Influenza A virus evolution and spatio-temporal dynamics in Eurasian wild birds: a phylogenetic and phylogeographical study of whole-genome sequence data.

Low pathogenic avian influenza A viruses (IAVs) have a natural host reservoir in wild waterbirds and the potential to spread to other host species. Here, we investigated the evolutionary, spatial and temporal dynamics of avian IAVs in Eurasian wild birds. We used whole-genome sequences collected as part of an intensive long-term Eurasian wild bird surveillance study, and combined this genetic data with temporal and spatial information to explore the virus evolutionary dynamics. Frequent reassortment and co-circulating lineages were observed for all eight genomic RNA segments over time. There was no apparent species-specific effect on the diversity of the avian IAVs. There was a spatial and temporal relationship between the Eurasian sequences and significant viral migration of avian IAVs from West Eurasia towards Central Eurasia. The observed viral migration patterns differed between segments. Furthermore, we discuss the challenges faced when analysing these surveillance and sequence data, and the caveats to be borne in mind when drawing conclusions from the apparent results of such analyses.We thank all ornithologists and other collaborators for their continuous support. We thank V. Munster, E. Skepner, O. Vuong, C. Baas, J. Guldemeester, M. Schutten, G. van der Water, D. Smith and E. Bortz for technical support and stimulating discussions. This manuscript was prepared while D.E. Wentworth was employed at the JCVI. The opinions expressed in this article are the author’s own and do not reflect the view of the Centers for Disease Control, the Department of Health and Human Services, or the United States government. This work was supported by NIAID/NIH contract HHSN266200700010C, HHSN272201400008C, HHSN272201400006C and HHSN272200900007C, a Wellcome Trust Fellowship Strategic Travel Award under contract WT089235MF, a DTRA FRCWMD Broad Agency Announcement under contract HDTRA1-09-14-FRCWMD GRANT11177182, by the EU Framework six program NewFluBird (044490) by contracts with the Dutch Ministry of Economic Affairs and a NIAID/NIH CEIRS travel grant under contract HHSN266200700010C. The Swedish sampling and analysis was supported by the Swedish Research Councils VR and FORMAS.This is the final version of the article. It first appeared from the Society for General Microbiology via http://dx.doi.org/10.1099/vir.0.00015

Avian Influenza Viruses in Wild Birds: Virus Evolution in a Multihost Ecosystem.

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, the East Africa/West Asia flyway, and the Black Sea/Mediterranean flyway. For six complete study years (2010 to 2016), we collected AIV samples from various duck and gull species that breed, migrate, and overwinter in Georgia. We found a substantial subtype diversity of viruses that varied in prevalence from year to year. Low-pathogenic AIV (LPAIV) subtypes included H1N1, H2N3, H2N5, H2N7, H3N8, H4N2, H6N2, H7N3, H7N7, H9N1, H9N3, H10N4, H10N7, H11N1, H13N2, H13N6, H13N8, and H16N3, and two highly pathogenic AIVs (HPAIVs) belonging to clade 2.3.4.4, H5N5 and H5N8, were found. Whole-genome phylogenetic trees showed significant host species lineage restriction for nearly all gene segments and significant differences in observed reassortment rates, as defined by quantification of phylogenetic incongruence, and in nucleotide sequence diversity for LPAIVs among different host species. Hemagglutinin clade 2.3.4.4 H5N8 viruses, which circulated in Eurasia during 2014 and 2015, did not reassort, but analysis after their subsequent dissemination during 2016 and 2017 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 AIVs in Georgia, whereas other lineages showed considerable genetic interrelationships with viruses circulating in other parts of Eurasia and Africa, despite relative undersampling in the area.IMPORTANCE Waterbirds (e.g., gulls and ducks) are natural reservoirs of avian influenza viruses (AIVs) and have been shown to mediate the dispersal of AIVs at intercontinental scales during seasonal migration. The segmented genome of influenza viruses enables viral RNA from different lineages to mix or reassort when two viruses infect the same host. Such reassortant viruses have been identified in most major human influenza pandemics and several poultry outbreaks. Despite their importance, we have only recently begun to understand AIV evolution and reassortment in their natural host reservoirs. This comprehensive study illustrates AIV evolutionary dynamics within a multihost ecosystem at a stopover site where three major migratory flyways intersect. Our analysis of this ecosystem over a 6-year period provides a snapshot of how these viruses are linked to global AIV populations. Understanding the evolution of AIVs in the natural host is imperative to mitigating both the risk of incursion into domestic poultry and the potential risk to mammalian hosts, including humans

Sequencing and analysis of globally obtained human parainfluenza viruses 1 and 3 genomes

