Isolation and Identification of a Large Green Alga Virus (Chlorella Virus XW01) of Mimiviridae and Its Virophage (Chlorella Virus Virophage SW01) by Using Unicellular Green Algal Cultures.

Virophages are a group of small double-stranded DNA viruses that infect protist hosts and parasitize the viral factory of host giant/large viruses to propagate. Here, we discover a novel cell-virus-virophage (CVv) tripartite interaction system by using unicellular micro-green algae (Chlorella sp.) as eukaryotic hosts for the first time. Viral particles, resembling known virophages and large alga viruses, are detected in culture supernatants and inside algal cells. Complete genomic sequences of the virophage (Chlorella virus virophage SW01 [CVv-SW01]; 24,744 bp) and large virus (Chlorella virus XW01 [CV-XW01]; 407,612 bp) are obtained from the cocultures. Both genomic and phylogenetic analyses show that CVv-SW01 is closely related to virophages previously found in Dishui Lake. CV-XW01 shares the greatest number of homologous genes (n = 82) with Cafeteria roenbergensis virus (CroV) and phylogenetically represents the closest relative to CroV. This is the first report of a large green alga virus being affiliated with a heterotrophic zooplankton-infecting Cafeteriavirus of the family Mimiviridae. Moreover, the codon usage preferences of CV-XW01 and CVv-SW01 are highly similar to those of CroV and its virophage Mavirus, respectively. The discovery of such a novel CVv system with the green alga Chlorella sp. as the single cellular eukaryotic host paves a way to further investigate the potential interaction mechanism of CVv and its significance in the ecology of green algae and the evolution of large/giant viruses and their parasitic viruses. IMPORTANCE Parasitic virophages are small unicellular eukaryotic dsDNA viruses that rely on the viral factories of coinfecting giant/large dsDNA viruses for propagation. Presently, the identified eukaryotic hosts of isolated virophages were restricted to a free-living amoeba, Acanthamoeba polyphaga, and a widespread marine heterotrophic flagellate, Cafeteria roenbergensis. In this study, we successfully discovered and identified a novel tripartite interaction system comprised of a micro-green alga (Chlorella sp.), Mimiviridae large green alga virus, and virophage at the coculture level, with Chlorella sp. as the eukaryotic host, based on combination analysis of infection, morphotype, genome, and phylogeny. The large green alga virus CV-XW01 represents the closest relative to the Mimiviridae giant virus Cafeteria roenbergensis virus, host virus of the virophage Mavirus, as well as a novel large virus of Mimiviridae that infects a non-protozoan protist host. The virophage CVv-SW01 highly resembles Mavirus in its codon usage frequency and preference, although they are phylogenetically distantly related. These findings give novel insights into the diversity of large/giant viruses and their virophages.

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.

Utilizing the VirIdAl Pipeline to Search for Viruses in the Metagenomic Data of Bat Samples.

According to various estimates, only a small percentage of existing viruses have been discovered, naturally much less being represented in the genomic databases. High-throughput sequencing technologies develop rapidly, empowering large-scale screening of various biological samples for the presence of pathogen-associated nucleotide sequences, but many organisms are yet to be attributed specific loci for identification. This problem particularly impedes viral screening, due to vast heterogeneity in viral genomes. In this paper, we present a new bioinformatic pipeline, VirIdAl, for detecting and identifying viral pathogens in sequencing data. We also demonstrate the utility of the new software by applying it to viral screening of the feces of bats collected in the Moscow region, which revealed a significant variety of viruses associated with bats, insects, plants, and protozoa. The presence of alpha and beta coronavirus reads, including the MERS-like bat virus, deserves a special mention, as it once again indicates that bats are indeed reservoirs for many viral pathogens. In addition, it was shown that alignment-based methods were unable to identify the taxon for a large proportion of reads, and we additionally applied other approaches, showing that they can further reveal the presence of viral agents in sequencing data. However, the incompleteness of viral databases remains a significant problem in the studies of viral diversity, and therefore necessitates the use of combined approaches, including those based on machine learning methods.

