A taxonomic proposal for cedratviruses, orpheoviruses, and pithoviruses.

Orpheoviruses, cedratviruses, and pithoviruses are large DNA viruses that cluster together taxonomically within the order Pimascovirales of the phylum Nucleocytoviricota. However, they were not classified previously by the International Committee on Taxonomy of Viruses (ICTV). Here, we present a comprehensive analysis of the gene content, morphology, and phylogenomics of these viruses, providing data that underpinned the recent proposal to establish new taxa for their initial classification. The new taxonomy, which has now been ratified by the ICTV, includes the family Orpheoviridae and genus Alphaorpheovirus, the family Pithoviridae and genus Alphapithovirus, and the family Cedratviridae and genus Alphacedratvirus, aiming to formally catalogue the isolates covered in this study. Additionally, as per the newly adopted rules, we applied standardized binomial names for the virus species created to classify isolates with complete genome sequences available in public databases at the time of the proposal. The specific epithet of each virus species was chosen as a reference to the location where the exemplar virus was isolated.

Gene duplication as a major force driving the genome expansion in some giant viruses.

IMPORTANCE: Giant viruses are noteworthy not only due to their enormous particles but also because of their gigantic genomes. In this context, a fundamental question has persisted: how did these genomes evolve? Here we present the discovery of cedratvirus pambiensis, featuring the largest genome ever described for a cedratvirus. Our data suggest that the larger size of the genome can be attributed to an unprecedented number of duplicated genes. Further investigation of this phenomenon in other viruses has illuminated gene duplication as a key evolutionary mechanism driving genome expansion in diverse giant viruses. Although gene duplication has been described as a recurrent event in cellular organisms, our data highlights its potential as a pivotal event in the evolution of gigantic viral genomes.

Analysis of the Genomic Features and Evolutionary History of Pithovirus-Like Isolates Reveals Two Major Divergent Groups of Viruses.

New representatives of the phylum Nucleocytoviricota have been rapidly described in the last decade. Despite this, not all viruses of this phylum are allocated to recognized taxonomic families, as is the case for orpheovirus, pithovirus, and cedratvirus, which form the proposed family Pithoviridae. In this study, we performed comprehensive comparative genomic analyses of 8 pithovirus-like isolates, aiming to understand their common traits and evolutionary history. Structural and functional genome annotation was performed de novo for all the viruses, which served as a reference for pangenome construction. The synteny analysis showed substantial differences in genome organization between these viruses, with very few and short syntenic blocks shared between orpheovirus and its relatives. It was possible to observe an open pangenome with a significant increase in the slope when orpheovirus was added, alongside a decrease in the core genome. Network analysis placed orpheovirus as a distant and major hub with a large fraction of unique clusters of orthologs, indicating a distant relationship between this virus and its relatives, with only a few shared genes. Additionally, phylogenetic analyses of strict core genes shared with other viruses of the phylum reinforced the divergence of orpheovirus from pithoviruses and cedratviruses. Altogether, our results indicate that although pithovirus-like isolates share common features, this group of ovoid-shaped giant viruses presents substantial differences in gene contents, genomic architectures, and the phylogenetic history of several core genes. Our data indicate that orpheovirus is an evolutionarily divergent viral entity, suggesting its allocation to a different viral family, Orpheoviridae. IMPORTANCE Giant viruses that infect amoebae form a monophyletic group named the phylum Nucleocytoviricota. Despite being genomically and morphologically very diverse, the taxonomic categories of some clades that form this phylum are not yet well established. With advances in isolation techniques, the speed at which new giant viruses are described has increased, escalating the need to establish criteria to define the emerging viral taxa. In this work, we performed a comparative genomic analysis of representatives of the putative family Pithoviridae. Based on the dissimilarity of orpheovirus from the other viruses of this putative family, we propose that orpheovirus be considered a member of an independent family, Orpheoviridae, and suggest criteria to demarcate families consisting of ovoid-shaped giant viruses.

A Brief History of Giant Viruses' Studies in Brazilian Biomes.

Almost two decades after the isolation of the first amoebal giant viruses, indubitably the discovery of these entities has deeply affected the current scientific knowledge on the virosphere. Much has been uncovered since then: viruses can now acknowledge complex genomes and huge particle sizes, integrating remarkable evolutionary relationships that date as early as the emergence of life on the planet. This year, a decade has passed since the first studies on giant viruses in the Brazilian territory, and since then biomes of rare beauty and biodiversity (Amazon, Atlantic forest, Pantanal wetlands, Cerrado savannas) have been explored in the search for giant viruses. From those unique biomes, novel viral entities were found, revealing never before seen genomes and virion structures. To celebrate this, here we bring together the context, inspirations, and the major contributions of independent Brazilian research groups to summarize the accumulated knowledge about the diversity and the exceptionality of some of the giant viruses found in Brazil.

Functional Genomic Analyses Reveal an Open Pan-genome for the Chloroviruses and a Potential for Genetic Innovation in New Isolates.

