Low-dose dengue virus 3 human challenge model: a phase 1 open-label study.

Dengue human infection models present an opportunity to explore the potential of a vaccine, anti-viral or immuno-compound for clinical benefit in a controlled setting. Here we report the outcome of a phase 1 open-label assessment of a low-dose dengue virus 3 (DENV-3) challenge model (NCT04298138), in which nine participants received a subcutaneous inoculation with 0.5 ml of a 1.4 × 103 plaque-forming unit per ml suspension of the attenuated DENV-3 strain CH53489. The primary and secondary endpoints of the study were to assess the safety of this DENV-3 strain in healthy flavivirus-seronegative individuals. All participants developed RNAaemia within 7 days after inoculation with peak titre ranging from 3.13 × 104 to 7.02 × 108 genome equivalents per ml. Solicited symptoms such as fever and rash, clinical laboratory abnormalities such as lymphopenia and thrombocytopenia, and self-reported symptoms such as myalgia were consistent with mild-to-moderate dengue in all volunteers. DENV-3-specific seroconversion and memory T cell responses were observed within 14 days after inoculation as assessed by enzyme-linked immunosorbent assay and interferon-gamma-based enzyme-linked immunospot. RNA sequencing and serum cytokine analysis revealed anti-viral responses that overlapped with the period of viraemia. The magnitude and frequency of clinical and immunologic endpoints correlated with an individual's peak viral titre.

Zika-specific neutralizing antibodies targeting inter-dimer envelope epitopes.

Zika virus (ZIKV) is an emerging pathogen that causes devastating congenital defects. The overlapping epidemiology and immunologic cross-reactivity between ZIKV and dengue virus (DENV) pose complex challenges to vaccine design, given the potential for antibody-dependent enhancement of disease. Therefore, classification of ZIKV-specific antibody targets is of notable value. From a ZIKV-infected rhesus macaque, we identify ZIKV-reactive B cells and isolate potent neutralizing monoclonal antibodies (mAbs) with no cross-reactivity to DENV. We group these mAbs into four distinct antigenic groups targeting ZIKV-specific cross-protomer epitopes on the envelope glycoprotein. Co-crystal structures of representative mAbs in complex with ZIKV envelope glycoprotein reveal envelope-dimer epitope and unique dimer-dimer epitope targeting. All four specificities are serologically identified in convalescent humans following ZIKV infection, and representative mAbs from all four groups protect against ZIKV replication in mice. These results provide key insights into ZIKV-specific antigenicity and have implications for ZIKV vaccine, diagnostic, and therapeutic development.

Safety and immunogenicity of a purified inactivated Zika virus vaccine candidate in adults primed with a Japanese encephalitis virus or yellow fever virus vaccine in the USA: a phase 1, randomised, double-blind, placebo-controlled clinical trial.

