RESUMO
The discovery of mimivirus in 2003 prompted the search for novel giant viruses worldwide. Despite increasing interest, the diversity and distribution of giant viruses is barely known. Here, we present data from a 2012-2022 study aimed at prospecting for amoebal viruses in water, soil, mud, and sewage samples across Brazilian biomes, using Acanthamoeba castellanii for isolation. A total of 881 aliquots from 187 samples covering terrestrial and marine Brazilian biomes were processed. Electron microscopy and PCR were used to identify the obtained isolates. Sixty-seven amoebal viruses were isolated, including mimiviruses, marseilleviruses, pandoraviruses, cedratviruses, and yaraviruses. Viruses were isolated from all tested sample types and almost all biomes. In comparison to other similar studies, our work isolated a substantial number of Marseillevirus and cedratvirus representatives. Taken together, our results used a combination of isolation techniques with microscopy, PCR, and sequencing and put highlight on richness of giant virus present in different terrestrial and marine Brazilian biomes.
Assuntos
Vírus Gigantes , Brasil , Vírus Gigantes/isolamento & purificação , Vírus Gigantes/genética , Vírus Gigantes/classificação , Vírus Gigantes/ultraestrutura , Filogenia , Reação em Cadeia da Polimerase , Acanthamoeba castellanii/virologia , Acanthamoeba castellanii/isolamento & purificação , Microbiologia do Solo , Esgotos/virologia , Análise de Sequência de DNA , Água do Mar/virologia , Microbiologia da ÁguaRESUMO
Giant viruses (Nucleocytoviricota) are significant lethality agents of various eukaryotic hosts. Although metagenomics indicates their ubiquitous distribution, available giant virus isolates are restricted to a very small number of protist and algal hosts. Here we report on the first viral isolate that replicates in the amoeboflagellate Naegleria. This genus comprises the notorious human pathogen Naegleria fowleri, the causative agent of the rare but fatal primary amoebic meningoencephalitis. We have elucidated the structure and infection cycle of this giant virus, Catovirus naegleriensis (a.k.a. Naegleriavirus, NiV), and show its unique adaptations to its Naegleria host using fluorescence in situ hybridization, electron microscopy, genomics, and proteomics. Naegleriavirus is only the fourth isolate of the highly diverse subfamily Klosneuvirinae, and like its relatives the NiV genome contains a large number of translation genes, but lacks transfer RNAs (tRNAs). NiV has acquired genes from its Naegleria host, which code for heat shock proteins and apoptosis inhibiting factors, presumably for host interactions. Notably, NiV infection was lethal to all Naegleria species tested, including the human pathogen N. fowleri. This study expands our experimental framework for investigating giant viruses and may help to better understand the basic biology of the human pathogen N. fowleri.
Assuntos
Genoma Viral , Vírus Gigantes , Naegleria , Genoma Viral/genética , Vírus Gigantes/genética , Vírus Gigantes/classificação , Vírus Gigantes/ultraestrutura , Vírus Gigantes/isolamento & purificação , Vírus Gigantes/fisiologia , Naegleria/genética , Naegleria/virologia , Naegleria fowleri/genética , Naegleria fowleri/isolamento & purificação , Filogenia , HumanosRESUMO
Medusavirus, a giant virus, is phylogenetically closer to eukaryotes than the other giant viruses and has been recently classified as an independent species. However, details of its morphology and maturation process in host cells remain unclear. Here, we investigated the particle morphology of medusavirus inside and outside infected cells using conventional transmission electron microscopy (C-TEM) and cryo-electron microscopy (cryo-EM). The C-TEM of amoebae infected with the medusavirus showed four types of particles, i.e., pseudo-DNA-empty (p-Empty), DNA-empty (Empty), semi-DNA-full (s-Full), and DNA-full (Full). Time-dependent changes in the four types of particles and their intracellular localization suggested a new maturation process for the medusavirus. Viral capsids and viral DNAs are produced independently in the cytoplasm and nucleus, respectively, and only the empty particles located near the host nucleus can incorporate the viral DNA into the capsid. Therefore, all four types of particles were found outside the cells. The cryo-EM of these particles showed that the intact virus structure, covered with three different types of spikes, was preserved among all particle types, although with minor size-related differences. The internal membrane exhibited a structural array similar to that of the capsid, interacted closely with the capsid, and displayed open membrane structures in the Empty and p-Empty particles. The results suggest that these open structures in the internal membrane are used for an exchange of scaffold proteins and viral DNA during the maturation process. This new model of the maturation process of medusavirus provides insight into the structural and behavioral diversity of giant viruses. IMPORTANCE Giant viruses exhibit diverse morphologies and maturation processes. In this study, medusavirus showed four types of particle morphologies, both inside and outside the infected cells, when propagated in amoeba culture. Time-course analysis and intracellular localization of the medusavirus in the infected cells suggested a new maturation process via the four types of particles. Like the previously reported pandoravirus, the viral DNA of medusavirus is replicated in the host's nucleus. However, viral capsids are produced independently in the host cytoplasm, and only empty capsids near the nucleus can take up viral DNA. As a result, many immature particles were released from the host cell along with the mature particles. The capsid structure is well conserved among the four types of particles, except for the open membrane structures in the empty particles, suggesting that they are used to exchange scaffold proteins for viral DNAs. These findings indicate that medusavirus has a unique maturation process.
