Influence of Sex on Respiratory Syncytial Virus Genotype Infection Frequency and Nasopharyngeal Microbiome.

Respiratory syncytial virus (RSV) has a significant health burden in children, older adults, and the immunocompromised. However, limited effort has been made to identify emergence of new RSV genotypes' frequency of infection and how the combination of nasopharyngeal microbiome and viral genotypes impact RSV disease outcomes. In an observational cohort designed to capture the first infant RSV infection, we employed multi-omics approaches to sequence 349 RSV complete genomes and matched nasopharyngeal microbiomes, during which the 2012/2013 season was dominated by RSV-A, whereas 2013 and 2014 was dominated by RSV-B. We found non-G-72nt-duplicated RSV-A strains were more frequent in male infants (P = 0.02), whereas G-72nt-duplicated genotypes (which is ON1 lineage) were seen equally in both males and females. DESeq2 testing of the nasal microbiome showed Haemophilus was significantly more abundant in infants with RSV-A infection compared to infants with RSV-B infection (adjusted P = 0.002). In addition, the broad microbial clustering of the abundant genera was significantly associated with infant sex (P = 0.03). Overall, we show sex differences in infection by RSV genotype and host nasopharyngeal microbiome, suggesting an interaction between host genetics, virus genotype, and associated nasopharyngeal microbiome. IMPORTANCE Respiratory syncytial virus (RSV) is one of the leading causes of lower respiratory tract infections in young children and is responsible for high hospitalization rates and morbidity in infants and the elderly. To understand how the emergence of RSV viral genotypes and viral-respiratory microbiome interactions contribute to infection frequency and severity, we utilized an observational cohort designed to capture the first infant RSV infection we employed multi-omics approaches to sequence 349 RSV complete genomes and matched nasopharyngeal microbiomes. We found non-G-72nt-duplicated RSV-A genotypes were more frequent in male infants, whereas G-72nt-duplicated RSV-A strains (ON1 lineage) were seen equally in both males and females. Microbiome analysis show Haemophilus was significantly more abundant in infants with RSV-A compared to infants with RSV-B infection and the microbial clustering of the abundant genera was associated with infant sex. Overall, we show sex differences in RSV genotype-nasopharyngeal microbiome, suggesting an interaction host genetics-virus-microbiome interaction.

Human Monoclonal Antibodies Potently Neutralize Zika Virus and Select for Escape Mutations on the Lateral Ridge of the Envelope Protein.

The mosquito-borne Zika virus (ZIKV) has been causing epidemic outbreaks on a global scale. Virus infection can result in severe disease in humans, including microcephaly in newborns and Guillain-Barré syndrome in adults. Here, we characterized monoclonal antibodies isolated from a patient with an active Zika virus infection that potently neutralized virus infection in Vero cells at the nanogram-per-milliliter range. In addition, these antibodies enhanced internalization of virions into human leukemia K562 cells in vitro, indicating their possible ability to cause antibody-dependent enhancement of disease. Escape variants of the ZIKV MR766 strain to a potently neutralizing antibody, AC10, exhibited an amino acid substitution at residue S368 in the lateral ridge region of the envelope protein. Analysis of publicly availably ZIKV sequences revealed the S368 site to be conserved among the vast majority (97.6%) of circulating strains. We validated the importance of this residue by engineering a recombinant virus with an S368R point mutation that was unable to be fully neutralized by AC10. Four out of the 12 monoclonal antibodies tested were also unable to neutralize the virus with the S368R mutation, suggesting this region to be an important immunogenic epitope during human infection. Last, a time-of-addition infection assay further validated the escape variant and showed that all monoclonal antibodies inhibited virus binding to the cell surface. Thus, the present study demonstrates that the lateral ridge region of the envelope protein is likely an immunodominant, neutralizing epitope.IMPORTANCE Zika virus (ZIKV) is a global health threat causing severe disease in humans, including microcephaly in newborns and Guillain-Barré syndrome in adults. Here, we analyzed the human monoclonal antibody response to acute ZIKV infection and found that neutralizing antibodies could not elicit Fc-mediated immune effector functions but could potentiate antibody-dependent enhancement of disease. We further identified critical epitopes involved with neutralization by generating and characterizing escape variants by whole-genome sequencing. We demonstrate that the lateral ridge region, particularly the S368 amino acid site, is critical for neutralization by domain III-specific antibodies.

Multiple Introductions and Antigenic Mismatch with Vaccines May Contribute to Increased Predominance of G12P[8] Rotaviruses in the United States.