BACKGROUND: Zika virus infection is a threat to at-risk populations, causing major birth defects and serious neurological complications. Development of a safe and efficacious Zika virus vaccine is, therefore, a global health priority. Assessment of heterologous flavivirus vaccination is important given co-circulation of Japanese encephalitis virus and yellow fever virus with Zika virus. We investigated the effect of priming flavivirus naive participants with a licensed flavivirus vaccine on the safety and immunogenicity of a purified inactivated Zika vaccine (ZPIV). METHODS: This phase 1, placebo-controlled, double-blind trial was done at the Walter Reed Army Institute of Research Clinical Trials Center in Silver Spring, MD, USA. Eligible participants were healthy adults aged 18-49 years, with no detectable evidence of previous flavivirus exposure (by infection or vaccination), as measured by a microneutralisation assay. Individuals with serological evidence of HIV, hepatitis B, or hepatitis C infection were excluded, as were pregnant or breastfeeding women. Participants were recruited sequentially into one of three groups (1:1:1) to receive no primer, two doses of intramuscular Japanese encephalitis virus vaccine (IXIARO), or a single dose of subcutaneous yellow fever virus vaccine (YF-VAX). Within each group, participants were randomly assigned (4:1) to receive intramuscular ZPIV or placebo. Priming vaccinations were given 72-96 days before ZPIV. ZPIV was administered either two or three times, at days 0, 28, and 196-234. The primary outcome was occurrence of solicited systemic and local adverse events along with serious adverse events and adverse events of special interest. These data were analysed in all participants receiving at least one dose of ZPIV or placebo. Secondary outcomes included measurement of neutralizing antibody responses following ZPIV vaccination in all volunteers with available post-vaccination data. This trial is registered at ClinicalTrials.gov, NCT02963909. FINDINGS: Between Nov 7, 2016, and Oct 30, 2018, 134 participants were assessed for eligibility. 21 did not meet inclusion criteria, 29 met exclusion criteria, and ten declined to participate. 75 participants were recruited and randomly assigned. 35 (47%) of 75 participants were male and 40 (53%) were female. 25 (33%) of 75 participants identified as Black or African American and 42 (56%) identified as White. These proportions and other baseline characteristics were similar between groups. There were no statistically significant differences in age, gender, race, or BMI between those who did and did not opt into the third dose. All participants received the planned priming IXIARO and YF-VAX vaccinations, but one participant who received YF-VAX dropped out before receipt of the first dose of ZPIV. 50 participants received a third dose of ZPIV or placebo, including 14 flavivirus-naive people, 17 people primed with Japanese encephalitis virus vaccine, and 19 participants primed with yellow fever vaccine. Vaccinations were well tolerated across groups. Pain at the injection site was the only adverse event reported more frequently in participants who received ZPIV than in those who received placebo (39 [65%] of 60 participants, 95% CI 51·6-76·9 who received ZPIV vs three [21·4%] of 14 who received placebo; 4·7-50·8; p=0·006). No patients had an adverse event of special interest or serious adverse event related to study treatment. At day 57, the flavivirus-naive volunteers had an 88% (63·6-98·5, 15 of 17) seroconversion rate (neutralising antibody titre ≥1:10) and geometric mean neutralising antibody titre (GMT) against Zika virus of 100·8 (39·7-255·7). In the Japanese encephalitis vaccine-primed group, the day 57 seroconversion rate was 31·6% (95% CI 12·6-56·6, six of 19) and GMT was 11·8 (6·1-22·8). Participants primed with YF-VAX had a seroconversion rate of 25% (95% CI 8·7-49·1, five of 20) and GMT of 6·6 (5·2-8·4). Humoral immune responses rose substantially following a third dose of ZPIV, with seroconversion rates of 100% (69·2-100; ten of ten), 92·9% (66·1-99·8; 13 of 14), and 60% (32·2-83·7, nine of 15) and GMTs of 511·5 (177·6-1473·6), 174·2 (51·6-587·6), and 79 (19·0-326·8) in the flavivirus naive, Japanese encephalitis vaccine-primed, and yellow fever vaccine-primed groups, respectively. INTERPRETATION: We found ZPIV to be well tolerated in flavivirus naive and primed adults but that immunogenicity varied significantly according to antecedent flavivirus vaccination status. Immune bias towards the flavivirus antigen of initial exposure and the timing of vaccination may have impacted responses. A third ZPIV dose overcame much, but not all, of the discrepancy in immunogenicity. The results of this phase 1 clinical trial have implications for further evaluation of ZPIV's immunisation schedule and use of concomitant vaccinations. FUNDING: Department of Defense, Defense Health Agency; National Institute of Allergy and Infectious Diseases; and Division of Microbiology and Infectious Disease.

Blocking NS3-NS4B interaction inhibits dengue virus in non-human primates.

Dengue is a major health threat and the number of symptomatic infections caused by the four dengue serotypes is estimated to be 96 million1 with annually around 10,000 deaths2. However, no antiviral drugs are available for the treatment or prophylaxis of dengue. We recently described the interaction between non-structural proteins NS3 and NS4B as a promising target for the development of pan-serotype dengue virus (DENV) inhibitors3. Here we present JNJ-1802-a highly potent DENV inhibitor that blocks the NS3-NS4B interaction within the viral replication complex. JNJ-1802 exerts picomolar to low nanomolar in vitro antiviral activity, a high barrier to resistance and potent in vivo efficacy in mice against infection with any of the four DENV serotypes. Finally, we demonstrate that the small-molecule inhibitor JNJ-1802 is highly effective against viral infection with DENV-1 or DENV-2 in non-human primates. JNJ-1802 has successfully completed a phase I first-in-human clinical study in healthy volunteers and was found to be safe and well tolerated4. These findings support the further clinical development of JNJ-1802, a first-in-class antiviral agent against dengue, which is now progressing in clinical studies for the prevention and treatment of dengue.