Assuntos
Vírus Gigantes , Capsídeo/metabolismo , Capsídeo/ultraestrutura , Microscopia Crioeletrônica , DNA Viral/metabolismo , Genoma Viral , Vírus Gigantes/genética , Vírus Gigantes/metabolismo , Vírus Gigantes/ultraestrutura , Microscopia Eletrônica de TransmissãoRESUMO
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.
Assuntos
Amoeba/virologia , Vírus Gigantes/genética , Vírus Gigantes/isolamento & purificação , Virologia/história , Biodiversidade , Brasil , Ecossistema , Genoma Viral , Vírus Gigantes/classificação , Vírus Gigantes/ultraestrutura , História do Século XXI , FilogeniaRESUMO
Large and giant DNA viruses are a monophyletic group constituting the recently established phylum Nucleocytoviricota. The virus particle morphogenesis of these viruses exhibit striking similarities. Viral factories are established in the host cells where new virions are assembled by recruiting host membranes, forming an inner lipid layer. An outer protein layer starts as a lamellar structure, commonly referred to as viral crescents, coded by the major capsid protein gene. Also, these viruses have a conserved ATPase-coding gene related to genome encapsidation. Similar properties are described for tectiviruses, putative small ancestors of giant viruses. Here we review the morphogenesis of giant viruses and discuss how the process similarities constitute additional evidence to the common origin of Nucleocytoviricota.
Assuntos
Amébidos/virologia , Vírus Gigantes/classificação , Vírus Gigantes/crescimento & desenvolvimento , Capsídeo/fisiologia , Capsídeo/ultraestrutura , Evolução Molecular , Vírus Gigantes/genética , Vírus Gigantes/ultraestrutura , Morfogênese , Filogenia , Proteínas Virais/genética , Proteínas Virais/metabolismo , Montagem de Vírus , Replicação ViralRESUMO
Although giant viruses have existed for millennia and possibly exerted great evolutionary influence in their environment. Their presence has only been noticed by virologists recently with the discovery of Acanthamoeba polyphaga mimivirus in 2003. Its virion with a diameter of 500 nm and its genome larger than 1 Mpb shattered preconceived standards of what a virus is and triggered world-wide prospection studies. Thanks to these investigations many giant virus families were discovered, each with its own morphological peculiarities and genomes ranging from 0.4 to 2.5 Mpb that possibly encode more than 400 viral proteins. This review aims to present the morphological diversity, the different aspects observed in host-virus interactions during replication, as well as the techniques utilized during their investigation.
