Search

Giant viruses of amoebas are a remarkable group of viruses. In addition to their large size and peculiar structures, the genetic content of these viruses is also special. Among the genetic features of these viruses that stand out is the presence of coding regions for elements involved in translation, a complex biological process that occurs in cellular organisms. No viral genome described so far has such a complex genetic arsenal as those of giant viruses, which code for several of these elements. Currently, tupanviruses have the most complete set of translation genes in the known virosphere. In this review, we have condensed what is currently known about translation genes in different groups of giant viruses and theorize about their biological importance, origin, and evolution, and what might possibly be found in the coming years.

International audience; The discovery of giant viruses in the last years has fascinated the scientific community due to virus particles size and genome complexity. Among such fantastic discoveries, we have recently described tupanviruses, which particles present a long tail, and has a genome that contains the most complete set of translation-related genes ever reported in the known virosphere. Here we describe a new kind of virus-host interaction involving tupanvirus. We observed that tupanvirus-infected amoebas were induced to aggregate with uninfected cells, promoting viral dissemination and forming giant host cell bunches. Even after mechanical breakdown of bunches, amoebas reaggregated within a few minutes. This remarkable interaction between infected and uninfected cells seems to be promoted by the expression of a mannose receptor gene. Our investigations demonstrate that the pre-treatment of amoebas with free mannose inhibits the formation of bunches, in a concentration-dependent manner, suggesting that amoebal-bunch formation correlates with mannose receptor gene expression. Finally, our data suggest that bunch-forming cells are able to interact with uninfected cells promoting the dissemination and increase of tupanvirus progeny. The recent discovery of tupanvirus, one of the largest and most complex viruses isolated to date, has reinforced the structural and genomic complexity of the giant viruses 1. Tupanviruses have been isolated from soda lakes, known as an extreme aquatic environments, and from ocean sediments collected at a depth of 3000 meters (m) 1,2. Phylogenetic analyses have shown the clustering of the tupanvirus with members of the family Mimiviridae. However, there are many peculiarities that make the tupanviruses unique entities in the known virosphere. Since its first observation, tupanviruses showed remarkable morphological characteristics; it has optically visible particles that average about 1.2 µm in size and can reach lengths up to 2.3 µm 1. Tupanvirus has the largest host range described so far among amoebal-infecting giant viruses and can causes a shutdown of host rRNA that is likely related to host-nucleolus degradation 1,3-9. The tupanviruses replication cycle is similar to those for other mim-iviruses, in which viral particles attach to the host-cell surface and enter through phagocytosis. The viral inner membrane then fuses with the phagosome membrane, releasing the genome. A viral factory (VF) is formed, where particle morphogenesis occurs; the cycle ends with cell lysis and the release of progeny viruses 1,10. The study of the tupanvirus genome further aroused the interest of virologists, not only due of its large size (~1.5 Mb), but also because these viruses show the largest translational apparatus described. It is composed of up to 70 tRNA, 20 aminoacyl-tRNA synthetases (aaRS), 11 factors associated with translation, and factors related to tRNA/mRNA maturation and ribosome protein modification 1. In addition to the robust translation apparatus, tupanvirus also contains a gene encoding mannose-specific lectin, also called mannose-binding protein (MBP) 1. Interestingly, previous studies revealed that Acanthamoeba castellanii expresses an MBP and that free-mannose can inhibit the adhesion of A. castellanii to surfaces, suggesting that the MBP plays a role in the pathogenesis of Acanthamoeba infection 11-16 .

The Acanthamoeba polyphaga mimivirus (APMV) was first isolated during a pneumonia outbreak in Bradford, England, and since its discovery many research groups devoted efforts to understand whether this virus could be associated to human diseases, in particular clinical signs and symptoms of pneumonia. In 2013, we observed cytopathic effect in amoebas (rounding and lysis) inoculated with APMV inoculated PBMCs (peripheral blood mononuclear cell) extracts, and at that point we interpreted those results as mimivirus replication in human PBMCs. Based on these results we decided to further investigate APMV replication in human PBMCs, by transmission electron microscopy (TEM) and qPCR. No viral factory was observed in APMV inoculated PBMCs, at any analyzed time and M.O.I.s (multiplicity of infection), by checking 550 cells per condition tested. We also measured the variation of viral DNA by qPCR targeting helicase gene during the course of the TEM experiment in PBMCs, but the DNA levels stayed the same as the first time-point post infection. In summary, our newest qPCR and TEM results do not support previous statements (including ours) that mimivirus is able to replicate in humans PBMCs. nonPeerReviewed

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.

International audience; In recent years, giant viruses belonging to the family Mimiviridae have been proposed to be infectious agents in humans. In this work we provide evidence of mimivirus genome and neutralizing antibodies detection in humans.

