RESUMO
Mimiviruses are giant viruses of amoeba that can be found in association with virophages. These satellite-like viruses are dependent on the mimivirus viral factory to replicate. Mimiviruses can also be associated with linear DNA molecules called transpovirons. Transpovirons and virophages are important drivers of giant virus evolution although they are still poorly studied elements. Here, we describe the isolation and genomic characterization of a mimivirus/virophage/transpoviron tripartite system from Brazil. We analyzed transmission electron microscopy images and performed genome sequencing and assembly, gene annotation, and phylogenetic analysis. Our data confirm the isolation of a lineage A mimivirus (1.2 Mb/1012 ORFs), called mimivirus argentum, and a sputnik virophage (18,880 bp/20 ORFs). We also detected a third sequence corresponding to a transpoviron from clade A (6365 bp/6 ORFs) that presents small terminal inverted repeats (77 nt). The main genomic features of mimivirus argentum and of its virophage/transpoviron elements corroborates with what is described for other known elements. This highlights that this triple genomic and biological interaction may be ancient and well-conserved. The results expand the basic knowledge about unique and little-known elements and pave the way to future studies that might contribute to a better understanding of this tripartite relationship.
Assuntos
Elementos de DNA Transponíveis , Evolução Molecular , Vírus Gigantes/genética , Mimiviridae/genética , Virófagos/genética , Brasil , Genoma Viral , Genômica , Vírus Gigantes/classificação , Mimiviridae/classificação , Fases de Leitura Aberta , Filogenia , Proteínas Virais/genética , Virófagos/classificaçãoRESUMO
Since its discovery, the first identified giant virus associated with amoebae, Acanthamoeba polyphaga mimivirus (APMV), has been rigorously studied to understand the structural and genomic complexity of this virus. In this work, we report the isolation and genomic characterization of a new mimivirus of lineage B, named "Borely moumouvirus". This new virus exhibits a structure and replicative cycle similar to those of other members of the family Mimiviridae. The genome of the new isolate is a linear double-strand DNA molecule of ~1.0 Mb, containing over 900 open reading frames. Genome annotation highlighted different translation system components encoded in the DNA of Borely moumouvirus, including aminoacyl-tRNA synthetases, translation factors, and tRNA molecules, in a distribution similar to that in other lineage B mimiviruses. Pan-genome analysis indicated an increase in the genetic arsenal of this group of viruses, showing that the family Mimiviridae is still expanding. Furthermore, phylogenetic analysis has shown that Borely moumouvirus is closely related to moumouvirus australiensis. This is the first mimivirus lineage B isolated from Brazilian territory to be characterized. Further prospecting studies are necessary for us to better understand the diversity of these viruses so a better classification system can be established.
Assuntos
Genoma Viral , Mimiviridae/isolamento & purificação , Rios/virologia , Brasil , Genômica , Mimiviridae/classificação , Mimiviridae/genética , Mimiviridae/fisiologia , Filogenia , Replicação ViralRESUMO
The inclusion of Mimiviridae members in the putative monophyletic nucleocytoplasmic large DNA virus (NCLDV) group is based on genomic and phylogenomic patterns. This shows that, along with other viral families, they share a set of genes known as core or "hallmark genes," including the gene for the major capsid protein (MCP). Although previous studies have suggested that the maturation of mimivirus MCP transcripts is dependent on splicing, there is little information about the processing of this transcript in other mimivirus isolates. Here we report the characterization of a new mimivirus isolate, called Kroon virus (KV) mimivirus. Analysis of the structure, synteny, and phylogenetic relationships of the MCP genes in many mimivirus isolates revealed a remarkable variation at position and types of intronic and exonic regions, even for mimiviruses belonging to the same lineage. In addition, sequencing of KV and Acanthamoeba polyphaga mimivirus (APMV) MCP transcripts has shown that inside the family, even related giant viruses may present different ways to process the MCP mRNA. These results contribute to the understanding of the genetic organization and evolution of the MCP gene in mimiviruses.IMPORTANCE Mimivirus isolates have been obtained by prospecting studies since 2003. Based on genomic and phylogenomic studies of conserved genes, these viruses have been clustered together with members of six other viral families. Although the major capsid protein (MCP) gene is an important member of the so-called "hallmark genes," there is little information about the processing and structure of this gene in many mimivirus isolates. In this work, we have analyzed the structure, synteny, and phylogenetic relationships of the MCP genes in many mimivirus isolates; these genes showed remarkable variation at position and types of intronic and exonic regions, even for mimiviruses belonging to the same lineage. These results contribute to the understanding of the genetic organization and evolution of the MCP gene in mimiviruses.
Assuntos
Proteínas do Capsídeo/genética , Evolução Molecular , Regulação Viral da Expressão Gênica , Mimiviridae/genética , Splicing de RNA , Transcrição Gênica , Genoma Viral , Mimiviridae/classificação , Mimiviridae/isolamento & purificação , Mimiviridae/ultraestrutura , Filogenia , RNA Viral , Replicação Viral , Microbiologia da ÁguaRESUMO
The natural history of mimiviruses (i.e., viruses that are members of the Mimivirus genus) is a challenge for modern biology. A new domain of life to include these organisms has been proposed from analysis of gene conservation. We analyzed the evolutionary relationship of proteins involved in the translation system, and our data show that mimiviruses are a sister group of Eukarya. New data about the origins of Eukarya, in which Eukarya appears as a branch derived from the Archaea domain, were discussed, and we suggest that the mimiviruses emerged from the initial population that gave origin to Eukarya and that, in this way are not part of a new domain of life.