Assuntos
Amébidos/virologia , Vírus Gigantes/fisiologia , Vírus Gigantes/ultraestrutura , Interações entre Hospedeiro e Microrganismos , Acanthamoeba castellanii/virologia , Genoma Viral , Vírus Gigantes/classificação , Vírus Gigantes/genética , Proteínas Virais/genética , Proteínas Virais/metabolismo , Compartimentos de Replicação Viral/fisiologia , Vírion/fisiologia , Vírion/ultraestrutura , Replicação ViralRESUMO
Bioconversion of organic materials is the foundation of many applications in chemical engineering, microbiology and biochemistry. Herein, we introduce a new methodology to quantitatively determine conversion of biomass in viral infections while simultaneously imaging morphological changes of the host cell. As proof of concept, the viral replication of an unidentified giant DNA virus and the cellular response of an amoebal host are studied using soft X-ray microscopy, titration dilution measurements and thermal gravimetric analysis. We find that virions produced inside the cell are visible from 18 h post infection and their numbers increase gradually to a burst size of 280-660 virions. Due to the large size of the virion and its strong X-ray absorption contrast, we estimate that the burst size corresponds to a conversion of 6-12% of carbonaceous biomass from amoebal host to virus. The occurrence of virion production correlates with the appearance of a possible viral factory and morphological changes in the phagosomes and contractile vacuole complex of the amoeba, whereas the nucleus and nucleolus appear unaffected throughout most of the replication cycle.
Assuntos
Acanthamoeba/virologia , Vírus de DNA/ultraestrutura , DNA Viral/genética , Genoma Viral , Vírus Gigantes/ultraestrutura , Vírion/ultraestrutura , Acanthamoeba/ultraestrutura , Biomassa , Vírus de DNA/genética , Vírus de DNA/crescimento & desenvolvimento , Vírus de DNA/isolamento & purificação , DNA Viral/biossíntese , Vírus Gigantes/genética , Vírus Gigantes/crescimento & desenvolvimento , Vírus Gigantes/isolamento & purificação , Interações Hospedeiro-Patógeno/genética , Fagossomos/ultraestrutura , Fagossomos/virologia , Microbiologia do Solo , Termogravimetria , Vacúolos/ultraestrutura , Vacúolos/virologia , Vírion/genética , Vírion/crescimento & desenvolvimento , Replicação Viral , Microtomografia por Raio-XRESUMO
Kaumoebavirus infects the amoeba Vermamoeba vermiformis and has recently been described as a distant relative of the African swine fever virus. To characterize the diversity and evolution of this novel viral genus, we report here on the isolation and genome sequencing of a second strain of Kaumoebavirus, namely LCC10. Detailed analysis of the sequencing data suggested that its 362-Kb genome is linear with covalently closed hairpin termini, so that DNA forms a single continuous polynucleotide chain. Comparative genomic analysis indicated that although the two sequenced Kaumoebavirus strains share extensive gene collinearity, 180 predicted genes were either gained or lost in only one genome. As already observed in another distant relative, i.e., Faustovirus, which infects the same host, the center and extremities of the Kaumoebavirus genome exhibited a higher rate of sequence divergence and the major capsid protein gene was colonized by type-I introns. A possible role of the Vermamoeba host in the genesis of these evolutionary traits is hypothesized. The Kaumoebavirus genome exhibited a significant gene strand bias over the two-third of genome length, a feature not seen in the other members of the "extended Asfarviridae" clade. We suggest that this gene strand bias was induced by a putative single origin of DNA replication located near the genome extremity that imparted a selective force favoring the genes positioned on the leading strand.
