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Marseilleviruses (MsV) are a group of viruses that compose the Marseilleviridae family within the Nucleocytoviricota phylum. They have been found in different samples, mainly in freshwater. MsV are classically organized into five phylogenetic lineages (A/B/C/D/E), but the current taxonomy does not fully represent all the diversity of the MsV lineages. Here, we describe a novel strain isolated from a Brazilian saltwater sample named Marseillevirus cajuinensis. Based on genomics and phylogenetic analyses, M. cajuinensis exhibits a 380,653-bp genome that encodes 515 open reading frames. Additionally, M. cajuinensis encodes a transfer RNA, a feature that is rarely described for Marseilleviridae. Phylogeny suggests that M. cajuinensis forms a divergent branch within the MsV lineage A. Furthermore, our analysis suggests that the common ancestor for the five classical lineages of MsV diversified into three major groups. The organization of MsV into three main groups is reinforced by a comprehensive analysis of clusters of orthologous groups, sequence identities, and evolutionary distances considering several MsV isolates. Taken together, our results highlight the importance of discovering new viruses to expand the knowledge about known viruses that belong to the same lineages or families. This work proposes a new perspective on the Marseilleviridae lineages organization that could be helpful to a future update in the taxonomy of the Marseilleviridae family., Importance: Marseilleviridae is a family of viruses whose members were mostly isolated from freshwater samples. In this work, we describe the first Marseillevirus isolated from saltwater samples, which we called Marseillevirus cajuinensis . Most of M. cajuinensis genomic features are comparable to other Marseilleviridae members, such as its high number of unknown proteins. On the other hand, M. cajuinensis encodes a transfer RNA, which is a gene category involved in protein translation that is rarely described in this viral family. Additionally, our phylogenetic analyses suggested the existence of, at least, three major Marseilleviridae groups. These observations provide a new perspective on Marseilleviridae lineages organization, which will be valuable in future updates to the taxonomy of the family since the current official classification does not capture all the Marseilleviridae known diversity., Competing Interests: The authors declare no conflict of interest.

The Mimivirus is a giant virus that infects amoebae and was long considered to be a bacterium due to its size. The viral particles are composed of a protein capsid of ~500 nm in diameter, which is enclosed in a polysaccharide layer in which ~120‑140 nm long fibers are embedded, resulting in an overall diameter of 700 nm. The virus has a genome size of 1.2 Mb DNA, and surprisingly, replicates only in the cytoplasm of the infected cells without entering the nucleus, which is a unique characteristic among DNA viruses. Their existence is undeniable; however, as with any novel discovery, there is still uncertainty concerning their pathogenicity mechanisms in humans and the nature of the Mimivirus virophage resistance element system (MIMIVIRE), a term given to describe the immune network of the Mimivirus, which closely resembles the CRISPR‑Cas system. The scope of the present review is to discuss the recent developments derived from structural and functional studies performed on the distinctive characteristics of the Mimivirus, and from studies concerning their putative clinical relevance in humans.

The family Marseilleviridae was the second family of giant viruses that was described in 2013, after the family Mimiviridae . Marseillevirus marseillevirus , isolated in 2007 by coculture on Acanthamoeba polyphaga , is the prototype member of this family. Afterward, the worldwide distribution of marseilleviruses was revealed through their isolation from samples of various types and sources. Thus, 62 were isolated from environmental water, one from soil, one from a dipteran, one from mussels, and two from asymptomatic humans, which led to the description of 67 marseillevirus isolates, including 21 by the IHU Méditerranée Infection in France. Recently, five marseillevirus genomes were assembled from deep sea sediment in Norway. Isolated marseilleviruses have ≈250 nm long icosahedral capsids and 348-404 kilobase long mosaic genomes that encode 386-545 predicted proteins. Comparative genomic analyses indicate that the family Marseilleviridae includes five lineages and possesses a pangenome composed of 3,082 clusters of genes. The detection of marseilleviruses in both symptomatic and asymptomatic humans in stool, blood, and lymph nodes, and an up-to-30-day persistence of marseillevirus in rats and mice, raise questions concerning their possible clinical significance that are still under investigation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Sahmi-Bounsiar, Rolland, Aherfi, Boudjemaa, Levasseur, La Scola and Colson.)