Characterization of B-cell and T-cell responses to a tetravalent dengue purified inactivated vaccine in healthy adults.

The increasing global impact of dengue underscores the need for a dengue virus (DENV) vaccine. We assessed B-cell and T-cell responses following vaccination with four formulations of a tetravalent dengue purified inactivated vaccine (DPIV) in dengue-primed and dengue-naive adults from two studies (NCT01666652, NCT01702857). Frequencies of DPIV-induced memory B cells specific to each DENV serotype remained high up to 12 months post-vaccination, and were higher in the dengue-primed than dengue-naive adults. A subsequent DPIV booster dose induced strong anamnestic B-cell responses. Multifunctional CD4+ T cells (predominantly expressing IL-2) were induced by DPIV, with higher frequencies in dengue-primed adults. DPIV-induced CD4+ T cells also demonstrated in vitro proliferative capacity and antigen-specific production of GM-CSF, IFN-γ, and IL-13. CD8+ T-cell responses were undetectable in dengue-naive adults and low in dengue-primed individuals. B- and T-cell responses persisted up to 12 months post-vaccination in both dengue-primed and dengue-naive adults.

The seroepidemiology of dengue in a US military population based in Puerto Rico during the early phase of the Zika pandemic.

Understanding the burden and risk factors of dengue virus (DENV) infection in Puerto Rico is important for the prevention of dengue in local, traveler and military populations. Using sera from the Department of Defense Serum Repository, we estimated the prevalence and predictors of DENV seropositivity in those who had served in Puerto Rico, stratified by birth or prior residence ("birth/residence") in dengue-endemic versus non-endemic regions. We selected sera collected in early 2015 from 500 U.S. military members, a time-point also permitting detection of early cryptic Zika virus (ZIKV) circulation. 87.2% were born or resided in a DENV-endemic area before their military service in Puerto Rico. A high-throughput, flow-cytometry-based neutralization assay was employed to screen sera for ZIKV and DENV neutralizing antibodies, and confirmatory testing was done by plaque-reduction neutralization test (PRNT). We identified one Puerto Rico resident who seroconverted to ZIKV by June 2015, suggesting cryptic ZIKV circulation in Puerto Rico at least 4 months before the first reported cases. A further six PRNT-positive presumptive ZIKV infections which were resolved as DENV infections only by the use of paired sera. We noted 66.8% of the total study sample was DENV seropositive by early 2015. Logistic regression analysis indicated that birth/residence in a dengue non-endemic region (before military service in Puerto Rico) was associated with a lower odds of DENV exposure by January-June 2015 (aOR = 0.28, p = 0.001). Among those with birth/residence in a non-endemic country, we noted moderate evidence to support increase in odds of DENV exposure for each year of military service in Puerto Rico (aOR = 1.58, p = 0.06), but no association with age. In those with birth/residence in dengue-endemic regions (before military service in Puerto Rico), we noted that age (aOR = 1.04, p = 0.02), rather than duration of Puerto Rico service, was associated with dengue seropositivity, suggesting earlier lifetime DENV exposure. Our findings provide insights into the burden and predictors of DENV infection in local, traveler and military populations in Puerto Rico. Our study also highlights substantial PRNT ZIKV false-positivity when paired sera are not available, even during periods of very low ZIKV prevalence.

Associations of human leukocyte antigen with neutralizing antibody titers in a tetravalent dengue vaccine phase 2 efficacy trial in Thailand.