Assuntos
Asfarviridae/genética , Genoma Viral , Vírus Gigantes/genética , Vírus não Classificados/genética , Asfarviridae/classificação , Proteínas do Capsídeo/genética , Replicação do DNA , Vírus de DNA/classificação , Vírus de DNA/genética , Vírus de DNA/isolamento & purificação , DNA Viral/química , DNA Viral/genética , DNA Viral/metabolismo , Evolução Molecular , Genes Virais , Vírus Gigantes/classificação , Vírus Gigantes/isolamento & purificação , Vírus Gigantes/ultraestrutura , Lobosea/virologia , Filogenia , Esgotos/virologia , Proteínas Virais/genética , Vírus não Classificados/isolamento & purificação , Vírus não Classificados/ultraestruturaRESUMO
"Giant" phages have genomes of >200 kbp, confined in correspondingly large capsids whose assembly and maturation are still poorly understood. Nevertheless, the first assembly product is likely to be, as in other tailed phages, a procapsid that subsequently matures and packages the DNA. The associated transformations include the cleavage of many proteins by the phage-encoded protease, as well as the thinning and angularization of the capsid. We exploited an amber mutation in the viral protease gene of the Salmonella giant phage SPN3US, which leads to the accumulation of a population of capsids with distinctive properties. Cryo-electron micrographs reveal patterns of internal density different from those of the DNA-filled heads of virions, leading us to call them "mottled capsids". Reconstructions show an outer shell with T = 27 symmetry, an embellishment of the HK97 prototype composed of the major capsid protein, gp75, which is similar to some other giant viruses. The mottled capsid has a T = 1 inner icosahedral shell that is a complex network of loosely connected densities composed mainly of the ejection proteins gp53 and gp54. Segmentation of this inner shell indicated that a number of densities (~12 per asymmetric unit) adopt a "twisted hook" conformation. Large patches of a proteinaceous tetragonal lattice with a 67 Å repeat were also present in the cell lysate. The unexpected nature of these novel inner shell and lattice structures poses questions as to their functions in virion assembly.
Assuntos
Capsídeo/metabolismo , Vírus Gigantes/fisiologia , Fagos de Salmonella/fisiologia , Montagem de Vírus , Capsídeo/ultraestrutura , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Microscopia Crioeletrônica , Empacotamento do DNA , Genoma Viral , Vírus Gigantes/genética , Vírus Gigantes/ultraestrutura , Salmonella/virologia , Fagos de Salmonella/genética , Fagos de Salmonella/ultraestrutura , Vírion/genética , Vírion/fisiologia , Vírion/ultraestruturaRESUMO
Microbes trapped in permanently frozen paleosoils (permafrost) are the focus of increasing research in the context of global warming. Our previous investigations led to the discovery and reactivation of two Acanthamoeba-infecting giant viruses, Mollivirus sibericum and Pithovirus sibericum, from a 30,000-year old permafrost layer. While several modern pithovirus strains have since been isolated, no contemporary mollivirus relative was found. We now describe Mollivirus kamchatka, a close relative to M. sibericum, isolated from surface soil sampled on the bank of the Kronotsky River in Kamchatka, Russian Federation. This discovery confirms that molliviruses have not gone extinct and are at least present in a distant subarctic continental location. This modern isolate exhibits a nucleocytoplasmic replication cycle identical to that of M. sibericum Its spherical particle (0.6 µm in diameter) encloses a 648-kb GC-rich double-stranded DNA genome coding for 480 proteins, of which 61% are unique to these two molliviruses. The 461 homologous proteins are highly conserved (92% identical residues, on average), despite the presumed stasis of M. sibericum for the last 30,000 years. Selection pressure analyses show that most of these proteins contribute to virus fitness. The comparison of these first two molliviruses clarify their evolutionary relationship with the pandoraviruses, supporting their provisional classification in a distinct family, the Molliviridae, pending the eventual discovery of intermediary missing links better demonstrating their common ancestry.IMPORTANCE Virology has long been viewed through the prism of human, cattle, or plant diseases, leading to a largely incomplete picture of the viral world. The serendipitous discovery of the first giant virus visible under a light microscope (i.e., >0.3 µm in diameter), mimivirus, opened a new era of environmental virology, now incorporating protozoan-infecting viruses. Planet-wide isolation studies and metagenome analyses have shown the presence of giant viruses in most terrestrial and aquatic environments, including upper Pleistocene frozen soils. Those systematic surveys have led authors to propose several new distinct families, including the Mimiviridae, Marseilleviridae, Faustoviridae, Pandoraviridae, and Pithoviridae We now propose to introduce one additional family, the Molliviridae, following the description of M. kamchatka, the first modern relative of M. sibericum, previously isolated from 30,000-year-old arctic permafrost.