Rotavirus is the leading global cause of diarrheal mortality for unvaccinated children under 5 years of age. The outer capsid of rotavirus virions consists of VP7 and VP4 proteins, which determine viral G and P types, respectively, and are primary targets of neutralizing antibodies. Successful vaccination depends upon generating broadly protective immune responses following exposure to rotaviruses presenting a limited number of G- and P-type antigens. Vaccine introduction resulted in decreased rotavirus disease burden but also coincided with the emergence of uncommon G and P genotypes, including G12. To gain insight into the recent predominance of G12P[8] rotaviruses in the United States, we evaluated 142 complete rotavirus genome sequences and metadata from 151 clinical specimens collected in Nashville, TN, from 2011 to 2013 through the New Vaccine Surveillance Network. Circulating G12P[8] strains were found to share many segments with other locally circulating strains but to have distinct constellations. Phylogenetic analyses of G12 sequences and their geographic sources provided evidence for multiple separate introductions of G12 segments into Nashville, TN. Antigenic epitopes of VP7 proteins of G12P[8] strains circulating in Nashville, TN, differ markedly from those of vaccine strains. Fully vaccinated children were found to be infected with G12P[8] strains more frequently than with other rotavirus genotypes. Multiple introductions and significant antigenic mismatch may in part explain the recent predominance of G12P[8] strains in the United States and emphasize the need for continued monitoring of rotavirus vaccine efficacy against emerging rotavirus genotypes.IMPORTANCE Rotavirus is an important cause of childhood diarrheal disease worldwide. Two immunodominant proteins of rotavirus, VP7 and VP4, determine G and P genotypes, respectively. Recently, G12P[8] rotaviruses have become increasingly predominant. By analyzing rotavirus genome sequences from stool specimens obtained in Nashville, TN, from 2011 to 2013 and globally circulating rotaviruses, we found evidence of multiple introductions of G12 genes into the area. Based on sequence polymorphisms, VP7 proteins of these viruses are predicted to present themselves to the immune system very differently than those of vaccine strains. Many of the sick children with G12P[8] rotavirus in their diarrheal stools also were fully vaccinated. Our findings emphasize the need for continued monitoring of circulating rotaviruses and the effectiveness of the vaccines against strains with emerging G and P genotypes.

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.

Large-Scale Complete-Genome Sequencing and Phylodynamic Analysis of Eastern Equine Encephalitis Virus Reveals Source-Sink Transmission Dynamics in the United States.

Eastern equine encephalitis virus (EEEV) has a high case-fatality rate in horses and humans, and Florida has been hypothesized to be the source of EEEV epidemics for the northeastern United States. To test this hypothesis, we sequenced complete genomes of 433 EEEV strains collected within the United States from 1934 to 2014. Phylogenetic analysis suggested EEEV evolves relatively slowly and that transmission is enzootic in Florida, characterized by higher genetic diversity and long-term local persistence. In contrast, EEEV strains in New York and Massachusetts were characterized by lower genetic diversity, multiple introductions, and shorter local persistence. Our phylogeographic analysis supported a source-sink model in which Florida is the major source of EEEV compared to the other localities sampled. In sum, this study revealed the complex epidemiological dynamics of EEEV in different geographic regions in the United States and provided general insights into the evolution and transmission of other avian mosquito-borne viruses in this region.IMPORTANCE Eastern equine encephalitis virus (EEEV) infections are severe in horses and humans on the east coast of the United States with a >90% mortality rate in horses, an ∼33% mortality rate in humans, and significant brain damage in most human survivors. However, little is known about the evolutionary characteristics of EEEV due to the lack of genome sequences. By generating large collection of publicly available complete genome sequences, this study comprehensively determined the evolution of the virus, described the epidemiological dynamics of EEEV in different states in the United States, and identified Florida as one of the major sources. These results may have important implications for the control and prevention of other mosquito-borne viruses in the Americas.

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

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.

Reassortment of Influenza A Viruses in Wild Birds in Alaska before H5 Clade 2.3.4.4 Outbreaks.

Sampling of mallards in Alaska during September 2014-April 2015 identified low pathogenic avian influenza A virus (subtypes H5N2 and H1N1) that shared ancestry with highly pathogenic reassortant H5N2 and H5N1 viruses. Molecular dating indicated reassortment soon after interhemispheric movement of H5N8 clade 2.3.4.4, suggesting genetic exchange in Alaska or surrounds before outbreaks.