Revealing the Viral Community in the Hadal Sediment of the New Britain Trench.

Marine viruses are widely distributed and influence matter and energy transformation in ecosystems by modulating hosts' metabolism. The hadal trenches represent the deepest marine habitat on Earth, for which the viral communities and related biogeochemical functions are least explored and poorly understood. Here, using the sediment samples (8720 m below sea level) collected from the New Britain Trench (NBT), we investigated the viral community, diversity, and genetic potentials in the hadal sediment habitat for the first time by deep shotgun metagenomic sequencing. We found the NBT sediment viral community was dominated by Siphoviridae, Myoviridae, Podoviridae, Mimiviridae, and Phycodnaviridae, which belong to the dsDNA viruses. However, the large majority of them remained uncharacterized. We found the hadal sediment virome had some common components by comparing the hadal sediment viruses with those of hadal aquatic habitats and those of bathypelagic and terrestrial habitats. It was also distinctive in community structure and had many novel viral clusters not associated with the other habitual virome included in our analyses. Further phylogenetic analysis on its Caudovirales showed novel diversities, including new clades specially evolved in the hadal sediment habitat. Annotation of the NBT sediment viruses indicated the viruses might influence microbial hydrocarbon biodegradation and carbon and sulfur cycling via metabolic augmentation through auxiliary metabolic genes (AMGs). Our study filled in the knowledge gaps on the virome of the hadal sediment habitats and provided insight into the evolution and the potential metabolic functions of the hadal sediment virome.

Seasonal Dynamics of Algae-Infecting Viruses and Their Inferred Interactions with Protists.

Viruses are a highly abundant, dynamic, and diverse component of planktonic communities that have key roles in marine ecosystems. We aimed to reveal the diversity and dynamics of marine large dsDNA viruses infecting algae in the Northern Skagerrak, South Norway through the year by metabarcoding, targeting the major capsid protein (MCP) and its correlation to protist diversity and dynamics. Metabarcoding results demonstrated a high diversity of algal viruses compared to previous metabarcoding surveys in Norwegian coastal waters. We obtained 313 putative algal virus operational taxonomic units (vOTUs), all classified by phylogenetic analyses to either the Phycodnaviridae or Mimiviridae families, most of them in clades without any cultured or environmental reference sequences. The viral community showed a clear temporal variation, with some vOTUs persisting for several months. The results indicate co-occurrences between abundant viruses and potential hosts during long periods. This study gives new insights into the virus-algal host dynamics and provides a baseline for future studies of algal virus diversity and temporal dynamics.

Unmasking cellular response of a bloom-forming alga to viral infection by resolving expression profiles at a single-cell level.

Infection by large dsDNA viruses can lead to a profound alteration of host transcriptome and metabolome in order to provide essential building blocks to support the high metabolic demand for viral assembly and egress. Host response to viral infection can typically lead to diverse phenotypic outcome that include shift in host life cycle and activation of anti-viral defense response. Nevertheless, there is a major bottleneck to discern between viral hijacking strategies and host defense responses when averaging bulk population response. Here we study the interaction between Emiliania huxleyi, a bloom-forming alga, and its specific virus (EhV), an ecologically important host-virus model system in the ocean. We quantified host and virus gene expression on a single-cell resolution during the course of infection, using automatic microfluidic setup that captures individual algal cells and multiplex quantitate PCR. We revealed high heterogeneity in viral gene expression among individual cells. Simultaneous measurements of expression profiles of host and virus genes at a single-cell level allowed mapping of infected cells into newly defined infection states and allowed detection specific host response in a subpopulation of infected cell which otherwise masked by the majority of the infected population. Intriguingly, resistant cells emerged during viral infection, showed unique expression profiles of metabolic genes which can provide the basis for discerning between viral resistant and susceptible cells within heterogeneous populations in the marine environment. We propose that resolving host-virus arms race at a single-cell level will provide important mechanistic insights into viral life cycles and will uncover host defense strategies.

The mutual interplay between calcification and coccolithovirus infection.