Human Parainfluenza viruses (HPIV) type 1 and 3 are important causes of respiratory tract infections in young children globally. HPIV infections do not confer complete protective immunity so reinfections occur throughout life. Since no effective vaccine is available for the two virus subtypes, comprehensive understanding of HPIV-1 and HPIV-3 genetic and epidemic features is important for diagnosis, prevention, and treatment of HPIV-1 and HPIV-3 infections. Relatively few whole genome sequences are available for both HPIV-1 and HPIV-3 viruses, so our study sought to provide whole genome sequences from multiple countries to further the understanding of the global diversity of HPIV at a whole-genome level. We collected HPIV-1 and HPIV-3 samples and isolates from Argentina, Australia, France, Mexico, South Africa, Switzerland, and USA from the years 2003–2011 and sequenced the genomes of 40 HPIV-1 and 75 HPIV-3 viruses with Sanger and next-generation sequencing with the Ion Torrent, Illumina, and 454 platforms. Phylogenetic analysis showed that the HPIV-1 genome is evolving at an estimated rate of 4.97 × 10−4 mutations/ site/year (95% highest posterior density 4.55 × 10−4 to 5.38 × 10−4) and the HPIV-3 genome is evolving at a similar rate (3.59 × 10−4 mutations/site/year, 95% highest posterior density 3.26 × 10−4 to 3.94 × 10−4). There were multiple genetically distinct lineages of both HPIV-1 and 3 circulating on a global scale. Further surveillance and whole-genome sequencing are greatly needed to better understand the spatial dynamics of these important respiratory viruses in humans.S1 Text. HPIV-1 Sanger sequencing primers.S2 Text. HPIV-3 Sanger sequencing primers.S1 Table. The sequence information of the 40 HPIV-1 genomes.S2 Table. The sequence information of the 75 HPIV-3 genomes.S3 Table. MEME episodic selection results for HPIV-1 and HPIV-3.The National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services under contract number HHSN272200900007C and grant numbers U19AI110819, with the sub-project directed by HAL, and grants U01AI070428 and U01AI077988 awarded to KJH.http://www.plosone.orgam2019Medical Virolog

Genomic analysis of a large set of currently—and historically—important human adenovirus pathogens

Human adenoviruses (HAdVs) are uniquely important “model organisms” as they have been used to elucidate fundamental biological processes, are recognized as complex pathogens, and are used as remedies for human health. As pathogens, HAdVs may effect asymptomatic or mild and severe symptomatic disease upon their infection of respiratory, ocular, gastrointestinal, and genitourinary systems. High-resolution genomic data have enhanced the understanding of HAdV epidemiology, with recombination recognized as an important and major pathway in the molecular evolution and genesis of emergent HAdV pathogens. To support this view and to actualize an algorithm for identifying, characterizing, and typing novel HAdVs, we determined the DNA sequence of 95 isolates from archives containing historically important pathogens and collections housing currently circulating strains to be sequenced. Of the 85 samples that were completely sequenced, 18 novel recombinants within species HAdV-B and D were identified. Two HAdV-D genomes were found to contain novel penton base and fiber genes with significant divergence from known molecular types. In this data set, we found additional isolates of HAdV-D53 and HAdV-D58, two novel genotypes recognized recently using genomics. This supports the thesis that novel HAdV genotypes are not limited to “one-time” appearances of the prototype but are of importance in HAdV epidemiology. These data underscore the significance of lateral genomic transfer in HAdV evolution and reinforce the potential public health impact of novel genotypes of HAdVs emerging in the population

Growth and adaptation of Zika virus in mammalian and mosquito cells.

The recent emergence of Zika virus (ZIKV) in the Americas coincident with increased caseloads of microcephalic infants and Guillain-Barre syndrome has prompted a flurry of research on ZIKV. Much of the research is difficult to compare or repeat because individual laboratories use different virus isolates, growth conditions, and quantitative assays. Here we obtained three readily available contemporary ZIKV isolates and the prototype Ugandan isolate. We generated stocks of each on Vero mammalian cells (ZIKVmam) and C6/36 mosquito cells (ZIKVmos), determined titers by different assays side-by-side, compared growth characteristics using one-step and multi-step growth curves on Vero and C6/36 cells, and examined plaque phenotype. ZIKV titers consistently peaked earlier on Vero cells than on C6/36 cells. Contemporary ZIKV isolates reached peak titer most quickly in a multi-step growth curve when the amplifying cell line was the same as the titering cell line (e.g., ZIKVmam titered on Vero cells). Growth of ZIKVmam on mosquito cells was particularly delayed. These data suggest that the ability to infect and/or replicate in insect cells is limited after growth in mammalian cells. In addition, ZIKVmos typically had smaller, more homogenous plaques than ZIKVmam in a standard plaque assay. We hypothesized that the plaque size difference represented early adaptation to growth in mammalian cells. We plaque purified representative-sized plaques from ZIKVmos and ZIKVmam. ZIKVmos isolates maintained the initial phenotype while plaques from ZIKVmam isolates became larger with passaging. Our results underscore the importance of the cells used to produce viral stocks and the potential for adaptation with minimal cell passages. In addition, these studies provide a foundation to compare current and emerging ZIKV isolates in vitro and in vivo