Comprehensive Genome Scale Phylogenetic Study Provides New Insights on the Global Expansion of Chikungunya Virus.

Since the India and Indian Ocean outbreaks of 2005 and 2006, the global distribution of chikungunya virus (CHIKV) and the locations of epidemics have dramatically shifted. First, the Indian Ocean lineage (IOL) caused sustained epidemics in India and has radiated to many other countries. Second, the Asian lineage has caused frequent outbreaks in the Pacific islands and in 2013 was introduced into the Caribbean, followed by rapid spread to nearly all of the neotropics. Further, CHIKV epidemics, as well as exported cases, have been reported in central Africa after a long period of perceived silence. To understand these changes and to anticipate the future of the virus, the exact distribution, genetic diversity, transmission routes, and future epidemic potential of CHIKV require further assessment. To do so, we conducted the most comprehensive phylogenetic analysis to date, examined CHIKV evolution and transmission, and explored distinct genetic factors associated with the emergence of the East/Central/South African (ECSA) lineage, the IOL, and the Asian lineage. Our results reveal contrasting evolutionary patterns among the lineages, with growing genetic diversities observed in each, and suggest that CHIKV will continue to be a major public health threat with the potential for further emergence and spread. IMPORTANCE: Chikungunya fever is a reemerging infectious disease that is transmitted by Aedes mosquitoes and causes severe health and economic burdens in affected populations. Since the unprecedented Indian Ocean and Indian subcontinent outbreaks of 2005 and 2006, CHIKV has further expanded its geographic range, including to the Americas in 2013. Its evolution and transmission during and following these epidemics, as well as the recent evolution and spread of other lineages, require optimal assessment. Using newly obtained genome sequences, we provide a comprehensive update of the global distribution of CHIKV genetic diversity and analyze factors associated with recent outbreaks. These results provide a solid foundation for future evolutionary studies of CHIKV that can elucidate emergence mechanisms and also may help to predict future epidemics.

Molecular Evolution and Intraclade Recombination of Enterovirus D68 during the 2014 Outbreak in the United States.

UNLABELLED: In August 2014, an outbreak of enterovirus D68 (EV-D68) occurred in North America, causing severe respiratory disease in children. Due to a lack of complete genome sequence data, there is only a limited understanding of the molecular evolution and epidemiology of EV-D68 during this outbreak, and it is uncertain whether the differing clinical manifestations of EV-D68 infection are associated with specific viral lineages. We developed a high-throughput complete genome sequencing pipeline for EV-D68 that produced a total of 59 complete genomes from respiratory samples with a 95% success rate, including 57 genomes from Kansas City, MO, collected during the 2014 outbreak. With these data in hand, we performed phylogenetic analyses of complete genome and VP1 capsid protein sequences. Notably, we observed considerable genetic diversity among EV-D68 isolates in Kansas City, manifest as phylogenetically distinct lineages, indicative of multiple introductions of this virus into the city. In addition, we identified an intersubclade recombination event within EV-D68, the first recombinant in this virus reported to date. Finally, we found no significant association between EV-D68 genetic variation, either lineages or individual mutations, and a variety of demographic and clinical variables, suggesting that host factors likely play a major role in determining disease severity. Overall, our study revealed the complex pattern of viral evolution within a single geographic locality during a single outbreak, which has implications for the design of effective intervention and prevention strategies. IMPORTANCE: Until recently, EV-D68 was considered to be an uncommon human pathogen, associated with mild respiratory illness. However, in 2014 EV-D68 was responsible for more than 1,000 disease cases in North America, including severe respiratory illness in children and acute flaccid myelitis, raising concerns about its potential impact on public health. Despite the emergence of EV-D68, a lack of full-length genome sequences means that little is known about the molecular evolution of this virus within a single geographic locality during a single outbreak. Here, we doubled the number of publicly available complete genome sequences of EV-D68 by performing high-throughput next-generation sequencing, characterized the evolutionary history of this outbreak in detail, identified a recombination event, and investigated whether there was any correlation between the demographic and clinical characteristics of the patients and the viral variant that infected them. Overall, these results will help inform the design of intervention strategies for EV-D68.

Spread and persistence of influenza A viruses in waterfowl hosts in the North American Mississippi migratory flyway.