Viruses are ubiquitous organisms, but their role in the ecosystem and their prevalence are still poorly understood. Mimiviruses are extremely complex and large DNA viruses. Although metagenomic studies have suggested that members of the family Mimiviridae are abundant in oceans, there is a lack of information about the association of mimiviruses with marine organisms. In this work, we demonstrate by molecular and virological methods that oysters are excellent sources for mimiviruses isolation. Our data not only provide new information about the biology of these viruses but also raise questions regarding the role of oyster consumption as a putative source of mimivirus infection in humans.

To investigate circulation of mimiviruses in the Amazon Region of Brazil, we surveyed 513 serum samples from domestic and wild mammals. Neutralizing antibodies were detected in 15 sample pools, and mimivirus DNA was detected in 9 pools of serum from capuchin monkeys and in 16 pools of serum from cattle.

International audience; In this review we discuss the role of mimiviruses as potential human pathogens focusing on clinical and evolutionary evidence. We also propose a novel antiviral immunomodulatory pathway controlled by interferon-beta (IFN-beta) and mediated by immune-responsive gene 1 (IRG1) and itaconic acid, its product. Acanthamoeba polyphaga Mimivirus (APMV) was isolated from amoebae in a hospital while investigating a pneumonia outbreak. Mimivirus ubiquity and role as protist pathogens are well understood, and its putative status as a human pathogen has been gaining strength as more evidence is being found. The study of APMV and human cells interaction revealed that the virus is able to evade the IFN system by inhibiting the regulation of interferon-stimulated genes, suggesting that the virus and humans have had host-pathogen interactions. It also has shown that the virus is capable of growing on IFN-alpha 2, but not on IFN-beta-treated cells, hinting at an exclusive IFN-beta antiviral pathway. Our hypothesis based on preliminary data and published articles is that IFN-beta preferentially upregulates IRG1 in human macrophagic cells, which in turn produces itaconic acid. This metabolite links metabolism to antiviral activity by inactivating the virus, in a novel immunomodulatory pathway relevant for APMV infections and probably to other infectious diseases as well.

The aim of this protocol is to describe the replication, purification, and titration of mimiviruses. These viruses belong to the Mimiviridae family, the first member of which was isolated in 1992 from a cooling tower water sample collected during an outbreak of pneumonia in a hospital in Bradford, England. In recent years, several new mimiviruses have been isolated from different environmental conditions. These giant viruses are easily replicated in amoeba of the Acanthamoeba genus, its natural host. Mimiviruses present peculiar features that make them unique viruses, such as the particle and genome size and the genome's complexity. The discovery of these viruses rekindled discussions about their origin and evolution, and the genetic and structural complexity opened up a new field of study. Here, we describe some methods utilized for mimiviruses replication, purification, and titration. © 2016 by John Wiley & Sons, Inc.

It is well recognized that gene duplication/acquisition is a key factor for molecular evolution, being directly related to the emergence of new genetic variants. The importance of such phenomena can also be expanded to the viral world, with impacts on viral fitness and environmental adaptations. In this work we describe the isolation and characterization of Niemeyer virus, a new mimivirus isolate obtained from water samples of an urban lake in Brazil. Genomic data showed that Niemeyer harbors duplicated copies of three of its four aminoacyl-tRNA synthetase genes (cysteinyl, methionyl, and tyrosyl RS). Gene expression analysis showed that such duplications allowed significantly increased expression of methionyl and tyrosyl aaRS mRNA by Niemeyer in comparison to APMV. Remarkably, phylogenetic data revealed that Niemeyer duplicated gene pairs are different, each one clustering with a different group of mimivirus strains. Taken together, our results raise new questions about the origins and selective pressures involving events of aaRS gain and loss among mimiviruses.

International audience; The complexity of giant virus genomes is intriguing, especially the presence of genes encoding components of the protein translation machinery such as transfer RNAs and aminoacyl-tRNA-synthetases; these features are uncommon among other viruses. Although orthologs of these genes are codified by their hosts, one can hypothesize that having these translation-related genes might represent a gain of fitness during infection. Therefore, the aim of this study was to evaluate the expression of translation-related genes by mimivirus during infection of Acanthamoeba castellanii under different nutritional conditions. In silico analysis of amino acid usage revealed remarkable differences between the mimivirus isolates and the A. castellanii host. Relative expression analysis by quantitative PCR revealed that mimivirus was able to modulate the expression of eight viral translation-related genes according to the amoebal growth condition, with a higher induction of gene expression under starvation. Some mimivirus isolates presented differences in translation-related gene expression; notably, polymorphisms in the promoter regions correlated with these differences. Two mimivirus isolates did not encode the tryptophanyl-tRNA in their genomes, which may be linked with low conservation pressure based on amino acid usage analysis. Taken together, our data suggest that mimivirus can modulate the expression of translation-related genes in response to nutrient availability in the host cell, allowing the mimivirus to adapt to different hosts growing under different nutritional conditions.