The recombinant, live, attenuated, tetravalent dengue vaccine CYD-TDV has shown efficacy against all four dengue serotypes. In this exploratory study (CYD59, NCT02827162), we evaluated potential associations of host human leukocyte antigen (HLA) alleles with dengue antibody responses, CYD-TDV vaccine efficacy, and virologically-confirmed dengue (VCD) cases. Children 4-11 years old, who previously completed a phase 2b efficacy study of CYD-TDV in a single center in Thailand, were included in the study. Genotyping of HLA class I and II loci was performed by next-generation sequencing from DNA obtained from 335 saliva samples. Dengue neutralizing antibody titers (NAb) were assessed as a correlate of risk and protection. Regression analyses were used to assess associations between HLA alleles and NAb responses, vaccine efficacy, and dengue outcomes. Month 13 NAb log geometric mean titers (GMTs) were associated with decreased risk of VCD. In the vaccine group, HLA-DRB1*11 was significantly associated with higher NAb log GMT levels (beta: 0.76; p = 0.002, q = 0.13). Additionally, in the absence of vaccination, HLA associations were observed between the presence of DPB1*03:01 and increased NAb log GMT levels (beta: 1.24; p = 0.005, q = 0.17), and between DPB1*05:01 and reduced NAb log GMT levels (beta: -1.1; p = 0.001, q = 0.07). This study suggests associations of HLA alleles with NAb titers in the context of dengue outcomes. This study was registered with clinicaltrials.gov: NCT02827162.

Designed, highly expressing, thermostable dengue virus 2 envelope protein dimers elicit quaternary epitope antibodies.

Dengue virus (DENV) is a worldwide health burden, and a safe vaccine is needed. Neutralizing antibodies bind to quaternary epitopes on DENV envelope (E) protein homodimers. However, recombinantly expressed soluble E proteins are monomers under vaccination conditions and do not present these quaternary epitopes, partly explaining their limited success as vaccine antigens. Using molecular modeling, we found DENV2 E protein mutations that induce dimerization at low concentrations (<100 pM) and enhance production yield by more than 50-fold. Cross-dimer epitope antibodies bind to the stabilized dimers, and a crystal structure resembles the wild-type (WT) E protein bound to a dimer epitope antibody. Mice immunized with the stabilized dimers developed antibodies that bind to E dimers and not monomers and elicited higher levels of DENV2-neutralizing antibodies compared to mice immunized with WT E antigen. Our findings demonstrate the feasibility of using structure-based design to produce subunit vaccines for dengue and other flaviviruses.

A comparative recombination analysis of human coronaviruses and implications for the SARS-CoV-2 pandemic.

The SARS-CoV-2 pandemic prompts evaluation of recombination in human coronavirus (hCoV) evolution. We undertook recombination analyses of 158,118 public seasonal hCoV, SARS-CoV-1, SARS-CoV-2 and MERS-CoV genome sequences using the RDP4 software. We found moderate evidence for 8 SARS-CoV-2 recombination events, two of which involved the spike gene, and low evidence for one SARS-CoV-1 recombination event. Within MERS-CoV, 229E, OC43, NL63 and HKU1 datasets, we noted 7, 1, 9, 14, and 1 high-confidence recombination events, respectively. There was propensity for recombination breakpoints in the non-ORF1 region of the genome containing structural genes, and recombination severely skewed the temporal structure of these data, especially for NL63 and OC43. Bayesian time-scaled analyses on recombinant-free data indicated the sampled diversity of seasonal CoVs emerged in the last 70 years, with 229E displaying continuous lineage replacements. These findings emphasize the importance of genomic based surveillance to detect recombination in SARS-CoV-2, particularly if recombination may lead to immune evasion.

Metagenomic analysis reveals Culex mosquito virome diversity and Japanese encephalitis genotype V in the Republic of Korea.