UNLABELLED: While geographic distance often restricts the spread of pathogens via hosts, this barrier may be compromised when host species are mobile. Migratory waterfowl in the order Anseriformes are important reservoir hosts for diverse populations of avian-origin influenza A viruses (AIVs) and are assumed to spread AIVs during their annual continental-scale migrations. However, support for this hypothesis is limited, and it is rarely tested using data from comprehensive surveillance efforts incorporating both the temporal and spatial aspects of host migratory patterns. We conducted intensive AIV surveillance of waterfowl using the North American Mississippi Migratory Flyway (MMF) over three autumn migratory seasons. Viral isolates (n = 297) from multiple host species were sequenced and analyzed for patterns of gene dispersal between northern staging and southern wintering locations. Using a phylogenetic and nucleotide identity framework, we observed a larger amount of gene dispersal within this flyway rather than between the other three longitudinally identified North American flyways. Across seasons, we observed patterns of regional persistence of diversity for each genomic segment, along with limited survival of dispersed AIV gene lineages. Reassortment increased with both time and distance, resulting in transient AIV constellations. This study shows that within the MMF, AIV gene flow favors spread along the migratory corridor within a season, and also that intensive surveillance during bird migration is important for identifying virus dispersal on time scales relevant to pandemic responsiveness. In addition, this study indicates that comprehensive monitoring programs to capture AIV diversity are critical for providing insight into AIV evolution and ecology in a major natural reservoir. IMPORTANCE: Migratory birds are a reservoir for antigenic and genetic diversity of influenza A viruses (AIVs) and are implicated in the spread of virus diversity that has contributed to previous pandemic events. Evidence for dispersal of avian-origin AIVs by migratory birds is rarely examined on temporal scales relevant to pandemic or panzootic threats. Therefore, characterizing AIV movement by hosts within a migratory season is important for implementing effective surveillance strategies. We conducted surveillance following birds along a major North American migratory route and observed that within a migratory season, AIVs rapidly reassorted and gene lineages were dispersed primarily within the migratory corridor. Patterns of regional persistence were observed across seasons for each gene segment. We show that dispersal of AIV gene lineages by migratory birds occurs quickly along migratory routes and that surveillance for AIVs threatening human and animal health should focus attention on these routes.

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.

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.

Isolation, propagation, genome analysis and epidemiology of HKU1 betacoronaviruses.

From 1 January 2009 to 31 May 2013, 15 287 respiratory specimens submitted to the Clinical Virology Laboratory at the Children's Hospital Colorado were tested for human coronavirus RNA by reverse transcription-PCR. Human coronaviruses HKU1, OC43, 229E and NL63 co-circulated during each of the respiratory seasons but with significant year-to-year variability, and cumulatively accounted for 7.4-15.6 % of all samples tested during the months of peak activity. A total of 79 (0.5 % prevalence) specimens were positive for human betacoronavirus HKU1 RNA. Genotypes HKU1 A and B were both isolated from clinical specimens and propagated on primary human tracheal-bronchial epithelial cells cultured at the air-liquid interface and were neutralized in vitro by human intravenous immunoglobulin and by polyclonal rabbit antibodies to the spike glycoprotein of HKU1. Phylogenetic analysis of the deduced amino acid sequences of seven full-length genomes of Colorado HKU1 viruses and the spike glycoproteins from four additional HKU1 viruses from Colorado and three from Brazil demonstrated remarkable conservation of these sequences with genotypes circulating in Hong Kong and France. Within genotype A, all but one of the Colorado HKU1 sequences formed a unique subclade defined by three amino acid substitutions (W197F, F613Y and S752F) in the spike glycoprotein and exhibited a unique signature in the acidic tandem repeat in the N-terminal region of the nsp3 subdomain. Elucidating the function of and mechanisms responsible for the formation of these varying tandem repeats will increase our understanding of the replication process and pathogenicity of HKU1 and potentially of other coronaviruses.

Genomic sequence of the pathogenic and allergenic filamentous fungus Aspergillus fumigatus.

Aspergillus fumigatus is exceptional among microorganisms in being both a primary and opportunistic pathogen as well as a major allergen. Its conidia production is prolific, and so human respiratory tract exposure is almost constant. A. fumigatus is isolated from human habitats and vegetable compost heaps. In immunocompromised individuals, the incidence of invasive infection can be as high as 50% and the mortality rate is often about 50% (ref. 2). The interaction of A. fumigatus and other airborne fungi with the immune system is increasingly linked to severe asthma and sinusitis. Although the burden of invasive disease caused by A. fumigatus is substantial, the basic biology of the organism is mostly obscure. Here we show the complete 29.4-megabase genome sequence of the clinical isolate Af293, which consists of eight chromosomes containing 9,926 predicted genes. Microarray analysis revealed temperature-dependent expression of distinct sets of genes, as well as 700 A. fumigatus genes not present or significantly diverged in the closely related sexual species Neosartorya fischeri, many of which may have roles in the pathogenicity phenotype. The Af293 genome sequence provides an unparalleled resource for the future understanding of this remarkable fungus.