Assuntos
Vírus Gigantes/classificação , Vírus Gigantes/genética , Vírus Gigantes/isolamento & purificação , Filogenia , Acanthamoeba/virologia , Vírus de DNA/classificação , Vírus de DNA/genética , Genoma Viral , Genômica , Vírus Gigantes/ultraestrutura , Mimiviridae/classificação , Mimiviridae/genética , Federação Russa , Microbiologia do Solo , Vírion/genética , Vírion/ultraestrutura , Vírus não Classificados/classificação , Vírus não Classificados/genética , Vírus não Classificados/isolamento & purificaçãoRESUMO
BACKGROUND: After the isolation of Acanthamoeba polyphaga mimivirus (APMV), the study and search for new giant viruses has been intensified. Most giant viruses are associated with free-living amoebae of the genus Acanthamoeba; however other giant viruses have been isolated in Vermamoeba vermiformis, such as Faustovirus, Kaumoebavirus and Orpheovirus. These studies have considerably expanded our knowledge about the diversity, structure, genomics, and evolution of giant viruses. Until now, there has been only one Orpheovirus isolate, and many aspects of its life cycle remain to be elucidated. METHODS: In this study, we performed an in-depth characterization of the replication cycle and particles of Orpheovirus by transmission and scanning electron microscopy, optical microscopy and IF assays. RESULTS: We observed, through optical and IF microscopy, morphological changes in V. vermiformis cells during Orpheovirus infection, as well as increased motility at 12 h post infection (h.p.i.). The viral factory formation and viral particle morphogenesis were analysed by transmission electron microscopy, revealing mitochondria and membrane recruitment into and around the electron-lucent viral factories. Membrane traffic inhibitor (Brefeldin A) negatively impacted particle morphogenesis. The first structure observed during particle morphogenesis was crescent-shaped bodies, which extend and are filled by the internal content until the formation of multi-layered mature particles. We also observed the formation of defective particles with different shapes and sizes. Virological assays revealed that viruses are released from the host by exocytosis at 12 h.p.i., which is associated with an increase of particle counts in the supernatant. CONCLUSIONS: The results presented here contribute to a better understanding of the biology, structures and important steps in the replication cycle of Orpheovirus.
Assuntos
Vírus de DNA/crescimento & desenvolvimento , Vírus Gigantes/crescimento & desenvolvimento , Replicação Viral , Antígenos Virais/análise , Vírus de DNA/ultraestrutura , Vírus Gigantes/ultraestrutura , Lobosea/virologia , Microscopia , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Vírion/química , Vírion/ultraestruturaRESUMO
The family of giant viruses is still expanding, and evidence of a translational machinery is emerging in the virosphere. The Klosneuvirinae group of giant viruses was first reconstructed from in silico studies, and then a unique member was isolated, Bodo saltans virus. Here we describe the isolation of a new member in this group using coculture with the free-living amoeba Vermamoeba vermiformis This giant virus, called Yasminevirus, has a 2.1-Mb linear double-stranded DNA genome encoding 1,541 candidate proteins, with a GC content estimated at 40.2%. Yasminevirus possesses a nearly complete translational machinery, with a set of 70 tRNAs associated with 45 codons and recognizing 20 amino acids (aa), 20 aminoacyl-tRNA synthetases (aaRSs) recognizing 20 aa, as well as several translation factors and elongation factors. At the genome scale, evolutionary analyses placed this virus in the Klosneuvirinae group of giant viruses. Rhizome analysis demonstrated that the genome of Yasminevirus is mosaic, with â¼34% of genes having their closest homologues in other viruses, followed by â¼13.2% in Eukaryota, â¼7.2% in Bacteria, and less than 1% in Archaea Among giant virus sequences, Yasminevirus shared 87% of viral hits with Klosneuvirinae. This description of Yasminevirus sheds light on the Klosneuvirinae group in a captivating quest to understand the evolution and diversity of giant viruses.IMPORTANCE Yasminevirus is an icosahedral double-stranded DNA virus isolated from sewage water by amoeba coculture. Here its structure and replicative cycle in the amoeba Vermamoeba vermiformis are described and genomic and evolutionary studies are reported. This virus belongs to the Klosneuvirinae group of giant viruses, representing the second isolated and cultivated giant virus in this group, and is the first isolated using a coculture procedure. Extended translational machinery pointed to Yasminevirus among the quasiautonomous giant viruses with the most complete translational apparatus of the known virosphere.
