Four principles to establish a universal virus taxonomy.

A universal taxonomy of viruses is essential for a comprehensive view of the virus world and for communicating the complicated evolutionary relationships among viruses. However, there are major differences in the conceptualisation and approaches to virus classification and nomenclature among virologists, clinicians, agronomists, and other interested parties. Here, we provide recommendations to guide the construction of a coherent and comprehensive virus taxonomy, based on expert scientific consensus. Firstly, assignments of viruses should be congruent with the best attainable reconstruction of their evolutionary histories, i.e., taxa should be monophyletic. This fundamental principle for classification of viruses is currently included in the International Committee on Taxonomy of Viruses (ICTV) code only for the rank of species. Secondly, phenotypic and ecological properties of viruses may inform, but not override, evolutionary relatedness in the placement of ranks. Thirdly, alternative classifications that consider phenotypic attributes, such as being vector-borne (e.g., "arboviruses"), infecting a certain type of host (e.g., "mycoviruses," "bacteriophages") or displaying specific pathogenicity (e.g., "human immunodeficiency viruses"), may serve important clinical and regulatory purposes but often create polyphyletic categories that do not reflect evolutionary relationships. Nevertheless, such classifications ought to be maintained if they serve the needs of specific communities or play a practical clinical or regulatory role. However, they should not be considered or called taxonomies. Finally, while an evolution-based framework enables viruses discovered by metagenomics to be incorporated into the ICTV taxonomy, there are essential requirements for quality control of the sequence data used for these assignments. Combined, these four principles will enable future development and expansion of virus taxonomy as the true evolutionary diversity of viruses becomes apparent.

The anti-immune dengue subgenomic flaviviral RNA is present in vesicles in mosquito saliva and is associated with increased infectivity.

Mosquito transmission of dengue viruses to humans starts with infection of skin resident cells at the biting site. There is great interest in identifying transmission-enhancing factors in mosquito saliva in order to counteract them. Here we report the discovery of high levels of the anti-immune subgenomic flaviviral RNA (sfRNA) in dengue virus 2-infected mosquito saliva. We established that sfRNA is present in saliva using three different methods: northern blot, RT-qPCR and RNA sequencing. We next show that salivary sfRNA is protected in detergent-sensitive compartments, likely extracellular vesicles. In support of this hypothesis, we visualized viral RNAs in vesicles in mosquito saliva and noted a marked enrichment of signal from 3'UTR sequences, which is consistent with the presence of sfRNA. Furthermore, we show that incubation with mosquito saliva containing higher sfRNA levels results in higher virus infectivity in a human hepatoma cell line and human primary dermal fibroblasts. Transfection of 3'UTR RNA prior to DENV2 infection inhibited type I and III interferon induction and signaling, and enhanced viral replication. Therefore, we posit that sfRNA present in salivary extracellular vesicles is delivered to cells at the biting site to inhibit innate immunity and enhance dengue virus transmission.

Recombinant vesicular stomatitis vaccine against Nipah virus has a favorable safety profile: Model for assessment of live vaccines with neurotropic potential.

