Chau, LN

Chau, NVV

Hung, NM

Ngoc, NM

Rahman, M

Thanh, TT

Thanh, TTT

That, BTT

Trang, NH

Rahman, Motiur

Oxford University Research Archive

relationships between mice and retroviruses. In particular, weestablish the presence of a more ancient ERV component in themurine genome, comprised of isolated, highly degraded inser-tions. These sequences evidence a transition in murine evolu-tionary history, beginning about one million years ago, whereinthe ancient ERV families that have counterparts in humans andother large mammals were overtaken by a wave of newlyacquired and/or transpositionally active ERVs.A15 Rapid radiation of treponema pallidum pertenue in wild non-human primatesA. Düx,1,* V.J. Schuenemann,2 J.F. Gogarten,1,3,4 H.M. De Nys,1,5K. Nieselt,6 M.A. Mayhew,7 F.H. Leendertz,1 S. Calvignac-Spencer,1J. Krause,2,8,91Project Group Epidemiology of Highly PathogenicMicroorganisms, Robert Koch Institute, Berlin, Germany, 2Institutefor Archaeological Sciences, Archaeo- and Palaeogenetics,University of Tübingen, Tübingen, Germany, 3Department ofBiology, McGill University, Montreal, Canada, 4Max-Planck-Institute for Evolutionary Anthropology, Leipzig, Germany, 5UMI233, Institut de Recherche pour le Développement (IRD) andUniversity of Montpellier, Montpellier, France, 6Center forBioinformatics, University of Tübingen, Tübingen, Germany,7Department of Science, Natural Resources and Outdoor Studies,University of Cumbria, Carlisle, UK, 8Max-Planck-Institute for theScience of Human History, Jena, Germany and 9SenckenbergCentre for Human Evolution and Palaeoenvironment, University ofTübingen, Tübingen, GermanyBacteria of the species Treponema pallidum are causative agentsof venereal syphilis (Treponema pallidum pallidum), Bejel (T. p.endemicum), and yaws (T. p. pertenue) in humans. We docu-mented Treponema pallidum infections associated with diseasein wild sooty mangabeys (Cercocebus atys) in Taı̈ National Park,Côte d’Ivoire, and green monkeys (Chlorocebus sabaeus) fromBijilo Forest Park, Gambia and Niokolo-Koba National Park,Senegal. To examine the evolutionary relatedness of these tre-ponemes to those responsible for diseases in humans and forpreviously documented infections in baboons (Papio papio), weconducted a hybridization capture experiment to enrichTreponema pallidum DNA from samples collected from sympto-matic individuals. This approach allowed us to sequence thefull genomes of Treponema pallidum strains infecting sooty man-gabeys (n¼ 2) and green monkeys (n¼ 4). Phylogenomic analy-ses revealed that all Treponema pallidum strains infecting non-human primates are most closely related to the sub-species T.p. pertenue. Strains infecting humans and non-human primatesdo not appear to be reciprocally monophyletic. The star-likephylogenetic branching pattern of the T. p. pertenue clade, withshort basal branches receiving low statistical support, suggestsa rapid initial radiation across humans and non-human pri-mates. These results greatly broaden the known host range ofT.p. pertenue and suggest the existence of a vast zoonotic reser-voir that could possibly contribute to the failure of global eradi-cation efforts.A16 A distributed pan-viral typing frameworkMichael Vilsker and Koen DeforcheEmweb, Herent, BelgiumWith the increasing amount of DNA sequence informationobtained from new sequencing methods, opening up the possi-bility for a complete viral screen of a host, there is an increasingneed for the rapid and accurate identification of the virus typesas well as their epidemiological background. While in the past,typing tools have been developed and made available (such asthe Rega HIV-1 subtyping tool) and hosted at multiple sites,these tools require maintenance to track the ongoing evolutionof the virus. The setup and the maintenance of typing tools,which are often deployed at multiple sites, has been a chal-lenge. Within the EU-funded VIROGENESIS project (Horizon2020), the Rega typing tool framework is being redesigned toseparate clearly the framework from the specifics for an individ-ual tool. This will (1) enable an expert to independently andeasily setup, create and maintain a typing tool for a new patho-gen; (2) establish an online repository of typing tools/versions towhich participants can push updates and from which up-to-date versions can be fetched to a distributed network of servershosting the typing tools; and (3) create a pan-viral typing tool toidentify the correct pathogen and which will allow further anal-ysis of the sequences using the specialized typing tool for thatpathogen. The transformation of the framework is expected tobe completed by September 2016, at the same time co-evolvingexisting typing tools already available (including HIV, HCV,HTLV, Enterovirus, Norovirus), and new typing tools (includingChikungunya, Coronaviruses, Dengue virus, Zika virus) that arebeing designed by partners within the VIROGENESIS project.Within the VIROGENESIS project, the existing typing tools basedupon the Rega typing tool framework will evolve into a distrib-uted pan-viral typing