bluuebunny/crossref_metadata_2025_split huggingface dataset:
A6 Does treatment cause virulence changes in HIV-1?
Authors: Hodgson David , Hué Stéphane , Panovska-Griffiths Jasmina , Iamarino Atila , Atkins Katherine E 
Published: August 2019

Abstract
Antiretroviral treatment (ART) has provided substantial benefits for HIV-1-infected patients and has reduced incidence in areas with high uptake since its introduction in the late 1980s. As ART has led to shifts in the worldwide epidemiology of HIV-1, it may also have the potential to cause concomitant selective pressure on the virus population. Evidence for changes in HIV-1 virulence since the introduction of ART appears to be inconsistent. As well as reviewing both empirical and theoretical studies on the likely impact of ART on HIV-1 virulence, we developed a mathematical framework to evaluate the likely impact of ART on virulence selection under the widespread treatment programs and the future impact of recent test-and-treat recommendations. By quantifying both the relationship between virulence changes with the transmissibility through disease progression and the speed of diagnosis and treatment, we reconcile observational studies on virulence changes with the mathematical model predictions. On adoption of new test-and-treat programs—synonymous with early detection and immediate treatment—it is likely that increased virulence will be observed. Our findings highlight the potential public health consequences of mass treatment and the ensuing requirement for greater access and adherence to nullify the public health effect of these virulence changes.
A12 Modeling residual HIV replication and the emergence of drug resistance on ART
Authors: Michael Bale J , Valerie Boltz F , Mellors John W , Coffin John M , Hughes Stephen H , Kearney Mary F 
Published: August 2019

Abstract
There are conflicting reports regarding the presence of low-level HIV replication during suppressive antiretroviral therapy (ART). We simulated varying levels of replication and estimated the number of generations needed to obtain linked, drug resistance mutations to explore the effects of replication during ART. HIV replication was simulated with varying population sizes (10 to 3,000,000). Each population size was modeled ten times. Each genome was given a Poisson-distributed number of mutations according to its length and the average replication error rate (3.4 × 10−5 sub/nt/cycle). Simulations were run a maximum of 20,000 generations with endpoints defined as detection of a variant with resistance mutations to at least two ARVs. In all simulations, variants that were resistant to all three ARVs emerged in less than 20,000 generations. The time to emergence ranged from 148–16,156 generations in the various simulations, depending on the replicating population size (4.8 months to 44.3 years if the generation time is 1 day). Clinically detectable virologic failure can result from linkage of two mutations conferring resistance to two ARVs in a regimen. In our simulations, two linked mutations emerged in from 9 to 6,429 generations (9 days to 17.6 years). Our simulations suggest that in patients continually suppressed on ART for at least 10 years, the replicating population size would have to be less than ten, or virologic failure would have occurred from emergence of two ARV-resistant variants. Because most patients on ART do not experience virologic failure, our simulations suggest that any residual replicating population on ART is very small and thus not likely to either sustain or significantly contribute to the HIV reservoir.
A19 The origin of acute HCV in HIV-infected men who have sex with men in the Netherlands
Authors: Popping S , Boucher C , Verjans G , Rijnders B , van de Vijver D 
Published: August 2019