Molecular characterization of respiratory syncytial viruses circulating in a paediatric cohort in Amman, Jordan.

Respiratory syncytial viruses (RSVs) are an important cause of mortality worldwide and a major cause of respiratory tract infections in children, driving development of vaccine candidates. However, there are large gaps in our knowledge of the local evolutionary and transmission dynamics of RSVs, particularly in understudied regions such as the Middle East. To address this gap, we sequenced the complete genomes of 58 RSVA and 27 RSVB samples collected in a paediatric cohort in Amman, Jordan, between 2010 and 2013. RSVA and RSVB co-circulated during each winter epidemic of RSV in Amman, and each epidemic comprised multiple independent viral introductions of RSVA and RSVB. However, RSVA and RSVB alternated in dominance across years, potential evidence of immunological interactions. Children infected with RSVA tended to be older than RSVB-infected children [30 months versus 22.4 months, respectively (P value = 0.02)], and tended to developed bronchopneumonia less frequently than those with RSVB, although the difference was not statistically significant (P value = 0.06). Differences in spatial patterns were investigated, and RSVA lineages were often identified in multiple regions in Amman, whereas RSVB introductions did not spread beyond a single region of the city, although these findings were based on small sample sizes. Multiple RSVA genotypes were identified in Amman, including GA2 viruses as well as three viruses from the ON1 sub-genotype that emerged in 2009 and are now the dominant genotype circulating worldwide. As vaccine development advances, further sequencing of RSV is needed to understand viral ecology and transmission, particularly in under-studied locations.

Genome sequence and identification of candidate vaccine antigens from the animal pathogen Dichelobacter nodosus.

Dichelobacter nodosus causes ovine footrot, a disease that leads to severe economic losses in the wool and meat industries. We sequenced its 1.4-Mb genome, the smallest known genome of an anaerobe. It differs markedly from small genomes of intracellular bacteria, retaining greater biosynthetic capabilities and lacking any evidence of extensive ongoing genome reduction. Comparative genomic microarray studies and bioinformatic analysis suggested that, despite its small size, almost 20% of the genome is derived from lateral gene transfer. Most of these regions seem to be associated with virulence. Metabolic reconstruction indicated unsuspected capabilities, including carbohydrate utilization, electron transfer and several aerobic pathways. Global transcriptional profiling and bioinformatic analysis enabled the prediction of virulence factors and cell surface proteins. Screening of these proteins against ovine antisera identified eight immunogenic proteins that are candidate antigens for a cross-protective vaccine.

Erratum: Evolutionary dynamics and molecular epidemiology of West Nile virus in New York State: 1999-2015.

[This corrects the article DOI: 10.1093/ve/vez020.].

Evolutionary dynamics and molecular epidemiology of West Nile virus in New York State: 1999-2015.

Following its introduction into New York State (NYS) in 1999, West Nile virus (WNV; Flavivirus, Flaviviridae) underwent a rapid expansion throughout the USA and into Canada and Latin America. WNV has been characterized as being evolutionarily stable, with weak geographic structure, a dominance of purifying selection and limited adaptive change. We analyzed all available full-genome WNV sequences, focusing on the 543 available sequences from NYS, which included 495 newly sequenced 2000-15 isolates. In addition, we analyzed deep-sequencing data from 317 of these isolates. While our data are generally in agreement with the limited pace of evolutionary change and broad geographic and temporal mixing identified in other studies, we have identified some important exceptions. Most notably, there are 14 codons which demonstrated evidence of positive selection as determined by multiple models, including some positions with evidence of selection in NYS exclusively. Coincident with increased WNV activity, genotypes possessing one or more of these mutations, designated NY01, NY07, and NY10, have increased in prevalence in recent years and displaced historic strains. In addition, we have found a geographical bias with many of these mutations, which suggests selective pressures and adaptations could be regional. Lastly, our deep-sequencing data suggest both increased overall diversity in avian tissue isolates relative to mosquito isolates and multiple non-synonymous minority variants that are both host-specific and retained over time and space. Together, these data provide novel insight into the evolutionary pressures on WNV and the need for continued genetic surveillance and characterization of emergent strains.

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.