Nipah virus (NiV) disease is a bat-borne zoonosis responsible for outbreaks with high lethality and is a priority for vaccine development. With funding from the Coalition of Epidemic Preparedness Innovations (CEPI), we are developing a chimeric vaccine (PHV02) composed of recombinant vesicular stomatitis virus (VSV) expressing the envelope glycoproteins of both Ebola virus (EBOV) and NiV. The EBOV glycoprotein (GP) mediates fusion and viral entry and the NiV attachment glycoprotein (G) is a ligand for cell receptors, and stimulates neutralizing antibody, the putative mediator of protection against NiV. PHV02 is identical in construction to the registered Ebola vaccine (Ervebo) with the addition of the NiV G gene. NiV ephrin B2 and B3 receptors are expressed on neural cells and the wild-type NiV is neurotropic and causes encephalitis in affected patients. It was therefore important to assess whether the NiV G alters tropism of the rVSV vector and serves as a virulence factor. PHV02 was fully attenuated in adult hamsters inoculated by the intramuscular (IM) route, whereas parental wild-type VSV was 100% lethal. Two rodent models (mice, hamsters) were infected by the intracerebral (IC) route with graded doses of PHV02. Comparator active controls in various experiments included rVSV-EBOV (representative of Ebola vaccine) and yellow fever (YF) 17DD commercial vaccine. These studies showed PHV02 to be more neurovirulent than both rVSV-EBOV and YF 17DD in infant animals. PHV02 was lethal for adult hamsters inoculated IC but not for adult mice. In contrast YF 17DD retained virulence for adult mice inoculated IC but was not virulent for adult hamsters. Because of the inconsistency of neurovirulence patterns in the rodent models, a monkey neurovirulence test (MNVT) was performed, using YF 17DD as the active comparator because it has a well-established profile of quantifiable microscopic changes in brain centers and a known reporting rate of neurotropic adverse events in humans. In the MNVT PHV02 was significantly less neurovirulent than the YF 17DD vaccine reference control, indicating that the vaccine will have an acceptable safety profile for humans. The findings are important because they illustrate the complexities of phenotypic assessment of novel viral vectors with tissue tropisms determined by transgenic proteins, and because it is unprecedented to use a heterologous comparator virus (YF vaccine) in a regulatory-enabling study. This approach may have value in future studies of other novel viral vectors.

Real-time RT-PCR for Venezuelan equine encephalitis complex, Madariaga, and Eastern equine encephalitis viruses: application in human and mosquito public health surveillance in Panama.

Eastern equine encephalitis virus (EEEV), Madariaga virus (MADV), and Venezuelan equine encephalitis virus complex (VEEV) are New World alphaviruses transmitted by mosquitoes. They cause febrile and sometimes severe neurological diseases in human and equine hosts. Detecting them during the acute phase is hindered by non-specific symptoms and limited diagnostic tools. We designed and clinically assessed real-time reverse transcription polymerase chain reaction assays (rRT-PCRs) for VEEV complex, MADV, and EEEV using whole-genome sequences. Validation involved 15 retrospective serum samples from 2015 to 2017 outbreaks, 150 mosquito pools from 2015, and 118 prospective samples from 2021 to 2022 surveillance in Panama. The rRT-PCRs detected VEEV complex RNA in 10 samples (66.7%) from outbreaks, with one having both VEEV complex and MADV RNAs. VEEV complex RNA was found in five suspected dengue cases from disease surveillance. The rRT-PCR assays identified VEEV complex RNA in three Culex (Melanoconion) vomerifer pools, leading to VEEV isolates in two. Phylogenetic analysis revealed the VEEV ID subtype in positive samples. Notably, 11.9% of dengue-like disease patients showed VEEV infections. Together, our rRT-PCR validation in human and mosquito samples suggests that this method can be incorporated into mosquito and human encephalitic alphavirus surveillance programs in endemic regions.

A Zika virus envelope mutation preceding the 2015 epidemic enhances virulence and fitness for transmission.

Arboviruses maintain high mutation rates due to lack of proofreading ability of their viral polymerases, in some cases facilitating adaptive evolution and emergence. Here we show that, just before its 2013 spread to the Americas, Zika virus (ZIKV) underwent an envelope protein V473M substitution (E-V473M) that increased neurovirulence, maternal-to-fetal transmission, and viremia to facilitate urban transmission. A preepidemic Asian ZIKV strain (FSS13025 isolated in Cambodia in 2010) engineered with the V473M substitution significantly increased neurovirulence in neonatal mice and produced higher viral loads in the placenta and fetal heads in pregnant mice. Conversely, an epidemic ZIKV strain (PRVABC59 isolated in Puerto Rico in 2015) engineered with the inverse M473V substitution reversed the pathogenic phenotypes. Although E-V473M did not affect oral infection of Aedes aegypti mosquitoes, competition experiments in cynomolgus macaques showed that this mutation increased its fitness for viremia generation, suggesting adaptive evolution for human viremia and hence transmission. Mechanistically, the V473M mutation, located at the second transmembrane helix of the E protein, enhances virion morphogenesis. Overall, our study revealed E-V473M as a critical determinant for enhanced ZIKV virulence, intrauterine transmission during pregnancy, and viremia to facilitate urban transmission.