Abstract
In recent years, several outbreaks of hepatitis C virus (HCV) infections have been observed among HIV-infected men who have sex with men (MSM) in Europe. In the Netherlands, high incidence rates of 10/1,000 person-years are reported. In this analysis, we describe whether acute HCV is linked to specific transmission networks. A total of 50 Dutch HIV-infected MSM, diagnosed with acute HCV genotype 1a between 2013 and 2014, were included. Target enrichment for viral nucleic acid separation and deep sequencing were used to recover whole HCV genomes. Phylogenetic trees were constructed by use of the maximum likelihood method. Pairwise distance matrices were generated by use of the Kimura-2 parameter distance estimation method. The consistency of the phylogenetic clustering was tested by bootstrap analysis with 1,000 replicates. A cluster was defined as having a genetic distance of at most 1.5 per cent and bootstrap values of 100 per cent. The most recent common ancestor was estimated with a coalescent-based model with a Bayesian statistical framework. Four transmission clusters were identified that included a total of 38 patients (76% of the total). The clusters were indicative of recent outbreaks, as highlighted by small genetic distances and a most recent common ancestor after the year 2000, when the first cases of HCV infection in HIV-infected MSM were reported. The HCV epidemic among HIV-infected MSM is a young epidemic, with most of the acute infections linked within the four major transmission networks. Prevention strategies identifying and targeting these transmission networks can potentially curb the epidemic.
A30 Avian influenza viruses in wild birds: Virus evolution in a multi-host ecosystem
Authors: Venkatesh Divya , Poen Marjolein J , Bestebroer Theo M , Scheuer Rachel D , Vuong Oanh , Chkhaidze Mzia , Machablishvili Anna , Mamuchadze Jimsher , Ninua Levan , Fedorova Nadia B , Halpin Rebecca A , Lin Xudong , Ransier Amy , Stockwell Timothy B , Wentworth David E , Kriti Divya , Dutta Jayeeta , van Bakel Harm , Puranik Anita , Slomka Marek J , Essen Steve , Brown Ian H , Fouchier Ron A M , Lewis Nicola S 
Published: August 2019

Abstract
Wild ducks and gulls are the major reservoirs for avian influenza A viruses (AIVs). The mechanisms that drive AIV evolution are complex at sites where various duck and gull species from multiple flyways breed, winter, or stage. The Republic of Georgia is located at the intersection of three migratory flyways: the Central Asian Flyway, East Asian/East African Flyway, and Black Sea/Mediterranean Flyway. For six consecutive years (2010–6), we collected AIV samples from various duck and gull species that breed, migrate, and overwinter in Georgia. We found substantial subtype diversity of viruses that varied in prevalence from year to year. Low pathogenic (LP)AIV subtypes included H1N1, H2N3, H2N5, H2N7, H3N8, H4N2, H6N2, H7N3, H7N7, H9N1, H9N3, H10N4, H10N7, H11N1, H13N2, H13N6, H13N8, and H16N3, plus two H5N5 and H5N8 highly pathogenic (HP)AIVs belonging to clade 2.3.4.4. Whole-genome phylogenetic trees showed significant host species lineage restriction for nearly all gene segments and significant differences for LPAIVs among different host species in observed reassortment rates, as defined by quantification of phylogenetic incongruence, and in nucleotide diversity. Hemagglutinin clade 2.3.4.4 H5N8 viruses, circulated in Eurasia during 2014–5 did not reassort, but analysis after its subsequent dissemination during 2016–7 revealed reassortment in all gene segments except NP and NS. Some virus lineages appeared to be unrelated to AIVs in wild bird populations in other regions with maintenance of local AIV viruses in Georgia, whereas other lineages showed considerable genetic inter-relationship with viruses circulating in other parts of Eurasia and Africa, despite relative under-sampling in the area.
A34 Molecular characterization of Zika virus in Cuba
Authors: Martínez P A , Díaz G , Castillo E M , Álvarez M , Santana E , Serrano S , Mederos D , Guzmán M G 
Published: August 2019

Abstract
Until now, three genotypes of Zika virus (ZIKV) have been detected (two African lineages and one Asian lineage). After the declaration of Public Health Emergency of International Concern issued by The Pan American Health Organization and the World Health Organization authors from some Latin American countries have identified the Asian genotype as the lineage responsible for the Zika epidemic in the western hemisphere. However, data from the Caribbean are sparse, and there is no published data regarding the genotypes that produced isolated outbreaks in Cuba. Aiming to realize the molecular characterization of ZIKV in Cuba, we will sequence by next-generation sequencing the full genome of the ZIKV identified in samples from Cuban patients of different provinces in which ZIKV produced outbreaks. All samples required for this study have been collected during the molecular surveillance of Arboviral diseases conducted at the National Reference Laboratory at Pedro Kourí Tropical Medicine Institute. Viral RNA will be purified from urine and serum samples collected from patients with confirmed ZIKV infection by real time PCR. Using evolutionary dynamics studies, we will map the spread of a virus or of particular variants in time and space in order to understand how frequently ZIKV has been introduced into Cuba. Moreover, we will evaluate the amino acid diversity of each ZIKV proteins. Further, we will evaluate the population dynamics of ZIKV in samples from patients with varying clinical outcomes. The results will allow us to characterize the ZIKV genome and its evolution into the Cuban population that would also have impact for vaccine development, diagnosis, and pathogenesis studies.
A43 Translational research: NGS metagenomics into clinical diagnostics
Authors: Schmitz D , Nooij S , Janssens T , Cremer J , Vennema H , Kroneman A , Koopmans M 
Published: August 2019