Assuntos
DNA Viral/genética , Regulação Viral da Expressão Gênica , Genoma Viral , Vírus Gigantes/genética , Mimiviridae/genética , Vírion/genética , Aminoácidos/genética , Aminoácidos/metabolismo , Aminoacil-tRNA Sintetases/classificação , Aminoacil-tRNA Sintetases/genética , Aminoacil-tRNA Sintetases/metabolismo , Composição de Bases , Mapeamento Cromossômico , Técnicas de Cocultura , Códon/química , Códon/metabolismo , DNA Viral/metabolismo , Tamanho do Genoma , Vírus Gigantes/classificação , Vírus Gigantes/metabolismo , Vírus Gigantes/ultraestrutura , Hartmannella/virologia , Mimiviridae/classificação , Mimiviridae/metabolismo , Mimiviridae/ultraestrutura , Fatores de Alongamento de Peptídeos/classificação , Fatores de Alongamento de Peptídeos/genética , Fatores de Alongamento de Peptídeos/metabolismo , Filogenia , Biossíntese de Proteínas , RNA de Transferência/classificação , RNA de Transferência/genética , RNA de Transferência/metabolismo , Análise de Sequência de DNA , Vírion/metabolismo , Vírion/ultraestruturaRESUMO
The last decade has been marked by two eminent discoveries that have changed our perception of the virology field: The discovery of giant viruses and a distinct new class of viral agents that parasitize their viral factories, the virophages. Coculture and metagenomics have actively contributed to the expansion of the virophage family by isolating dozens of new members. This increase in the body of data on virophage not only revealed the diversity of the virophage group, but also the relevant ecological impact of these small viruses and their potential role in the dynamics of the microbial network. In addition, the isolation of virophages has led us to discover previously unknown features displayed by their host viruses and cells. In this review, we present an update of all the knowledge on the isolation, biology, genomics, and morphological features of the virophages, a decade after the discovery of their first member, the Sputnik virophage. We discuss their parasitic lifestyle as bona fide viruses of the giant virus factories, genetic parasites of their genomes, and then their role as a key component or target for some host defense mechanisms during the tripartite virophage-giant virus-host cell interaction. We also present the latest advances regarding their origin, classification, and definition that have been widely discussed.
Assuntos
Vírus Gigantes/fisiologia , Virófagos/fisiologia , Animais , Evolução Biológica , Genoma Viral , Genômica/métodos , Vírus Gigantes/isolamento & purificação , Vírus Gigantes/ultraestrutura , História do Século XXI , Interações Hospedeiro-Patógeno , Humanos , Sequências Repetitivas Dispersas , Estágios do Ciclo de Vida , Metagenômica/métodos , Pesquisa/história , Virologia/história , Virófagos/classificação , Virófagos/isolamento & purificação , Virófagos/ultraestruturaRESUMO
The history of giant viruses began in 2003 with the identification of Acanthamoeba polyphaga mimivirus. Since then, giant viruses of amoeba enlightened an unknown part of the viral world, and every discovery and characterization of a new giant virus modifies our perception of the virosphere. This notably includes their exceptional virion sizes from 200 nm to 2 µm and their genomic complexity with length, number of genes, and functions such as translational components never seen before. Even more surprising, Mimivirus possesses a unique mobilome composed of virophages, transpovirons, and a defense system against virophages named Mimivirus virophage resistance element (MIMIVIRE). From the discovery and isolation of new giant viruses to their possible roles in humans, this review shows the active contribution of the University Hospital Institute (IHU) Mediterranee Infection to the growing knowledge of the giant viruses' field.
Assuntos
Amoeba/virologia , Pesquisa Biomédica/tendências , Vírus Gigantes/isolamento & purificação , Virologia/tendências , Vírus Gigantes/classificação , Vírus Gigantes/genética , Vírus Gigantes/ultraestrutura , Interações entre Hospedeiro e MicrorganismosRESUMO
Although the nucleocytoplasmic large DNA viruses (NCLDVs) are one of the largest group of viruses that infect many eukaryotic hosts, the near-atomic resolution structures of these viruses have remained unknown. Here we describe a 3.5 Å resolution icosahedrally averaged capsid structure of Paramecium bursaria chlorella virus 1 (PBCV-1). This structure consists of 5040 copies of the major capsid protein, 60 copies of the penton protein and 1800 minor capsid proteins of which there are 13 different types. The minor capsid proteins form a hexagonal network below the outer capsid shell, stabilizing the capsid by binding neighboring capsomers together. The size of the viral capsid is determined by a tape-measure, minor capsid protein of which there are 60 copies in the virion. Homologs of the tape-measure protein and some of the other minor capsid proteins exist in other NCLDVs. Thus, a similar capsid assembly pathway might be used by other NCLDVs.