Marseilleviridae is a family of large double-stranded DNA viruses that is currently divided into five subgroups, lineages A-E. Hokutovirus and kashiwazakivirus, both of which belong to lineage B, have been reported to induce host acanthamoeba cells to form aggregations called "bunches". This putatively results in increased opportunities to infect acanthamoeba cells, in contrast to lineage A, which has been reported to not form "bunches". In the present study, we isolated 14 virus strains of the family Marseilleviridae from several Japanese water samples, 11 of which were identified as lineage B viruses. All 11 lineage B strains caused infected amoeba cells to form bunches. We then investigated the involvement of monosaccharides in bunch formation by amoeba cells infected with hokutovirus. Galactose inhibited bunch formation, thereby allowing amoeba cells to delay the process, whereas mannose and glucose did not. A kinetic image analysis of hokutovirus-infected amoeba cells confirmed the inhibition of bunch formation by galactose. The number of hokutovirus-infected amoeba cells increased more rapidly than that of tokyovirus-infected cells, which belongs to lineage A. This result suggests that bunch formation by infected amoeba cells is advantageous for lineage B viruses.

Marseilleviridae is a family of viruses which have only been propagated in acanthamoeba. Marseillevirus sequences have been recently detected in different human matrices by viral metagenomics. Single-center studies worldwide have estimated a low prevalence of marseillevirus both in symptomatic patients and in healthy donors but, to date, no informations are available on the prevalence of this giant virus in Italy. By a polymerase chain reaction targeting the ORF152 viral sequence, we tested sera from 197 immunosuppressed patients and 285 healthy donors, and 63 and 30 respiratory and cerebrospinal fluid samples, respectively, of patients with various clinical conditions and referring the Virology Division for diagnostic purposes. We observed no evidence of Marseillevirus DNA in all 575 samples tested. Marseillevirus probably does not cause infection in human., (© 2019 Wiley Periodicals, Inc.)

Nucleocytoplasmic large DNA viruses (NCLDVs) blur the line between viruses and cells. Melbournevirus (MelV, family Marseilleviridae) belongs to a new family of NCLDVs. Here we present an electron cryo-microscopy structure of the MelV particle, with the large triangulation number T = 309 constructed by 3080 pseudo-hexagonal capsomers. The most distinct feature of the particle is a large and dense body (LDB) consistently found inside all particles. Electron cryo-tomography of 147 particles shows that the LDB is preferentially located in proximity to the probable lipid bilayer. The LDB is 30 nm in size and its density matches that of a genome/protein complex. The observed LDB reinforces the structural complexity of MelV, setting it apart from other NCLDVs., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: While the genomes of eukaryotes and Archaea both encode the histone-fold domain, only eukaryotes encode the core histone paralogs H2A, H2B, H3, and H4. With DNA, these core histones assemble into the nucleosomal octamer underlying eukaryotic chromatin. Importantly, core histones for H2A and H3 are maintained as neofunctionalized paralogs adapted for general bulk chromatin (canonical H2 and H3) or specialized chromatin (H2A.Z enriched at gene promoters and cenH3s enriched at centromeres). In this context, the identification of core histone-like "doublets" in the cytoplasmic replication factories of the Marseilleviridae (MV) is a novel finding with possible relevance to understanding the origin of eukaryotic chromatin. Here, we analyze and compare the core histone doublet genes from all known MV genomes as well as other MV genes relevant to the origin of the eukaryotic replisome., Results: Using different phylogenetic approaches, we show that MV histone domains encode obligate H2B-H2A and H4-H3 dimers of possible proto-eukaryotic origin. MV core histone moieties form sister clades to each of the four eukaryotic clades of canonical and variant core histones. This suggests that MV core histone moieties diverged prior to eukaryotic neofunctionalizations associated with paired linear chromosomes and variant histone octamer assembly. We also show that MV genomes encode a proto-eukaryotic DNA topoisomerase II enzyme that forms a sister clade to eukaryotes. This is a relevant finding given that DNA topo II influences histone deposition and chromatin compaction and is the second most abundant nuclear protein after histones., Conclusions: The combined domain architecture and phylogenomic analyses presented here suggest that a primitive origin for MV histone genes is a more parsimonious explanation than horizontal gene transfers + gene fusions + sufficient divergence to eliminate relatedness to eukaryotic neofunctionalizations within the H2A and H3 clades without loss of relatedness to each of the four core histone clades. We thus suggest MV histone doublet genes and their DNA topo II gene possibly were acquired from an organism with a chromatinized replisome that diverged prior to the origin of eukaryotic core histone variants for H2/H2A.Z and H3/cenH3. These results also imply that core histones were utilized ancestrally in viral DNA compaction and/or protection from host endonucleases.