Two prominent characteristics of marine coccolithophores are their secretion of coccoliths and their susceptibility to infection by coccolithoviruses (EhVs), both of which display variation among cells in culture and in natural populations. We examined the impact of calcification on infection by challenging a variety of Emiliania huxleyi strains at different calcification states with EhVs of different virulence. Reduced cellular calcification was associated with increased infection and EhV production, even though calcified cells and associated coccoliths had significantly higher adsorption coefficients than non-calcified (naked) cells. Sialic acid glycosphingolipids, molecules thought to mediate EhV infection, were generally more abundant in calcified cells and enriched in purified, sorted coccoliths, suggesting a biochemical link between calcification and adsorption rates. In turn, viable EhVs impacted cellular calcification absent of lysis by inducing dramatic shifts in optical side scatter signals and a massive release of detached coccoliths in a subpopulation of cells, which could be triggered by resuspension of healthy, calcified host cells in an EhV-free, 'induced media'. Our findings show that calcification is a key component of the E. huxleyi-EhV arms race and an aspect that is critical both to the modelling of these host-virus interactions in the ocean and interpreting their impact on the global carbon cycle.

Phaeoviral Infections Are Present in Macrocystis, Ecklonia and Undaria (Laminariales) and Are Influenced by Wave Exposure in Ectocarpales.

Two sister orders of the brown macroalgae (class Phaeophyceae), the morphologically complex Laminariales (commonly referred to as kelp) and the morphologically simple Ectocarpales are natural hosts for the dsDNA phaeoviruses (family Phycodnaviridae) that persist as proviruses in the genomes of their hosts. We have previously shown that the major capsid protein (MCP) and DNA polymerase concatenated gene phylogeny splits phaeoviruses into two subgroups, A and B (both infecting Ectocarpales), while MCP-based phylogeny suggests that the kelp phaeoviruses form a distinct third subgroup C. Here we used MCP to better understand the host range of phaeoviruses by screening a further 96 and 909 samples representing 11 and 3 species of kelp and Ectocarpales, respectively. Sporophyte kelp samples were collected from their various natural coastal habitats spanning five continents: Africa, Asia, Australia, Europe, and South America. Our phylogenetic analyses showed that while most of the kelp phaeoviruses, including one from Macrocystispyrifera, belonged to the previously designated subgroup C, new lineages of Phaeovirus in 3 kelp species, Ecklonia maxima, Ecklonia radiata, Undaria pinnatifida, grouped instead with subgroup A. In addition, we observed a prevalence of 26% and 63% in kelp and Ectocarpales, respectively. Although not common, multiple phaeoviral infections per individual were observed, with the Ectocarpales having both intra- and inter-subgroup phaeoviral infections. Only intra-subgroup phaeoviral infections were observed in kelp. Furthermore, prevalence of phaeoviral infections within the Ectocarpales is also linked to their exposure to waves. We conclude that phaeoviral infection is a widely occurring phenomenon in both lineages, and that phaeoviruses have diversified with their hosts at least since the divergence of the Laminariales and Ectocarpales.

Is the Virus Important? And Some Other Questions.

The motivation for focusing on a specific virus is often its importance in terms of impact on human interests. The chlorella viruses are a notable exception and 40 years of research has made them the undisputed model system for large icosahedral dsDNA viruses infecting eukaryotes. Their status has changed from inconspicuous and rather odd with no ecological relevance to being the Phycodnaviridae type strain possibly affecting humans and human cognitive functioning in ways that remain to be understood. The Van Etten legacy is the backbone for research on Phycodnaviridae. After highlighting some of the peculiarities of chlorella viruses, we point to some issues and questions related to the viruses we choose for our research, our prejudices, what we are still missing, and what we should be looking for.

Variation in the Genetic Repertoire of Viruses Infecting Micromonas pusilla Reflects Horizontal Gene Transfer and Links to Their Environmental Distribution.