Acanthamoeba polyphaga mimivirus (APMV) was described in 2003, and due to its unique structural and genetic complexity, the viral family Mimiviridae was created. APMV prompted the creation of an open field of study on the function of hundreds of never-before-seen open reading frames (ORFs) and their roles in virus-host interactions. In recent years, several giant viruses have been isolated from different environments and specimens. Although the scientific community has experienced a remarkable advancement in the comprehension of the mimivirus replication cycle in the last years, few studies have been devoted to the investigation of the methodological features and conditions for mimivirus cultivation. In this work, conditions for the cultivation of mimivirus isolates were investigated to obtain relevant information about the production of infectious particles, total viral particles and viral DNA. The results suggest that low viral doses are more efficient for the production of infectious particles, yielding up to 5000 TCID50 for each inoculated TCID50. Besides methodological information, these data also reveal, for the first time, the ratio between total and infectious particles (in TCID50) that are produced during mimivirus cultivation in laboratory conditions. All of this information can be used as a worldwide guide for the production of mimiviruses and can help prompt mimivirological studies in different fields.

Amoebas of the genus Acanthamoeba are protists that are associated with human disease and represent a public health concern. They can harbor pathogenic microorganisms, acting as a platform for pathogen replication. Acanthamoeba polyphaga mimivirus (APMV), the type species of the genus Mimivirus, family Mimiviridae, represents the largest group of amoeba-associated viruses that has been described to date. Recent studies have demonstrated that APMV and other giant viruses may cause pneumonia. Amoebas can survive in most environments and tolerate various adverse conditions, including UV light irradiation, high concentrations of disinfectants, and a broad range of temperatures. However, it is unknown how the amoebal intracellular environment influences APMV stability and resistance to adverse conditions. Therefore, in this work, we evaluated the stability of APMV, either purified or carried by the amoeba host, under extreme conditions, including UV irradiation, heat and exposure to six different chemical biocides. After each treatment, the virus was titrated in amoebas using the TCID50 method. APMV was more stable in all resistance tests performed when located inside its host. Our results demonstrate that Acanthamoeba acts as a natural bunker for APMV, increasing viral resistance to extreme physical and chemical conditions. The data raise new questions regarding the survival of APMV in nature and in hospital environments.

Viruses are extremely diverse and abundant and are present in countless environments. Giant viruses of the Megavirales order have emerged as a fascinating research topic for virologists around the world. As evidence of their ubiquity and ecological impact, mimiviruses have been found in multiple environmental samples. However, isolation of these viruses from environmental samples is inefficient, mainly due to methodological limitations and lack of information regarding the interactions between viruses and substrates. In this work, we demonstrate the long-lasting stability of mimivirus in environmental (freshwater and saline water) and hospital (ventilator plastic device tube) substrates, showing the detection of infectious particles after more than 9 months. In addition, an enrichment protocol was implemented that remarkably increased mimivirus detection from all tested substrates, including field tests. Moreover, biological, morphological and genetic tests revealed that the enrichment protocol maintained mimivirus particle integrity. In conclusion, our work demonstrated the stability of APMV in samples of environmental and health interest and proposed a reliable and easy protocol to improve giant virus isolation. The data presented here can guide future giant virus detection and isolation studies.

Acanthamoeba polyphaga mimivirus (APMV) is a giant, double-stranded virus of the Mimiviridae family that was discovered in 2003. Recent studies have shown that this virus is able to replicate in murine and human phagocytes and might be considered a putative human pathogen that causes pneumonia. However, there is little data regarding APMV and its host defense relationship. In the present study, we investigated how some components of the interferon (IFN) system are stimulated by APMV in human peripheral blood mononuclear cells (PBMCs) and how APMV replication is affected by IFN treatment. Our results demonstrated that APMV is able to replicate in human PBMCs, inducing type I Interferons (IFNs) but inhibiting interferon stimulated genes (ISG) induction by viroceptor and STAT-1 and STAT-2 dephosphorylation independent mechanisms. We also showed that APMV is resistant to the antiviral action of interferon-alpha2 (IFNA2) but is sensitive to the antiviral action of interferon-beta (IFNB1). Our results demonstrated the productive infection of professional phagocytes with APMV and showed that this virus is recognized by the immune system of vertebrates and inhibits it. It provides the first data regarding APMV and the IFN system interaction and raise new and relevant evolutional questions about the relationship between APMV and vertebrate hosts.

In 2003, Acanthamoeba polyphaga mimivirus (APMV) was first described and began to impact researchers around the world, due to its structural and genetic complexity. This virus founded the family Mimiviridae. In recent years, several new giant viruses have been isolated from different environments and specimens. Giant virus research is in its initial phase and information that may arise in the coming years may change current conceptions of life, diversity and evolution. Thus, this review aims to condense the studies conducted so far about the features and peculiarities of APMV, from its discovery to its clinical relevance.

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.