Chloroviruses (family Phycodnaviridae) are large double-stranded DNA (dsDNA) viruses that infect unicellular green algae present in inland waters. These viruses have been isolated using three main chlorella-like green algal host cells, traditionally called NC64A, SAG, and Pbi, revealing extensive genetic diversity. In this study, we performed a functional genomic analysis on 36 chloroviruses that infected the three different hosts. Phylogenetic reconstruction based on the DNA polymerase B family gene clustered the chloroviruses into three distinct clades. The viral pan-genome consists of 1,345 clusters of orthologous groups of genes (COGs), with 126 COGs conserved in all viruses. Totals of 368, 268, and 265 COGs are found exclusively in viruses that infect NC64A, SAG, and Pbi algal hosts, respectively. Two-thirds of the COGs have no known function, constituting the "dark pan-genome" of chloroviruses, and further studies focusing on these genes may identify important novelties. The proportions of functionally characterized COGs composing the pan-genome and the core-genome are similar, but those related to transcription and RNA processing, protein metabolism, and virion morphogenesis are at least 4-fold more represented in the core genome. Bipartite network construction evidencing the COG sharing among host-specific viruses identified 270 COGs shared by at least one virus from each of the different host groups. Finally, our results reveal an open pan-genome for chloroviruses and a well-established core genome, indicating that the isolation of new chloroviruses can be a valuable source of genetic discovery. IMPORTANCE Chloroviruses are large dsDNA viruses that infect unicellular green algae distributed worldwide in freshwater environments. They comprise a genetically diverse group of viruses; however, a comprehensive investigation of the genomic evolution of these viruses is still missing. Here, we performed a functional pan-genome analysis comprising 36 chloroviruses associated with three different algal hosts in the family Chlorellaceae, referred to as zoochlorellae because of their endosymbiotic lifestyle. We identified a set of 126 highly conserved genes, most of which are related to essential functions in the viral replicative cycle. Several genes are unique to distinct isolates, resulting in an open pan-genome for chloroviruses. This profile is associated with generalist organisms, and new insights into the evolution and ecology of chloroviruses are presented. Ultimately, our results highlight the potential for genetic diversity in new isolates.

In-depth characterization of congenital Zika syndrome in immunocompetent mice: Antibody-dependent enhancement and an antiviral peptide therapy.

BACKGROUND: Zika virus (ZIKV) infection during pregnancy may cause major congenital defects, including microcephaly, ocular, articular and muscle abnormalities, which are collectively defined as Congenital Zika Syndrome. Here, we performed an in-depth characterization of the effects of congenital ZIKV infection (CZI) in immunocompetent mice. METHODS: Pregnant dams were inoculated with ZIKV on embryonic day 5.5 in the presence or absence of a sub-neutralizing dose of a pan-flavivirus monoclonal antibody (4G2) to evaluate the potential role of antibody-dependent enhancement phenomenon (ADE) during short and long outcomes of CZI. FINDINGS: ZIKV infection induced maternal immune activation (MIA), which was associated with occurrence of foetal abnormalities and death. Therapeutic administration of AH-D antiviral peptide during the early stages of pregnancy prevented ZIKV replication and death of offspring. In the post-natal period, CZI was associated with a decrease in whole brain volume, ophthalmologic abnormalities, changes in testicular morphology, and disruption in bone microarchitecture. Some alterations were enhanced in the presence of 4G2 antibody. INTERPRETATION: Our results reveal that early maternal ZIKV infection causes several birth defects in immunocompetent mice, which can be potentiated by ADE phenomenon and are associated with MIA. Additionally, antiviral treatment with AH-D peptide may be beneficial during early maternal ZIKV infection. FUND: This work was supported by the Brazilian National Science Council (CNPq, Brazil), Minas Gerais Foundation for Science (FAPEMIG), Funding Authority for Studies and Projects (FINEP), Coordination of Superior Level Staff Improvement (CAPES), National Research Foundation of Singapore and Centre for Precision Biology at Nanyang Technological University.

Therapeutic treatment of Zika virus infection using a brain-penetrating antiviral peptide.

Zika virus is a mosquito-borne virus that is associated with neurodegenerative diseases, including Guillain-Barré syndrome1 and congenital Zika syndrome2. As Zika virus targets the nervous system, there is an urgent need to develop therapeutic strategies that inhibit Zika virus infection in the brain. Here, we have engineered a brain-penetrating peptide that works against Zika virus and other mosquito-borne viruses. We evaluated the therapeutic efficacy of the peptide in a lethal Zika virus mouse model exhibiting systemic and brain infection. Therapeutic treatment protected against mortality and markedly reduced clinical symptoms, viral loads and neuroinflammation, as well as mitigated microgliosis, neurodegeneration and brain damage. In addition to controlling systemic infection, the peptide crossed the blood-brain barrier to reduce viral loads in the brain and protected against Zika-virus-induced blood-brain barrier injury. Our findings demonstrate how engineering strategies can be applied to develop peptide therapeutics and support the potential of a brain-penetrating peptide to treat neurotropic viral infections.