Whole genome sequencing, variant analysis, phylogenetics, and deep sequencing of Zika virus strains

Abstract The recent emergence of Zika virus (ZIKV) has been concentrated in the Caribbean, Southeastern United States, and South- and Central America; resulting in travel-based cases being reported around the globe. As multi-disciplinary collaborations are combatting the ZIKV outbreak, the need to validate the sequence of existing strains has become apparent. Here, we report high-quality sequence data for multiple ZIKV strains made publicly available through the National Institutes of Health- (NIH) funded biorepository, BEI Resources (www.beiresources.org). Next-generation sequencing, 3′ rapid amplification of cDNA ends (RACE), and viral genome annotation pipelines generated GenBank sequence records for 16 BEI Resources strains. Minor variants, consensus mutations, and consensus insertions/deletions were identified within the viral stocks using next-generation sequencing (NGS) and consensus changes were confirmed with Sanger sequencing. Bioinformatics analyses of the sequencing results confirm that the virus stocks available to the scientific research community through BEI Resources adequately represent the viral population diversity of ZIKV

Sequencing Bacillus anthracis Typing Phages Gamma and Cherry Reveals a Common Ancestry

The genetic relatedness of the Bacillus anthracis typing phages Gamma and Cherry was determined by nucleotide sequencing and comparative analysis. The genomes of these two phages were identical except at three variable loci, which showed heterogeneity within individual lysates and among Cherry, Wβ, Fah, and four Gamma bacteriophage sequences

Phylogeography of Influenza A(H3N2) Virus in Peru, 2010–2012

It remains unclear whether lineages of influenza A(H3N2) virus can persist in the tropics and seed temperate areas. We used viral gene sequence data sampled from Peru to test this source–sink model for a Latin American country. Viruses were obtained during 2010–2012 from influenza surveillance cohorts in Cusco, Tumbes, Puerto Maldonado, and Lima. Specimens positive for influenza A(H3N2) virus were randomly selected and underwent hemagglutinin sequencing and phylogeographic analyses. Analysis of 389 hemagglutinin sequences from Peru and 2,192 global sequences demonstrated interseasonal extinction of Peruvian lineages. Extensive mixing occurred with global clades, but some spatial structure was observed at all sites; this structure was weakest in Lima and Puerto Maldonado, indicating that these locations may experience greater viral traffic. The broad diversity and co-circulation of many simultaneous lineages of H3N2 virus in Peru suggests that this country should not be overlooked as a potential source for novel pandemic strains

Zika Virus Antagonizes Type I Interferon Responses during Infection of Human Dendritic Cells

<div><p>Zika virus (ZIKV) is an emerging mosquito-borne flavivirus that is causally linked to severe neonatal birth defects, including microcephaly, and is associated with Guillain-Barre syndrome in adults. Dendritic cells (DCs) are an important cell type during infection by multiple mosquito-borne flaviviruses, including dengue virus, West Nile virus, Japanese encephalitis virus, and yellow fever virus. Despite this, the interplay between ZIKV and DCs remains poorly defined. Here, we found human DCs supported productive infection by a contemporary Puerto Rican isolate with considerable variability in viral replication, but not viral binding, between DCs from different donors. Historic isolates from Africa and Asia also infected DCs with distinct viral replication kinetics between strains. African lineage viruses displayed more rapid replication kinetics and infection magnitude as compared to Asian lineage viruses, and uniquely induced cell death. Infection of DCs with both contemporary and historic ZIKV isolates led to minimal up-regulation of T cell co-stimulatory and MHC molecules, along with limited secretion of inflammatory cytokines. Inhibition of type I interferon (IFN) protein translation was observed during ZIKV infection, despite strong induction at the RNA transcript level and up-regulation of other host antiviral proteins. Treatment of human DCs with RIG-I agonist potently restricted ZIKV replication, while type I IFN had only modest effects. Mechanistically, we found all strains of ZIKV antagonized type I IFN-mediated phosphorylation of STAT1 and STAT2. Combined, our findings show that ZIKV subverts DC immunogenicity during infection, in part through evasion of type I IFN responses, but that the RLR signaling pathway is still capable of inducing an antiviral state, and therefore may serve as an antiviral therapeutic target.</p></div

Sequencing and analysis of globally obtained human parainfluenza viruses 1 and 3 genomes

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