Dis. 17:2270‐6, 2011). The isolates from outbreaks of 1955, 1974 and 1975 all which fall into a third cluster designated Z. A majority (87.5%) of 2008 outbreak isolates are in Clade C (Grobbelaar et al +2011), which contains only one isolate from the 2009 outbreak. A majority of the 2009 (33%) and 2010 (67%) outbreak isolates clustered in clade H. Rift Valley Fever viral genome sequences determined by Bird et al (J Virol. 81:2805‐16, 2007), included in the analyses for comparison, did not cluster with the 2008‐2010 isolates from the disease outbreaks; instead, they clustered with the isolates from 1955, 1974 and 1975 South African outbreaks. The data will be presented and discussed in the context of phylogenetic relationships among the isolates, and implicit recombination, if any, in the genes encoding glycoproteins Gc and Gn, which have a role in protective host immune response. 96
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Reliable solutions for focused NGS
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Sequencing

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