Abstract
As research next-generation sequencing (NGS) metagenomic pipelines transition to clinical diagnostics, the user-base changes from bioinformaticians to biologists, medical doctors, and lab-technicians. Besides the obvious need for benchmarking and assessment of diagnostic outcomes of the pipelines and tools, other focus points remain: reproducibility, data immutability, user-friendliness, portability/scalability, privacy, and a clear audit trail. We have a research metagenomics pipeline that takes raw fastq files and produces annotated contigs, but it is too complicated for non-bioinformaticians. Here, we present preliminary findings in adapting this pipeline for clinical diagnostics. We used information available on relevant fora (www.bioinfo-core.org) and experiences and publications from colleague bioinformaticians in other institutes (COMPARE, UBC, and LUMC). From this information, a robust and user-friendly storage and analysis workflow was designed for non-bioinformaticians in a clinical setting. Via Conda [https://conda.io] and Docker containers [http://www.docker.com], we made our disparate pipeline processes self-contained and reproducible. Furthermore, we moved all pipeline settings into a separate JSON file. After every analysis, the pipeline settings and virtual-environment recipes will be archived (immutably) under a persistent unique identifier. This allows long-term precise reproducibility. Likewise, after every run the raw data and final products will be automatically archived, complying with data retention laws/guidelines. All the disparate processes in the pipeline are parallelized and automated via Snakemake1 (i.e. end-users need no coding skills). In addition, interactive web-reports such as MultiQC [http://multiqc.info] and Krona2 are generated automatically. By combining Snakemake, Conda, and containers, our pipeline is highly portable and easily scaled up for outbreak situations, or scaled down to reduce costs. Since patient privacy is a concern, our pipeline automatically removes human genetic data. Moreover, all source code will be stored on an internal Gitlab server, and, combined with the archived data, ensures a clear audit trail. Nevertheless, challenges remain: (1) reproducible reference databases, e.g. being able to revert to an older version to reproduce old analyses. (2) A user-friendly GUI. (3) Connecting the pipeline and NGS data to in-house LIMS. (4) Efficient long-term storage, e.g. lossless compression algorithms. Nevertheless, this work represents a step forward in making user-friendly clinical diagnostic workflows.
A50 Whole-genome sequencing of African swine fever isolates from Sardinia
Authors: Torresi C , Granberg F , Bertolotti L , Oggiano A , Colitti B , Giammarioli M , De Mia G M 
Published: August 2019