Assuntos
Proteínas do Capsídeo/química , Proteínas do Capsídeo/ultraestrutura , Capsídeo/química , Capsídeo/ultraestrutura , Vírus Gigantes/ultraestrutura , Phycodnaviridae/ultraestrutura , Vírus de DNA/ultraestrutura , Modelos Moleculares , Estrutura Quaternária de Proteína , Homologia de Sequência de Aminoácidos , Proteínas Virais/química , Proteínas Virais/ultraestrutura , Vírion/ultraestrutura , Montagem de VírusRESUMO
Most known giant viruses, i.e., viruses producing giant virions, parasitize amoebae and other unicellular eukaryotes. Although they vary in the level of dependence on host nuclear functions, their virions self-assemble in the host cell's cytoplasm. Here we report the discovery of a new prototype of giant virus infecting epidermal cells of the marine arrow worm Adhesisagitta hispida. Its 1.25 µm-long virions self-assemble and accumulate in the host cell's nucleus. Conventional transmission electron microscopy reveals that the virions have a unique bipartite structure. An ovoid nucleocapsid, situated in a broad "head" end of the virion is surrounded by a thin envelope. The latter extends away from the head to form a voluminous conical "tail" filled with electron-dense extracapsidular material. The 31nm-thick capsid wall has a distinctive substructure resulting from a patterned arrangement of subunits; it bears no ultrastructural resemblance to the virion walls of other known giant viruses. The envelope self-assembles coincident with the capsid and remotely from all host membranes. We postulate that transmission to new hosts occurs by rupture of protruding virion-filled nuclei when infected arrow worms mate. Future genomic work is needed to determine the phylogenetic position of this new virus, which we have provisionally named Meelsvirus.
Assuntos
Vírus Gigantes/ultraestrutura , Animais , Núcleo Celular/ultraestrutura , Núcleo Celular/virologia , Vírus Gigantes/isolamento & purificação , Vírus Gigantes/patogenicidade , Interações entre Hospedeiro e Microrganismos , Microscopia Eletrônica de Transmissão , Filogenia , Vírion/ultraestrutura , Montagem de Vírus , Zooplâncton/ultraestrutura , Zooplâncton/virologiaRESUMO
BACKGROUND: Since the discovery of giant viruses infecting amoebae in 2003, many dogmas of virology have been revised and the search for these viruses has been intensified. Over the last few years, several new groups of these viruses have been discovered in various types of samples and environments.In this work, we describe the isolation of 68 giant viruses of amoeba obtained from environmental samples from Brazil and Antarctica. METHODS: Isolated viruses were identified by hemacolor staining, PCR assays and electron microscopy (scanning and/or transmission). RESULTS: A total of 64 viruses belonging to the Mimiviridae family were isolated (26 from lineage A, 13 from lineage B, 2 from lineage C and 23 from unidentified lineages) from different types of samples, including marine water from Antarctica, thus being the first mimiviruses isolated in this extreme environment to date. Furthermore, a marseillevirus was isolated from sewage samples along with two pandoraviruses and a cedratvirus (the third to be isolated in the world so far). CONCLUSIONS: Considering the different type of samples, we found a higher number of viral groups in sewage samples. Our results reinforce the importance of prospective studies in different environmental samples, therefore improving our comprehension about the circulation anddiversity of these viruses in nature.