Viruses display a wide range of genomic profiles and, consequently, a variety of gene expression strategies. Specific sequences associated with transcriptional processes have been described in viruses, and putative promoter motifs have been elucidated for some nucleocytoplasmic large DNA viruses (NCLDV). Among NCLDV, the Marseilleviridae is a well-recognized family because of its genomic mosaicism. The marseilleviruses have an ability to incorporate foreign genes, especially from sympatric organisms inhabiting Acanthamoeba , its main known host. Here, we identified for the first time an eight-nucleotide A/T-rich promoter sequence (AAATATTT) associated with 55% of marseillevirus genes that is conserved in all marseilleviruses lineages, a higher level of conservation than that of any giant virus described to date. We instigated our prediction about the promoter motif by biological assays and by evaluating how single mutations in this octamer can impact gene expression. The investigation of sequences that regulate the expression of genes relative to lateral transfer revealed that the promoter motifs do not appear to be incorporated by marseilleviruses from donor organisms. Indeed, analyses of the intergenic regions that regulate lateral gene transfer-related genes have revealed an independent origin of the marseillevirus intergenic regions that does not match gene-donor organisms. About 50% of AAATATTT motifs spread throughout intergenic regions of the marseilleviruses are present as multiple copies. We believe that such multiple motifs are associated with increased expression of a given gene or are related to incorporation of foreign genes into the mosaic genome of marseilleviruses. IMPORTANCE The marseilleviruses draw attention because of the peculiar features of their genomes; however, little is known about their gene expression patterns or the factors that regulate those expression patterns. The limited published research on the expression patterns of the marseilleviruses and their unique genomes has led us to study the promoter motif sequences in the intergenic regions of the marseilleviruses. This work is the first to analyze promoter sequences in the genomes of the marseilleviruses. We also suggest a strong capacity to acquire foreign genes and to express those genes mediated by multiple copies of the promoter motifs available in intergenic regions. These findings contribute to an understanding of genomic expansion and plasticity observed in these giant viruses., (Copyright © 2017 American Society for Microbiology.)