Abstract
In order to assess the molecular epidemiology of African swine fever (ASF) in Sardinia, we analyzed a wide range of isolates from wild and domestic pigs over a 31-year period (1978–2009) by genotyping sequence data from the genes encoding the p54 and the p72 proteins and the CVR. On this basis, the analysis of the B602L gene revealed a minor difference, placing the Sardinian isolates into two clusters according to their temporal distribution. As an extension of this study, in order to achieve a higher level of discrimination, three further variable genome regions, namely p30, CD2v, and I73R/I329L, of a large number of isolates collected from outbreaks in the years 2002–14 have been investigated. Sequence analysis of the CD2v region revealed a temporal subdivision of the viruses into two subgroups. These data, together with those from the B602L gene analysis, demonstrated that the viruses circulating in Sardinia belong to p72/genotype I, but since 1990 have undergone minor genetic variations in respect to its ancestor, thus making it impossible to trace isolates, enabling a more accurate assessment of the origin of outbreaks, and extending knowledge of virus evolution. To solve this problem, we have sequenced and annotated the complete genome of nine ASF isolates collected in Sardinia between 1978 and 2012. This was achieved using sequence data determined by next-generation sequencing. The results showed a very high identity with range of nucleotide similarity among isolates of 99.5 per cent to 99.9 per cent. The ASF virus (ASFV) genomes were composed of terminal inverted repeats and conserved and non-conserved ORFs. Among the conserved ORFs, B385R, H339R, and O61R-p12 showed 100 per cent amino acid identity. The same was true for the hypervariable ORFs, with regard to X69R, DP96R, DP60R, EP153R, B407L, I10L, and L60L genes. The EP402R and B602L genes showed, as expected, an amino acid identity range of 98.5 per cent to 100 per cent and 91 per cent to 100 per cent, respectively. In addition, all of the isolates displayed variable intergenic sequences. As a whole, the results from our studies confirmed a remarkable genetic stability of the ASFV/p72 genotype I viruses circulating in Sardinia.
A51 Genetic variability of small ruminant lentiviruses in sheep and goats from single-species flocks from Poland
Authors: Olech Monika , Kuźmak Jacek 
Published: August 2019

Abstract
Previous phylogenetic analyses of small ruminant lentivirus (SRLV) sequences found in Poland revealed the circulation of subtype A1 in both sheep and goats, subtypes B1 in goats, and subtypes B2, A12, and A13 in sheep only. This study aimed to analyze the genetic nature of SRLV circulating in sheep and goats from single-species flocks. In order to analyze the degree of genetic variability, the fragments of gag and env genes of 24 SRLV strains were amplified by PCR, cloned into plasmid vectors, sequenced, and consensus sequences were aligned to each other and to reference sequences available from GenBank. Phylogenetic analysis was performed using the Geneious tree-builder tool, and phylogenetic trees were constructed using Mr Bayes (using the general time reversible substitution model) within Geneious Pro 5.3. Pairwise genetic distances were calculated in MEGA 6. Phylogenetic analysis revealed that the strains were highly heterogeneous and represented ovine strains belonging to subtypes A12 and B2 and caprine strains grouped in subtypes B1, B2, A1, and A12. In addition, two novel subtypes, A16 and A17, were found in goats. The mean pairwise genetic distances of gag and env sequences of both clusters were above 15 per cent nucleotide divergence when compared to all other subtypes within group A, which is a criterion required to distinguish a new subtype. Additionally, the existence of two separated clusters was confirmed by high bootstrap values. Co-infections with strains belonging to different subtypes within A and B groups were detected in one sheep and four goats originating from four flocks. Since the co-infection with more than one lentivirus genotype offers an opportunity for viral recombination, the possible recombination events were tested based on RDP analysis. For all co-infected animals, no evidence of recombination was found within the gag gene; however, env sequences showed some recombination patterns in three samples. In conclusion, we have demonstrated extended genetic variability of SRLV in sheep and goats from Poland with the existence of co-infection and recombination events.
A54 Genomic analysis of camel-HKU23 in Nigeria dromedary camels reveals strain-specific cross-species recombination
Authors: So R T Y , Oladipo J O , Chu D K W , Peiris M 
Published: August 2019