Assuntos
Microbiologia Ambiental , Vírus Gigantes/genética , Vírus Gigantes/isolamento & purificação , Amoeba , Animais , Regiões Antárticas , Brasil , DNA Viral , Genoma Viral , Geografia , Vírus Gigantes/classificação , Vírus Gigantes/ultraestrutura , Humanos , Filogenia , Análise de Sequência de DNARESUMO
Whereas the protein composition and overall shape of several giant virus capsids have been described, the mechanism by which these large capsids assemble remains enigmatic. Here, we present a reconstruction of the capsid of Cafeteria roenbergensis virus (CroV), one of the largest viruses analyzed by cryo-electron microscopy (cryo-EM) to date. The CroV capsid has a diameter of 3,000 Å and a Triangulation number of 499. Unlike related mimiviruses, the CroV capsid is not decorated with glycosylated surface fibers, but features 30 Å-long surface protrusions that are formed by loops of the major capsid protein. Based on the orientation of capsomers in the cryo-EM reconstruction, we propose that the capsids of CroV and related giant viruses are assembled by a newly conceived assembly pathway that initiates at a five-fold vertex and continuously proceeds outwards in a spiraling fashion.
Assuntos
Capsídeo/ultraestrutura , Microscopia Crioeletrônica , Vírus Gigantes/fisiologia , Vírus Gigantes/ultraestrutura , Mimiviridae/fisiologia , Mimiviridae/ultraestrutura , Montagem de Vírus/fisiologia , Sequência de Aminoácidos , Proteínas do Capsídeo/química , Proteínas do Capsídeo/metabolismo , Genoma Viral , Vírus Gigantes/genética , Mimiviridae/genética , Vírion/ultraestruturaRESUMO
Amoeba-infecting viruses have raised scientists' interest due to their novel particle morphologies, their large genome size and their genomic content challenging previously established dogma. We report here the discovery and the characterization of Cedratvirus lausannensis, a novel member of the Megavirales, with a 0.75-1 µm long amphora-shaped particle closed by two striped plugs. Among numerous host cell types tested, the virus replicates only in Acanthamoeba castellanii leading to host cell lysis within 24 h. C. lausannensis was resistant to ethanol, hydrogen peroxide and heating treatments. Like 30 000-year-old Pithovirus sibericum, C. lausannensis enters by phagocytosis, releases its genetic content by fusion of the internal membrane with the inclusion membrane and replicates in intracytoplasmic viral factories. The genome encodes 643 proteins that confirmed the grouping of C. lausannensis with Cedratvirus A11 as phylogenetically distant members of the family Pithoviridae. The 575,161 bp AT-rich genome is essentially devoid of the numerous repeats harbored by Pithovirus, suggesting that these non-coding repetitions might be due to a selfish element rather than particular characteristics of the Pithoviridae family. The discovery of C. lausannensis confirms the contemporary worldwide distribution of Pithoviridae members and the characterization of its genome paves the way to better understand their evolution.
Assuntos
Vírus de DNA/classificação , Vírus Gigantes/classificação , Acanthamoeba castellanii/virologia , Vírus de DNA/genética , Vírus de DNA/ultraestrutura , Variação Genética , Genoma Viral , Vírus Gigantes/genética , Vírus Gigantes/ultraestrutura , FilogeniaRESUMO
The accidental discovery of the giant virus of amoeba - Acanthamoeba polyphaga mimivirus (APMV; more commonly known as mimivirus) - in 2003 changed the field of virology. Viruses were previously defined by their submicroscopic size, which probably prevented the search for giant viruses, which are visible by light microscopy. Extended studies of giant viruses of amoebae revealed that they have genetic, proteomic and structural complexities that were not thought to exist among viruses and that are comparable to those of bacteria, archaea and small eukaryotes. The giant virus particles contain mRNA and more than 100 proteins, they have gene repertoires that are broader than those of other viruses and, notably, some encode translation components. The infection cycles of giant viruses of amoebae involve virus entry by amoebal phagocytosis and replication in viral factories. In addition, mimiviruses are infected by virophages, defend against them through the mimivirus virophage resistance element (MIMIVIRE) system and have a unique mobilome. Overall, giant viruses of amoebae, including mimiviruses, marseilleviruses, pandoraviruses, pithoviruses, faustoviruses and molliviruses, challenge the definition and classification of viruses, and have increasingly been detected in humans.