Recent outbreaks of emerging and re-emerging viruses have shown that timely detection of novel arboviruses with epidemic potential is essential to mitigate human health risks. There are rising concerns that emergent JEV genotype V (GV) is circulating in Asia, against which current vaccines may not be efficacious. To ascertain if JEV GV and other arboviruses are circulating in East Asia, we conducted next-generation sequencing on 260 pools of Culex tritaeniorhynchus and Culex bitaeniorhynchus mosquitoes (6540 specimens) collected at Camp Humphreys, Republic of Korea (ROK) in 2018. Interrogation of our data revealed a highly abundant and diverse virosphere that contained sequences from 122 distinct virus species. Our statistical and hierarchical analysis uncovered correlates of potential health, virological, and ecological relevance. Furthermore, we obtained evidence that JEV GV was circulating in Pyeongtaek and, retrospectively, in Seoul in 2016 and placed these findings within the context of human and fowl reservoir activity. Sequence-based analysis of JEV GV showed a divergent genotype that is the most distant from the GIII-derived live attenuated SA14-14-2 vaccine strain and indicated regions probably responsible for reduced antibody affinity. These results emphasize recent concerns of shifting JEV genotype in East Asia and highlight the critical need for a vaccine proven efficacious against this re-emergent virus. Together, our one-health approach to Culex viral metagenomics uncovered novel insights into virus ecology and human health.

Evaluation of the extended efficacy of the Dengvaxia vaccine against symptomatic and subclinical dengue infection.

More than half of the world's population lives in areas at risk for dengue virus infection. A vaccine will be pivotal to controlling spread, however, the only licensed vaccine, Dengvaxia, has been shown to increase the risk of severe disease in a subset of individuals. Vaccine efforts are hampered by a poor understanding of antibody responses, including those generated by vaccines, and whether antibody titers can be used as a marker of protection from infection or disease. Here we present the results of an ancillary study to a phase III vaccine study (n = 611). All participants received three doses of either Dengvaxia or placebo and were followed for 6 years. We performed neutralization tests on annual samples and during confirmed dengue episodes (n = 16,508 total measurements). We use mathematical models to reconstruct long-term antibody responses to vaccination and natural infection, and to identify subclinical infections. There were 87 symptomatic infections reported, and we estimated that there were a further 351 subclinical infections. Cumulative vaccine efficacy was positive for both subclinical and symptomatic infection, although the protective effect of the vaccine was concentrated in the first 3 years following vaccination. Among individuals with the same antibody titer, we found no difference between the risk of subsequent infection or disease between placebo and vaccine recipients, suggesting that antibody titers are a good predictor of both protection and disease risk.

Correlation between reported dengue illness history and seropositivity in rural Thailand.

In the latest World Health Organization (WHO) recommendation for Dengvaxia implementation, either serological testing or a person's history of prior dengue illness may be used as supporting evidence to identify dengue virus (DENV)-immune individuals eligible for vaccination, in areas with limited capacity for laboratory confirmation. This analysis aimed to estimate the concordance between self-reported dengue illness histories and seropositivity in a prospective cohort study for dengue virus infection in Kamphaeng Phet province, a dengue-endemic area in northern Thailand. The study enrolled 2,076 subjects from 516 multigenerational families, with a median age of 30.6 years (range 0-90 years). Individual and family member dengue illness histories were obtained by questionnaire. Seropositivity was defined based on hemagglutination inhibition (HAI) assays. Overall seropositivity for DENV was 86.5% among those aged 9-45 years, which increased with age. 18.5% of participants reported a history of dengue illness prior to enrollment; 30.1% reported a previous DENV infection in the family, and 40.1% reported DENV infection in either themselves or a family member. Relative to seropositivity by HAI in the vaccine candidate group, the sensitivity and specificity of individual prior dengue illness history were 18.5% and 81.6%, respectively; sensitivity and specificity of reported dengue illness in a family member were 29.8% and 68.0%, and of either the individual or a family member were 40.1% and 60.5%. Notably, 13.4% of individuals reporting prior dengue illness were seronegative. Given the high occurrence of asymptomatic and mild DENV infection, self-reported dengue illness history is poorly sensitive for prior exposure and may misclassify individuals as 'exposed' when they were not. This analysis highlights that a simple, highly sensitive, and highly specific test for determining serostatus prior to Dengvaxia vaccination is urgently needed.

Precision Tracing of Household Dengue Spread Using Inter- and Intra-Host Viral Variation Data, Kamphaeng Phet, Thailand.