ICTV Virus Taxonomy Profile: Rhabdoviridae 2022.

The family Rhabdoviridae comprises viruses with negative-sense (-) RNA genomes of 10-16 kb. Virions are typically enveloped with bullet-shaped or bacilliform morphology but can also be non-enveloped filaments. Rhabdoviruses infect plants or animals, including mammals, birds, reptiles, amphibians or fish, as well as arthropods, which serve as single hosts or act as biological vectors for transmission to animals or plants. Rhabdoviruses include important pathogens of humans, livestock, fish or agricultural crops. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Rhabdoviridae, which is available at ictv.global/report/rhabdoviridae.

Is zoonotic Plasmodium vivax malaria an obstacle for disease elimination?

BACKGROUND: The groundwork for malaria elimination does not currently consider the potential of Plasmodium zoonotic cycles that involve non-human primates (NHPs) in sylvatic environments. Since vivax malaria is less responsive to control measures, finding Plasmodium vivax infected NHPs adds even more concern. METHODS: Both Free-living monkeys in forest fragments inside the urban area and captive monkeys from a local zoo had blood samples tested for Plasmodium species. RESULTS: In this study, among the Neotropical monkeys tested, three (4.4%), one captive and two free-living, were found to be naturally infected by P. vivax. CONCLUSION: This important finding indicates that it is necessary to estimate the extent to which P. vivax NHP infection contributes to the maintenance of malaria transmission to humans. Therefore, the discussion on wildlife conservation and management must be incorporated into the malaria elimination agenda.

ICTV Virus Taxonomy Profile: Nyamiviridae 2021.

Nyamiviridae is a family of viruses in the order Mononegavirales, with unsegmented (except for members of the genus Tapwovirus), negative-sense RNA genomes of 10-13 kb. Nyamviruses have a genome organisation and content similar to that of other mononegaviruses. Nyamiviridae includes several genera that form monophyletic clades on phylogenetic analysis of the RNA polymerase. Nyamiviruses have been found associated with diverse invertebrates as well as land- and seabirds. Members of the genera Nyavirus and Socyvirus produce enveloped, spherical virions. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Nyamiviridae, which is available at ictv.global/report/nyamiviridae.

Presentation of fatal stroke due to SARS-CoV-2 and dengue virus coinfection.

Herein, we report a case of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and dengue coinfection, presented as a fatal stroke in our hospital, in São José do Rio Preto, São Paulo State, a Brazilian city hyperendemic for dengue viruses and other arthropod-borne viruses (arboviruses) and currently facing a surge of SARS-CoV-2 cases. This case is the first described in the literature and contributes to the better understanding of clinical presentations of two important diseases in a tropical setting.

Zika, Chikungunya, and Other Emerging Vector-Borne Viral Diseases.

Arthropod-borne viruses (arboviruses) have a long history of emerging to infect humans, but during recent decades, they have been spreading more widely and affecting larger populations. This is due to several factors, including increased air travel and uncontrolled mosquito vector populations. Emergence can involve simple spillover from enzootic (wildlife) cycles, as in the case of West Nile virus accompanying geographic expansion into the Americas; secondary amplification in domesticated animals, as seen with Japanese encephalitis, Venezuelan equine encephalitis, and Rift Valley fever viruses; and urbanization, in which humans become the amplification hosts and peridomestic mosquitoes, mainly Aedes aegypti, mediate human-to-human transmission. Dengue, yellow fever, chikungunya, and Zika viruses have undergone such urban emergence. We focus mainly on the latter two, which are recent arrivals in the Western Hemisphere. We also discuss a few other viruses with the potential to emerge through all of these mechanisms.

Strengthening the Interaction of the Virology Community with the International Committee on Taxonomy of Viruses (ICTV) by Linking Virus Names and Their Abbreviations to Virus Species.