tool.A17 Molecular characteristics of hepatitis B virus (HBV) isolatedfrom chronic hepatitis B patients in South VietnamNguyen Hoa Trang,1 Bui Thi Ton That,2 Tran Thi Thanh Thanh,1Le Ngoc Chau,1 Tran Tan Thanh,1 Nghiem My Ngoc,2Nguyen Manh Hung,2 Nguyen Van Vinh Chau,2 Motiur Rahman,11Oxford University Clinical Research Unit, Wellcome Trust MajorOverseas Programme, Ho Chi Minh City, Viet Nam and 2TheHospital for Tropical Diseases, Ho Chi Minh City, Viet NamChronic infection with hepatitis B virus (HBV) is a major healthproblem due to its worldwide distribution and its long-termeffects. Vietnam is country with a high HBV burden and theprevalence of chronic HBV infection in general population is8.8–12.3%. In East Asia, the most common HBV genotypes are Band C. Despite high prevalence of HBV, data on HBV (genotypeand subgenotype, virulence markers, drug resistance mutationsand prevalence of recombinant strains) is limited in Vietnam.There are only few reports on HBV genotypes in Vietnam,mostly based on pre-S/S gene sequences. We have analyzedwhole genome sequence of 98 HBV isolates from chronic HBVpatients attending at Hospital for Tropical Disease, Ho Chi MinhCity, Vietnam, who were under treatment for 1–6 years. HBVgenome was amplified in 4 overlapping fragments (777–1,136 bp) and the amplicons were subjected to deep sequencingby using Illumina MiSeq system. Sequence assembly, genomeanalysis and phylogenetic analysis were performed withinGeneious package. A sequence was assigned to a certain geno-type and subgenotype if it was contained within a well-sup-ported phylogenetic cluster (bootstrap value>75%) and theintra-genotypic nucleotide divergence was<7.5 and>4.5%,respectively. Mutations in Basal core promoter (BCP), pre-CORE,and CORE gene regions were determined by comparing withreference sequences. Finally, screening of minor (sub-consen-sus) variants was performed using the SNP detection tool avail-able in Geneious. 1% frequency and 500-fold coverage werechosen as cut-off values. Among the isolates, 71.43% were geno-type B, 27.55% were genotype C and one isolate was a recombi-nant (between B and C). Among genotype B isolates, 65 weresubgenotype B4 (92.86%) and 5 were B2 (7.14%). 92.6% of subge-notype C belong to C1, 3.7% is subgenotype C2 and the remain-ing 3.7% to C3. Mutations G1752A, T1753C, G1757A, A1762G/T,G1764A and C1766G on BCP and CORE were found in 76 of 98S6 | Virus Evolution, 2017, Vol. 3, Suppl. 1(77.55%) strains; and in 96.3% of subgenotype C (26/27) isolates.Stop codon mutation on Pre-Core, G1896A, was found in 23 of 98patients (23.47%); among them, 20 of 23 (86.96%) are genotype B.Mutations related to antiviral drug resistance with frequencymore than 1% in the reverse transcriptase (RT) domain ofPolymerase gene were identified. 83.67% sequences have morethan one mutation. 50% sequences have mutation at rtI169V/M,and all belong to sub-genotype B. Mutations at rtM204I/V andrtM250I were present in 23.47% and 11.22% of isolates (23 and 11of 98, respectively), and was higher in sub-genotype C (P-values¼ 0.06 and<0.01, respectively). Our study provides highprecision whole genome sequences of the Vietnamese HBV pop-ulation and shows potential information related to treatmentprediction for HBV infection as well as viral evolution. A furtherstudy with larger sample size and intensive analysis combinedwith clinical information promises more useful data for treat-ment and invention.A18 Random amplification with next-generation sequencing tocover HIV and HCV full-length genomesL. Cuypers,1,* N. Conceição-Neto,1 T. Dierickx,1 Y. Schrooten,1,2 B.Vrancken,1 E. Van Wijngaerden,3,4 F. Nevens,5,6 A.-M. Vandamme,1,7J. Matthijnssens,1 K. Van Laethem,1,21Department of Microbiology and Immunology, Rega Institute forMedical Research, Clinical and Epidemiological Virology, KULeuven – University of Leuve, Leuven, Belgium, 2AIDS ReferenceLaboratory, University Hospitals Leuven, Leuven, Belgium,3Department of Microbiology and Immunology, Clinical andInflammatory Disorders, KU Leuven – University of Leuven,Leuven, Belgium, 4Department of General Internal Medicine,University Hospitals Leuven, Leuven, Belgium, 5 Department ofClinical and Experimental Medicine, KU Leuven – University ofLeuven, Leuven, Belgium Hepatology, 6 Department ofGastroenterology and Hepatology, University Hospitals Leuven,Leuven, Belgium and 7Center for Global Health and TropicalMedicine, Microbiology Unit, Institute for Hygiene and TropicalMedicine, University Nova of Lisboa, Rua da Junqueira 100 Lisbon1349-008, PortugalSince both HIV and HCV are known to be highly variable andinfections are treated with combination therapies that targetdifferent viral proteins, random amplification followed by next-generation sequencing could be an attractive alternative toSanger sequencing for obtaining complete viral genomes.Moreover, one experimental approach could enable the simulta-neous mapping of virus and human origin sequence reads,which is of particular interest for the detection of single nucleo-tide polymorphisms (SNPs) associated with disease progressionand treatment response. A plasma sample obtained from apatient infected with HIV-1 recombinant subtype B and F1, anddiagnosed with AIDS; and another sample from an HCV infectedpatient who progressed to fibrosis stage F1 were included in thispilot study. They were both characterized by a high viral load,i.e.