Prasinophytes, a group of eukaryotic phytoplankton, has a global distribution and is infected by large double-stranded DNA viruses (prasinoviruses) in the family Phycodnaviridae. This study examines the genetic repertoire, phylogeny, and environmental distribution of phycodnaviruses infecting Micromonas pusilla, other prasinophytes and chlorophytes. Based on comparisons among the genomes of viruses infecting M. pusilla and other phycodnaviruses, as well as the genome from a host isolate of M. pusilla, viruses infecting M. pusilla (MpVs) share a limited set of core genes, but vary strongly in their flexible pan-genome that includes numerous metabolic genes, such as those associated with amino acid synthesis and sugar manipulation. Surprisingly, few of these presumably host-derived genes are shared with M. pusilla, but rather have their closest non-viral homologue in bacteria and other eukaryotes, indicating horizontal gene transfer. A comparative analysis of full-length DNA polymerase (DNApol) genes from prasinoviruses with their overall gene content, demonstrated that the phylogeny of DNApol gene fragments reflects the gene content of the viruses; hence, environmental DNApol gene sequences from prasinoviruses can be used to infer their overall genetic repertoire. Thus, the distribution of virus ecotypes across environmental samples based on DNApol sequences implies substantial underlying differences in gene content that reflect local environmental conditions. Moreover, the high diversity observed in the genetic repertoire of prasinoviruses has been driven by horizontal gene transfer throughout their evolutionary history, resulting in a broad suite of functional capabilities and a high diversity of prasinovirus ecotypes.

Change in Emiliania huxleyi Virus Assemblage Diversity but Not in Host Genetic Composition during an Ocean Acidification Mesocosm Experiment.

Effects of elevated pCO2 on Emiliania huxleyi genetic diversity and the viruses that infect E. huxleyi (EhVs) have been investigated in large volume enclosures in a Norwegian fjord. Triplicate enclosures were bubbled with air enriched with CO2 to 760 ppmv whilst the other three enclosures were bubbled with air at ambient pCO2; phytoplankton growth was initiated by the addition of nitrate and phosphate. E. huxleyi was the dominant coccolithophore in all enclosures, but no difference in genetic diversity, based on DGGE analysis using primers specific to the calcium binding protein gene (gpa) were detected in any of the treatments. Chlorophyll concentrations and primary production were lower in the three elevated pCO2 treatments than in the ambient treatments. However, although coccolithophores numbers were reduced in two of the high-pCO2 treatments; in the third, there was no suppression of coccolithophores numbers, which were very similar to the three ambient treatments. In contrast, there was considerable variation in genetic diversity in the EhVs, as determined by analysis of the major capsid protein (mcp) gene. EhV diversity was much lower in the high-pCO2 treatment enclosure that did not show inhibition of E. huxleyi growth. Since virus infection is generally implicated as a major factor in terminating phytoplankton blooms, it is suggested that no study of the effect of ocean acidification in phytoplankton can be complete if it does not include an assessment of viruses.

Characterization of a new chlorovirus type with permissive and non-permissive features on phylogenetically related algal strains.

A previous report indicated that prototype chlorovirus PBCV-1 replicated in two Chlorella variabilis algal strains, NC64A and Syngen 2-3, that are ex-endosymbionts isolated from the protozoan Paramecium bursaria. Surprisingly, plaque-forming viruses on Syngen 2-3 lawns were often higher than on NC64A lawns from indigenous water samples. These differences led to the discovery of viruses that exclusively replicate in Syngen 2-3 cells, named Only Syngen (OSy) viruses. OSy-NE5, the prototype virus for the proposed new species, had a linear dsDNA genome of 327kb with 44-nucleotide-long, incompletely base-paired, covalently closed hairpin ends. Each hairpin structure was followed by an identical 2612 base-paired inverted sequence after which the DNA sequence diverged. OSy-NE5 encoded 357 predicted CDSs and 13 tRNAs. Interestingly, OSy-NE5 attached to and initiated infection in NC64A cells but infectious progeny viruses were not produced; thus OSy-NE5 replication in NC64A is blocked at some later stage of replication.

Three-year survey of abundance, prevalence and genetic diversity of chlorovirus populations in a small urban lake.