The International Committee on Taxonomy of Viruses (ICTV) is tasked with classifying viruses into taxa (phyla to species) and devising taxon names. Virus names and virus name abbreviations are currently not within the ICTV's official remit and are not regulated by an official entity. Many scientists, medical/veterinary professionals, and regulatory agencies do not address evolutionary questions nor are they concerned with the hierarchical organization of the viral world, and therefore, have limited use for ICTV-devised taxa. Instead, these professionals look to the ICTV as an expert point source that provides the most current taxonomic affiliations of viruses of interests to facilitate document writing. These needs are currently unmet as an ICTV-supported, easily searchable database that includes all published virus names and abbreviations linked to their taxa is not available. In addition, in stark contrast to other biological taxonomic frameworks, virus taxonomy currently permits individual species to have several members. Consequently, confusion emerges among those who are not aware of the difference between taxa and viruses, and because certain well-known viruses cannot be located in ICTV publications or be linked to their species. In addition, the number of duplicate names and abbreviations has increased dramatically in the literature. To solve this conundrum, the ICTV could mandate listing all viruses of established species and all reported unclassified viruses in forthcoming online ICTV Reports and create a searchable webpage using this information. The International Union of Microbiology Societies could also consider changing the mandate of the ICTV to include the nomenclature of all viruses in addition to taxon considerations. With such a mandate expansion, official virus names and virus name abbreviations could be catalogued and virus nomenclature could be standardized. As a result, the ICTV would become an even more useful resource for all stakeholders in virology.

ICTV Virus Taxonomy Profile: Artoviridae.

The family Artoviridae was created in 2018 for the established monospecific genus Peropuvirus and six new species of invertebrate viruses that had all been discovered by high-throughput sequencing. Artoviruses have negative-sense RNA genomes of about 12 kb and produce enveloped, spherical particles that are 100-130 nm in diameter. Hosts include parasitoid wasps, barnacles, pillworms, woodlice, copepods and odonates. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Artoviridae, which is available at www.ictv.global/report/artoviridae.

Characterization of Triniti virus supports its reclassification in the family Peribunyaviridae.

Triniti virus (TNTV) has been isolated in Trinidad and Tobago and in Brazil. To date little is known about this virus, which is classified as an ungrouped virus within the family Togaviridae. Here, three isolates of TNTV were characterized both genetically and antigenically. The genome was shown to contain three RNA segments: small (S), medium (M) and large (L). Genome organization, protein sizes and protein motifs were similar to those of viruses in the genus Orthobunyavirus, family Peribunyaviridae. Antigenic reactivity revealed the three TNTV isolates to be closely related, but no serologic cross-reaction with other orthobunyaviruses. Morphological observation by transmission electron microscopy indicated that virus size and symmetry were compatible with those of viruses in the family Peribunyaviridae. Our serological, morphological and molecular results support the taxonomic reclassification of TNTV as a member of the genus Orthobunyavirus, family Peribunyaviridae.

Genomic characterisation of Cuiaba and Charleville viruses: arboviruses (family Rhabdoviridae, genus Sripuvirus) infecting reptiles and amphibians.

Viruses in the family Rhabdoviridae are ecologically very diverse, infecting mammals, birds, reptiles, fish, plants and a wide range of other terrestrial and aquatic invertebrates. The genus Sripuvirus currently comprises five viruses that appear to circulate in transmission cycles involving reptiles and sandflies. Here, we report an analysis of the complete coding sequences of Cuiaba virus (CUIV), isolated from an amphibian (Bufo marinus) collected in Brazil, and Charleville virus (CHVV), isolated from sandflies (Phlebotomus sp.) and lizards (Gehyra australis), collected in Australia. CUIV and CHVV cluster phylogenetically with the sripuviruses in maximum likelihood trees generated from complete L protein (RdRp) sequences. They also share with sripuviruses unique features in genome organisation, including an additional gene (U1) between the matrix protein (M) gene and glycoprotein (G) gene, and an alternative long open reading frame near the start of the G ORF that encodes a predicted transmembrane protein. In view of their phylogenetic relationships, similar genome organisations and similar ecological characteristics, we propose the assignment of CUIV and CHVV as novel members of the genus Sripuvirus.

ICTV Virus Taxonomy Profile: Rhabdoviridae.