Dengue control approaches are best informed by granular spatial epidemiology of these viruses, yet reconstruction of inter- and intra-household transmissions is limited when analyzing case count, serologic, or genomic consensus sequence data. To determine viral spread on a finer spatial scale, we extended phylogenomic discrete trait analyses to reconstructions of house-to-house transmissions within a prospective cluster study in Kamphaeng Phet, Thailand. For additional resolution and transmission confirmation, we mapped dengue intra-host single nucleotide variants on the taxa of these time-scaled phylogenies. This approach confirmed 19 household transmissions and revealed that dengue disperses an average of 70 m per day between households in these communities. We describe an evolutionary biology framework for the resolution of dengue transmissions that cannot be differentiated based on epidemiologic and consensus genome data alone. This framework can be used as a public health tool to inform control approaches and enable precise tracing of dengue transmissions.

Enhanced dengue vaccine virus replication and neutralizing antibody responses in immune primed rhesus macaques.

Antibody-dependent enhancement (ADE) is suspected to influence dengue virus (DENV) infection, but the role ADE plays in vaccination strategies incorporating live attenuated virus components is less clear. Using a heterologous prime-boost strategy in rhesus macaques, we examine the effect of priming with DENV purified inactivated vaccines (PIVs) on a tetravalent live attenuated vaccine (LAV). Sera exhibited low-level neutralizing antibodies (NAb) post PIV priming, yet moderate to high in vitro ADE activity. Following LAV administration, the PIV primed groups exhibited DENV-2 LAV peak viremias up to 1,176-fold higher than the mock primed group, and peak viremia correlated with in vitro ADE. Furthermore, PIV primed groups had more balanced and higher DENV-1-4 NAb seroconversion and titers than the mock primed group following LAV administration. These results have implications for the development of effective DENV vaccine prime-boost strategies and for our understanding of the role played by ADE in modulating DENV replication.

Pre-existing Immunity to Japanese Encephalitis Virus Alters CD4 T Cell Responses to Zika Virus Inactivated Vaccine.

The epidemic spread of Zika virus (ZIKV), associated with devastating neurologic syndromes, has driven the development of multiple ZIKV vaccines candidates. An effective vaccine should induce ZIKV-specific T cell responses, which are shown to improve the establishment of humoral immunity and contribute to viral clearance. Here we investigated how previous immunization against Japanese encephalitis virus (JEV) and yellow fever virus (YFV) influences T cell responses elicited by a Zika purified-inactivated virus (ZPIV) vaccine. We demonstrate that three doses of ZPIV vaccine elicited robust CD4 T cell responses to ZIKV structural proteins, while ZIKV-specific CD4 T cells in pre-immunized individuals with JEV vaccine, but not YFV vaccine, were more durable and directed predominantly toward conserved epitopes, which elicited Th1 and Th2 cytokine production. In addition, T cell receptor repertoire analysis revealed preferential expansion of cross-reactive clonotypes between JEV and ZIKV, suggesting that pre-existing immunity against JEV may prime the establishment of stronger CD4 T cell responses to ZPIV vaccination. These CD4 T cell responses correlated with titers of ZIKV-neutralizing antibodies in the JEV pre-vaccinated group, but not in flavivirus-naïve or YFV pre-vaccinated individuals, suggesting a stronger contribution of CD4 T cells in the generation of neutralizing antibodies in the context of JEV-ZIKV cross-reactivity.

Reconstructing unseen transmission events to infer dengue dynamics from viral sequences.

For most pathogens, transmission is driven by interactions between the behaviours of infectious individuals, the behaviours of the wider population, the local environment, and immunity. Phylogeographic approaches are currently unable to disentangle the relative effects of these competing factors. We develop a spatiotemporally structured phylogenetic framework that addresses these limitations by considering individual transmission events, reconstructed across spatial scales. We apply it to geocoded dengue virus sequences from Thailand (N = 726 over 18 years). We find infected individuals spend 96% of their time in their home community compared to 76% for the susceptible population (mainly children) and 42% for adults. Dynamic pockets of local immunity make transmission more likely in places with high heterotypic immunity and less likely where high homotypic immunity exists. Age-dependent mixing of individuals and vector distributions are not important in determining spread. This approach provides previously unknown insights into one of the most complex disease systems known and will be applicable to other pathogens.