Abstract
Coronaviruses (CoVs) are enveloped, single stranded, positive-sense RNA viruses with a large genomic size of 26–32 kilobases. The first human CoV identified in the 1960s was isolated from patients presenting with common cold symptoms. Subsequent epidemic outbreaks of novel zoonotic CoV transmission were reported, examples including HCoV-229E (229E), HCoV-OC43 (OC43), severe acute respiratory syndrome, and Middle East respiratory syndrome (MERS). The ongoing outbreak of MERS in the Middle East is originating from a zoonotic source of dromedary camels. Surveillance later revealed that three CoV species—HCoV-229E (229E), camel-HKU23, and MERS-CoV—were co-circulating in Saudi Arabia dromedary camels. Camel-HKU23 belongs to Group 2a CoV, which also includes human coronavirus OC43, bovine coronavirus, and porcine hemagglutinating encephalomyelitis virus. Recombination, resulting in the generation of different novel genotypes, has been reported previously among these CoVs. Our surveillance of dromedary camels slaughtered in a major abattoir in Nigeria identified camel-HKU23 from nasal swab samples with a prevalence of 2.2 per cent. Phylogenetic analysis showed Nigeria camel-HKU23 is distinct from those previously identified in Saudi Arabia, while still genetically similar, as they share a monophyletic origin. Recombination analysis of Nigeria camel-HKU23 revealed two recombination breakpoints at positions of 22774–24100 base pairs (bp) and 28224–29362 bp. Recombination breakpoint at position 22774, encoding the Group 2a CoV-specific hemagglutinin esterase gene, exhibited high bootstrap support for clustering with RbCoV HKU14, which was previously detected in domestic rabbits in China. The recombination signal is only observed in Nigeria camel-HKU23, suggesting a regional varied evolutionary history of camel-HKU23. Our findings extended the knowledge of the evolutionary relationship among Group 2a CoVs. Further surveillance in other African camels will be important to elucidate the evolution of camel-HKU23.
A56 Visualization of recombination in deformed wing virus infecting bees
Authors: Bigot Diane , Gogol-Döring Andreas , Koch Peter , Paxton Robert J 
Published: August 2019

Abstract
Honey bees suffer increasing colony mortality worldwide, partially caused by the spread of viral pathogens. Among these pathogens, deformed wing virus (DWV) is one of the major, widespread viruses of honey bees resulting in wing deformities and weakening colonies. DWV can be found in honey bees, bumble bees, and other wild bees as three major genotypes named DWV-A, -B (also named Varroa destructor virus 1), and -C. Various recombinants of DWV-A and -B have been previously found in honey bees, some of which have been suggested to have higher virulence over non-recombinant, parental virus. In most of these cases, recombinants were only shown as consensus sequences from previous assemblies and alignments and may not reflect the biological reality of all variants present within a host bee. It is therefore important to build a method of recombinant detection and quantification within mixed infections in single-host individuals, including both parental and various recombinant genomes, so as to evaluate the relevance of recombinants for viral genome evolution and the impact on hosts. Here, we propose to visualize and quantify these recombinants using next-generation sequencing data to better understand how these genomes evolve within bees. Our method will be performed directly from raw sequence reads from various datasets (including field and lab experiments as well as screening of public databases) in order to obtain an overview of DWV recombination in various in vivo and in vitro conditions. Recombination of viral genomes is a key point for virus evolution. The detection and quantification of recombination will facilitate analysis of the determinants of recombination and help in understanding the routes by which new viral variants emerge. The emergence of new (more virulent) recombinant viruses can result from acquisition of new capabilities, such as escape from host immunity or increased transmission rates. Recombination can also lead to adaptation to new environments and new hosts by a change in cell tropism, allowing cross-species transmission, which may be particularly relevant for bumble bees and wild bees infected by honey bee-derived DWV.
A62 A major likelihood-based approach gives problematic estimates of diversification dynamics and rates
Authors: Di Chenlu , Meyer Andreas L S , Wiens John J 
Published: August 2019

Abstract
The diversity of life is shaped by rates of speciation and extinction, and so estimating these rates correctly is crucial for understanding diversity patterns among clades, regions, and habitats. In 2011, Morlon and collaborators developed a promising likelihood-based approach to estimate speciation and extinction and to infer the model describing how these rates change over time based on AICc. This approach is now implemented in an R package (RPANDA). Here, we test the accuracy of this approach under simulated conditions, to evaluate its ability to correctly estimate rates of speciation, extinction, and diversification (speciation—extinction) and to choose the correct underlying model of diversification (e.g. constant or changing rates of speciation and extinction over time). We found that this likelihood-based approach frequently picked the incorrect model. For example, with changing speciation rates over time, the correct model was chosen in only ∼10 per cent of replicates. There were significant relationships between true and estimated speciation rates using this approach, but relationships were weak when speciation rates were constant within clades. Relationships were consistently weak between true and estimated rates of extinction and of diversification. Overall, we suggest that results from this approach should be interpreted with considerable caution.