The family Rhabdoviridae comprises viruses with negative-sense (-) single-stranded RNA genomes of 10.8-16.1 kb. Virions are typically enveloped with bullet-shaped or bacilliform morphology but can also be non-enveloped filaments. Rhabdoviruses infect plants and animals including mammals, birds, reptiles and fish, as well as arthropods which serve as single hosts or act as biological vectors for transmission to animals or plants. Rhabdoviruses include important pathogens of humans, livestock, fish and agricultural crops. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of Rhabdoviridae, which is available at www.ictv.global/report/rhabdoviridae.

History and Emergence of Zika Virus.

Zika virus was discovered in East Africa in 1947 by the Rockefeller Foundation during investigations on the ecology of yellow fever. Although it was subsequently shown to have widespread distribution in Africa and Asia, it was not known to cause epidemics until 2007. This paper describes the history of the virus discovery, emergence and evolution as an epidemic virus, and the its evolving clinical spectrum.

Viral Load and Cytokine Response Profile Does Not Support Antibody-Dependent Enhancement in Dengue-Primed Zika Virus-Infected Patients.

Background: The pathogenesis of severe dengue disease involves immune components as biomarkers. The mechanism by which some dengue virus (DENV)-infected individuals progress to severe disease is poorly understood. Most studies on the pathogenesis of severe dengue disease focus on the process of antibody-dependent enhancement (ADE) as a primary risk factor. With the circulation of Zika virus (ZIKV) in DENV-endemic areas, many people infected by ZIKV were likely exposed to DENV. The influence of such exposure on Zika disease outcomes remains unknown. Methods: We investigated whether patients previously exposed to DENV exhibited higher viremia when exposed to a subsequent, heterologous dengue or Zika infection than those patients not previously exposed to dengue. We measured viral loads and cytokine profile during patients' acute infections. Results: Neither dengue nor Zika viremia was higher in patients with prior DENV infection, although the power to detect such a difference was only adequate in the ZIKV analysis. Of the 10 cytokines measured, only 1 significant difference was detected: Levels of interleukin 1ß (IL-1ß) were lower in dengue-infected patients who had experienced a previous dengue infection than patients infected with dengue for the first time. However, power to detect differences between groups was low. In Zika-infected patients, levels of IL-1ß showed a significant, positive correlation with viral load. Conclusions: No signs of ADE were observed in vivo in patients with acute ZIKV infection who had prior exposure to DENV.

Transient Hearing Loss in Adults Associated With Zika Virus Infection.

In 2015, during the outbreak of Zika virus (ZIKV) in Brazil, we identified 3 cases of acute hearing loss after exanthematous illness. Serology yielded finding compatible with ZIKV as the cause of a confirmed (n = 1) and a probable (n = 2) flavivirus infection, indicating an association between ZIKV infection and transient hearing loss.

Zika Virus Vector Competency of Mosquitoes, Gulf Coast, United States.

Zika virus has recently spread throughout the Americas. Although Aedes aegypti mosquitoes are considered the primary vector, Culex quinquefasciatus and mosquitoes of other species may also be vectors. We tested Cx. quinquefasciatus and Ae. taeniorhynchus mosquitoes from the US Gulf Coast; both were refractory to infection and incapable of transmission.

Variation in Aedes aegypti Mosquito Competence for Zika Virus Transmission.

To test whether Zika virus has adapted for more efficient transmission by Aedes aegypti mosquitoes, leading to recent urban outbreaks, we fed mosquitoes from Brazil, the Dominican Republic, and the United States artificial blood meals containing 1 of 3 Zika virus strains (Senegal, Cambodia, Mexico) and monitored infection, dissemination, and virus in saliva. Contrary to our hypothesis, Cambodia and Mexica strains were less infectious than the Senegal strain. Only mosquitoes from the Dominican Republic transmitted the Cambodia and Mexica strains. However, blood meals from viremic mice were more infectious than artificial blood meals of comparable doses; the Cambodia strain was not transmitted by mosquitoes from Brazil after artificial blood meals, whereas 61% transmission occurred after a murine blood meal (saliva titers up to 4 log 10 infectious units/collection). Although regional origins of vector populations and virus strain influence transmission efficiency, Ae. aegypti mosquitoes appear to be competent vectors of Zika virus in several regions of the Americas.