Effect of low-passage number on dengue consensus genomes and intra-host variant frequencies.

Intra-host single nucleotide variants (iSNVs) have been increasingly used in genomic epidemiology to increase phylogenetic resolution and reconstruct fine-scale outbreak dynamics. These analyses are preferably done on sequence data from direct clinical samples, but in many cases due to low viral loads, there might not be enough genetic material for deep sequencing and iSNV determination. Isolation of the virus from clinical samples with low-passage number increases viral load, but few studies have investigated how dengue virus (DENV) culture isolation from a clinical sample impacts the consensus sequence and the intra-host virus population frequencies. In this study, we investigate consensus and iSNV frequency differences between DENV sequenced directly from clinical samples and their corresponding low-passage isolates. Twenty five DENV1 and DENV2 positive sera and their corresponding viral isolates (T. splendens inoculation and C6/36 passage) were obtained from a prospective cohort study in the Philippines. These were sequenced on MiSeq with minimum nucleotide depth of coverage of 500×, and iSNVs were detected using LoFreq. For both DENV1 and DENV2, we found a maximum of one consensus nucleotide difference between clinical sample and isolate. Interestingly, we found that iSNVs with frequencies ≥5 % were often preserved between the samples, and that the number of iSNV positions, and sample diversity, at this frequency cutoff did not differ significantly between the sample pairs (clinical sample and isolate) in either DENV1 or DENV2 data. Our results show that low-passage DENV isolate consensus genomes are largely representative of their direct sample parental viruses, and that low-passage isolates often mirror high frequency within-host variants from direct samples.

Recent African strains of Zika virus display higher transmissibility and fetal pathogenicity than Asian strains.

The global emergence of Zika virus (ZIKV) revealed the unprecedented ability for a mosquito-borne virus to cause congenital birth defects. A puzzling aspect of ZIKV emergence is that all human outbreaks and birth defects to date have been exclusively associated with the Asian ZIKV lineage, despite a growing body of laboratory evidence pointing towards higher transmissibility and pathogenicity of the African ZIKV lineage. Whether this apparent paradox reflects the use of relatively old African ZIKV strains in most laboratory studies is unclear. Here, we experimentally compare seven low-passage ZIKV strains representing the recently circulating viral genetic diversity. We find that recent African ZIKV strains display higher transmissibility in mosquitoes and higher lethality in both adult and fetal mice than their Asian counterparts. We emphasize the high epidemic potential of African ZIKV strains and suggest that they could more easily go unnoticed by public health surveillance systems than Asian strains due to their propensity to cause fetal loss rather than birth defects.

Complete Genome Sequences of Two Human Adenovirus Type 55 Isolates from South Korea and the United States.

Here, we report two complete genome sequences of human adenovirus 55 (HAdV-55) isolates, from a patient in Pennsylvania in 2006 and a U.S. military member in South Korea in 2019. The findings demonstrate the continued global transmission of HAdV-55 viruses in both military and civilian populations.

Temporally integrated single cell RNA sequencing analysis of PBMC from experimental and natural primary human DENV-1 infections.

Dengue human infection studies present an opportunity to address many longstanding questions in the field of flavivirus biology. However, limited data are available on how the immunological and transcriptional response elicited by an attenuated challenge virus compares to that associated with a wild-type DENV infection. To determine the kinetic transcriptional signature associated with experimental primary DENV-1 infection and to assess how closely this profile correlates with the transcriptional signature accompanying natural primary DENV-1 infection, we utilized scRNAseq to analyze PBMC from individuals enrolled in a DENV-1 human challenge study and from individuals experiencing a natural primary DENV-1 infection. While both experimental and natural primary DENV-1 infection resulted in overlapping patterns of inflammatory gene upregulation, natural primary DENV-1 infection was accompanied with a more pronounced suppression in gene products associated with protein translation and mitochondrial function, principally in monocytes. This suggests that the immune response elicited by experimental and natural primary DENV infection are similar, but that natural primary DENV-1 infection has a more pronounced impact on basic cellular processes to induce a multi-layered anti-viral state.