Assuntos
Evolução Molecular , Mimiviridae/genética , Filogenia , Proteínas Virais/genética , Mimiviridae/classificação , Modelos Genéticos , Biossíntese de Proteínas/genética , Homologia de Sequência , Proteínas Virais/metabolismoRESUMO
Since the recent discovery of Samba virus, the first representative of the family Mimiviridae from Brazil, prospecting for mimiviruses has been conducted in different environmental conditions in Brazil. Recently, we isolated using Acanthamoeba sp. three new mimiviruses, all of lineage A of amoebal mimiviruses: Kroon virus from urban lake water; Amazonia virus from the Brazilian Amazon river; and Oyster virus from farmed oysters. The aims of this work were to sequence and analyze the genome of these new Brazilian mimiviruses (mimi-BR) and update the analysis of the Samba virus genome. The genomes of Samba virus, Amazonia virus and Oyster virus were 97%-99% similar, whereas Kroon virus had a low similarity (90%-91%) with other mimi-BR. A total of 3877 proteins encoded by mimi-BR were grouped into 974 orthologous clusters. In addition, we identified three new ORFans in the Kroon virus genome. Additional work is needed to expand our knowledge of the diversity of mimiviruses from Brazil, including if and why among amoebal mimiviruses those of lineage A predominate in the Brazilian environment.
Assuntos
Água Doce/virologia , Genoma Viral , Mimiviridae/genética , Sequência de Bases , Brasil , Mimiviridae/química , Mimiviridae/classificação , Mimiviridae/isolamento & purificação , Dados de Sequência Molecular , Filogenia , Alinhamento de SequênciaRESUMO
BACKGROUND: The identification of novel giant viruses from the nucleocytoplasmic large DNA viruses group and their virophages has increased in the last decade and has helped to shed light on viral evolution. This study describe the discovery, isolation and characterization of Samba virus (SMBV), a novel giant virus belonging to the Mimivirus genus, which was isolated from the Negro River in the Brazilian Amazon. We also report the isolation of an SMBV-associated virophage named Rio Negro (RNV), which is the first Mimivirus virophage to be isolated in the Americas. METHODS/RESULTS: Based on a phylogenetic analysis, SMBV belongs to group A of the putative Megavirales order, possibly a new virus related to Acanthamoeba polyphaga mimivirus (APMV). SMBV is the largest virus isolated in Brazil, with an average particle diameter about 574 nm. The SMBV genome contains 938 ORFs, of which nine are ORFans. The 1,213.6 kb SMBV genome is one of the largest genome of any group A Mimivirus described to date. Electron microscopy showed RNV particle accumulation near SMBV and APMV factories resulting in the production of defective SMBV and APMV particles and decreasing the infectivity of these two viruses by several logs. CONCLUSION: This discovery expands our knowledge of Mimiviridae evolution and ecology.
Assuntos
Mimiviridae/isolamento & purificação , Filogenia , Rios/virologia , Brasil , DNA Viral/química , DNA Viral/genética , Microscopia Eletrônica de Transmissão , Mimiviridae/classificação , Mimiviridae/genética , Mimiviridae/ultraestrutura , Dados de Sequência Molecular , Fases de Leitura Aberta , Floresta Úmida , Análise de Sequência de DNA , Vírion/ultraestruturaRESUMO
Acanthamoeba is a free-living protozoan widely distributed in the environment, occurring in vegetative trophozoite and resistance cyst stages during its life cycle. It constitutes an etiological agent of Acanthamoeba keratitis, a disease that may cause severe ocular inflammation and blindness. New drugs can be developed from molecules found in plants and thus help in its difficult treatment. Acanthospermum australe (Asteraceae), a plant used in folk medicine, had its effect tested on Acanthamoeba polyphaga. Aqueous and ethanolic extracts of A. austral were obtained from aerial parts for infusion and static maceration, respectively. Concentrations of 10, 5, 2.5, 1.25 and 0.625 mg/ml of the extract were tested against Acanthamoeba polyphaga trophozoites. The cytotoxic effect of the extracts was tested in mammalian cells using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. RESULTS: The 10 mg/ml concentration of ethanolic extract was lethal to 100% of the A. polyphaga trophozoites in 24 h and both extracts presented cytotoxic effect against mammalian cells. These findings suggest that the A. austral ethanolic extract may have compounds with relevance to the development of new amoebicidal drugs.
Acanthamoeba é um protozoário de vida livre amplamente distribuído no ambiente, ocorrendo sob a forma trofozoítica (metabolicamente ativa) e cística (de resistência), durante seu ciclo de vida. O protozoário constitui um agente etiológico da Ceratite Amebiana, uma doença que pode causar inflamação ocular severa e cegueira. Novos fármacos podem ser desenvolvidos a partir de moléculas encontradas em plantas e assim ajudar em seu difícil tratamento. Aqui, Acanthospermum australe (Asteraceae), uma planta utilizada na medicina popular, teve seu efeito sobre trofozoítos de Acanthamoeba polyphaga testado. O extrato aquoso e etanólico de A. australe foram obtidos das partes aéreas por infusão e maceração estática, respectivamente. As concentrações 10, 5, 2,5, 1,25 e 0,625 mg/ml dos extratos foram testadas contra trofozoítos do protozoário. O efeito citotóxico dos extratos foi testado em células de mamífero utilizando o ensaio de brometo de 3-[4,5-dimetiltiazol-2-il]-2,5-difeniltetrazólio (MTT). A concentração de 10 mg/ml do extrato etanólico foi letal a 100% dos trofozoítos de A. polyphaga em 24 h e ambos os extratos apresentaram efeito citotóxico contra as células de mamífero. Estes resultados sugerem que o extrato etanólico de A. australe pode ter componentes com relevância para o desenvolvimento de novos fármacos amebicidas.