Amoeba-resisting microorganisms raised a great interest during the last decade. Among them, some large DNA viruses present huge genomes up to 2.5 Mb long, exceeding the size of small bacterial genomes. The rate of genome evolution in terms of mutation, deletion, and gene acquisition in these genomes is yet unknown. Given the suspected high plasticity of viral genomes, the microevolution of the 346 kb genome of Lausannevirus, a member of Megavirales, was studied. Hence, Lausannevirus was co-cultured within the amoeba Acanthamoeba castellanii over one year. Despite a low number of mutations, the virus showed a genome reduction of 3.7% after 12 months. Lausannevirus genome evolution in sympatric conditions was investigated by its co-culture with Estrella lausannensis, an obligate intracellular bacterium, in the amoeba A. castellanii during one year. Cultures were split every 3 months. Genome sequencing revealed that in these conditions both, Lausannevirus and E. lausannensis, show stable genome, presenting no major rearrangement. In fact, after one year they acquired from 2 to 7 and from 4 to 10 mutations per culture for Lausannevirus and E. lausannensis, respectively. Interestingly, different mutations in the endonuclease encoding genes of Lausannevirus were observed in different subcultures, highlighting the importance of this gene product in the replication of Lausannevirus. Conversely, mutations in E. lausannensis were mainly located in a gene encoding for a phosphoenolpyruvate-protein phosphotransferase (PtsI), implicated in sugar metabolism. Moreover, in our conditions and with our analyses we detected no horizontal gene transfer during one year of co-culture., (© The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

In 2003, Acanthamoeba polyphaga mimivirus (APMV) was discovered as parasitizing Acanthamoeba. It was revealed to exhibit remarkable features, especially odd genomic characteristics, and founded viral family Mimiviridae. Subsequently, a second family of giant amoebal viruses was described, Marseilleviridae, whose prototype member is Marseillevirus, discovered in 2009. Currently, the genomes of seven different members of this family have been fully sequenced. Previous phylogenetic analysis suggested the existence of three Marseilleviridae lineages: A, B and C. Here, we describe a new member of this family, Brazilian Marseillevirus (BrMV), which was isolated from a Brazilian sample and whose genome was fully sequenced and analyzed. Surprisingly, data from phylogenetic analyses and comparative genomics, including mean amino acid identity between BrMV and other Marseilleviridae members and the analyses of the core genome and pan-genome of marseilleviruses, indicated that this virus can be assigned to a new Marseilleviridae lineage. Even if the BrMV genome is one of the smallest among Marseilleviridae members, it harbors the second largest gene content into this family. In addition, the BrMV genome encodes 29 ORFans. Here, we describe the isolation and genome analyses of the BrMV strain, and propose its classification as the prototype virus of a new lineage D within the family Marseilleviridae.

The family Marseilleviridae encompasses giant viruses that replicate in free-living Acanthamoeba amoebae. Since the discovery of the founding member Marseillevirus in 2007, 7 new marseilleviruses have been observed, including 3 from environmental freshwater, one from a dipteran, and two from symptom-free humans. Marseilleviruses have ≈250-nm-large icosahedral capsids and 346-386-kb-long mosaic genomes that encode 444-497 predicted proteins. They share a small set of core genes with Mimivirus and other large and giant DNA viruses that compose a monophyletic group, first described in 2001. Comparative genomics analyses indicate that the family Marseilleviridae currently includes three lineages and a pan-genome composed of ≈600 genes. Antibodies against marseilleviruses and viral DNA have been observed in a significant proportion of asymptomatic individuals and in the blood and lymph nodes of a child with adenitis; these observations suggest that these giant viruses may be blood borne and question if they may be pathogenic in humans., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Aim: During the last decade it became obvious that viruses belonging to Mimiviridae and Marseilleviridae families (order Megavirales), may be potential causative agents of pneumonia. Thus, we have performed a review of the association of Mimiviridae, Marseilleviridae, and virophages with pneumonia, particularly healthcare-associated pneumonia, and other infections of the respiratory tract., Results and Discussion: According to the analysis of the published articles, viruses belonging to Mimiviridae family can be potential agents of both community-acquired and healthcare-associated pneumonia. In particular, these viruses may be associated with poor outcome in patients of intensive care units. The exact mechanism of their pathogenicity, however, still remains unclear. The discrepancies between the results obtained by serological and genomic methods could be explained by the high polymorphism of nucleotide sequences of Mimiviridae family representatives. Further investigations on the Mimiviridae pathogenicity and on the determination of Mimiviridae-caused pneumonia risk groups are required. However, the pathogenicity of the viruses belonging to Marseilleviridae family and virophages is unclear up to now.