>10M copies/ml for HIV and>3.5M IU/ml for HCV. Sampleswere directly extracted or pre-treated with homogenisation,centrifugation and filtration. Subsequently, purified nucleicacids were subjected to targeted (HIV-1 PR-RT and HCV NS3 pro-tease) amplification and Sanger sequencing, or to randomamplification followed by Illumina sequencing (2M paired endreads attributed to each sample). Quality of the Illumina readswas checked using FastQC, followed by a de novo assembly ofsample-specific contigs using the VICUNA and V-FAT softwarepackages. In total, 0.3% (n¼ 5557) and 0.02% (n¼ 421) of thereads mapped to the HCV genome, respectively with or withoutvirus enrichment steps. This sufficed for the de novo generatedcontigs to cover 98.3 and 51.3% of the HCV genome with andwithout pre-treatment steps, respectively. Lower numbers ofreads could be mapped for HIV-1 (821 versus 196 without virusenrichment), resulting in only 45.6 and 21.7% coverage of thefull-genome. For both samples, the number of reads (<2M) wastoo low to map SNPs of interest within the human genome.Subject to adaptations in the protocol, such as improved enrich-ment strategies and deeper sequencing, this approach could besuitable for the surveillance of local HIV-1 and HCV epidemics.More extensive studies are required to investigate its usefulnessin clinical research.A19 Genetic markers for protease inhibitor drug resistance inregions outside of the protease geneJ. Giandhari,1,* M. Gordon,2 C. van Vuuren,3 P. Moodley,4T. de Oliveira,11Africa Centre for Health and Population Studies, College of HealthSciences, University of KwaZulu-Natal, 2HIV PathogenesisProgramme, University of Kwa-Zulu Natal, Durban, South Africa,3Military Hospital Division of Infectious Diseases, University of theFree State, South Africa and 4Department of Virology, NationalHealth Laboratory Service, University of KwaZulu-Natal, Durban,South AfricaThere are currently 6.8 million individuals infected with HIV-1 inSouth Africa. Nearly half of these individuals are on antiretroviraltherapy. Protease inhibitors (PIs) form part of the South Africannational treatment guidelines as a first-line regimen for paediat-ric patients and a second-line regimen for adults. However, suc-cessful treatment is often hindered by the development of drugresistance. Resistance to PIs is characterised by a stepwise accu-mulation of mutations in the protease gene. However, studieshave shown a low frequency of mutations in patients failing PIs.Regions outside of the protease, such as gag and more recentlyenv, have been associated with PI drug resistance in the absenceof protease mutations. We aim to identify genetic markers in thegag and envelope genes which are associated with PI treatmentfailure. Stored plasma samples from HIV-1-infected patients fail-ing PI-based therapy (n ¼ 500) will be collected from collaboratorsin South Africa and whole genome sequences from PI-naı̈vepatients will be downloaded from the Los Alamos SequenceDatabase. Failure will be defined as having two consecutive viralloads>1,000 cpm after being on a PI-based regimen for6months. The whole HIV-1 genome will be amplified from the PIfailures and sequenced using Illumina Miseq. Sequences will bealigned to a reference and analysed for single nucleotide poly-morphisms (SNPs) in all HIV genes using Geneious v 8.1.8 andGATK software application. Genome wide association analysisusing PLINK will be performed to identify SNPs in PI treatedpatients that are associated with treatment failure. Episodicdirectional selection model such as MEDS and IFEL will be used toidentify mutations that occur at specific amino acid positionsand confer resistance to PIs. The previously mentioned methodsallow the identification of drug resistance mutations without theneed of baseline samples. Preliminary data from GWAS analysison 26 patients failing PI-based therapy has identified codons inenvelope and gag which are significantly associated with PI fail-ure. Further examination of these sites in the viral minority popu-lation has shown that they are present at>2% of the viralpopulation. By increasing our sample size we will obtain a morecomprehensive and robust analysis of the role of regions outsidethe protease gene in PI resistance. Analysis of the control samplesis currently being performed.21st International BioInformatics Workshop on Virus Evolution and Molecular Epidemiology | S7

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A17 Molecular characteristics of hepatitis B virus (HBV) isolated from chronic hepatitis B patients in South Vietnam

A17 Molecular characteristics of hepatitis B virus (HBV) isolated from chronic hepatitis B patients in South Vietnam

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