Inland water environments cover about 2.5 percent of our planet and harbor huge numbers of known and still unknown microorganisms. In this report, we examined water samples for the abundance, prevalence, and genetic diversity of a group of infectious viruses (chloroviruses) that infect symbiotic chlorella-like green algae. Samples were collected on a weekly basis for a period of 24 to 36 months from a recreational freshwater lake in Lincoln, Nebraska, and assayed for infectious viruses by plaque assay. The numbers of infectious virus particles were both host- and site-dependent. The consistent fluctuations in numbers of viruses suggest their impact as key factors in shaping microbial community structures in the water surface. Even in low-viral-abundance months, infectious chlorovirus populations were maintained, suggesting either that the viruses are very stable or that there is ongoing viral production in natural hosts.

Seasonal determinations of algal virus decay rates reveal overwintering in a temperate freshwater pond.

To address questions about algal virus persistence (i.e., continued existence) in the environment, rates of decay of infectivity for two viruses that infect Chlorella-like algae, ATCV-1 and CVM-1, and a virus that infects the prymnesiophyte Chrysochromulina parva, CpV-BQ1, were estimated from in situ incubations in a temperate, seasonally frozen pond. A series of experiments were conducted to estimate rates of decay of infectivity in all four seasons with incubations lasting 21 days in spring, summer and autumn, and 126 days in winter. Decay rates observed across this study were relatively low compared with previous estimates obtained for other algal viruses, and ranged from 0.012 to 11% h(-1). Overall, the virus CpV-BQ1 decayed most rapidly whereas ATCV-1 decayed most slowly, but for all viruses the highest decay rates were observed during the summer and the lowest were observed during the winter. Furthermore, the winter incubations revealed the ability of each virus to overwinter under ice as ATCV-1, CVM-1 and CpV-BQ1 retained up to 48%, 19% and 9% of their infectivity after 126 days, respectively. The observed resilience of algal viruses in a seasonally frozen freshwater pond provides a mechanism that can support the maintenance of viral seed banks in nature. However, the high rates of decay observed in the summer demonstrate that virus survival and therefore environmental persistence can be subject to seasonal bottlenecks.

Diversity and dynamics of algal Megaviridae members during a harmful brown tide caused by the pelagophyte, Aureococcus anophagefferens.

Many giant dsDNA algal viruses share a common ancestor with Mimivirus--one of the largest viruses, in terms of genetic content. Together, these viruses form the proposed 'Megaviridae' clade of nucleocytoplasmic large DNA viruses. To gauge Megaviridae diversity, we designed degenerate primers targeting the major capsid protein genes of algae-infecting viruses within this group and probed the clade's diversity during the course of a brown tide bloom caused by the harmful pelagophyte,Aureococcus anophagefferens We amplified target sequences in water samples from two distinct locations (Weesuck Creek and Quantuck Bay, NY) covering 12 weeks concurrent with the proliferation and demise of a bloom. In total, 475 amplicons clustered into 145 operational taxonomic units (OTUs) at 97% identity. One OTU contained 19 sequences with ≥97% identity to AaV, a member of the Megaviridae clade that infects A. anophagefferens, suggesting AaV was present during the bloom. Unifrac analysis showed clear temporal patterns in algal Megaviridae dynamics, with a shift in the virus community structure that corresponded to the Aureococcus bloom decline in both locations. Our data provide insights regarding the environmental relevance of algal Megaviridae members and raise important questions regarding their phylodynamics across different environmental gradients.

First Insights into the Viral Communities of the Deep-sea Anoxic Brines of the Red Sea.

The deep-sea brines of the Red Sea include some of the most extreme and unique environments on Earth. They combine high salinities with increases in temperature, heavy metals, hydrostatic pressure, and anoxic conditions, creating unique settings for thriving populations of novel extremophiles. Despite a recent increase of studies focusing on these unusual biotopes, their viral communities remain unexplored. The current survey explores four metagenomic datasets obtained from different brine-seawater interface samples, focusing specifically on the diversity of their viral communities. Data analysis confirmed that the particle-attached viral communities present in the brine-seawater interfaces were diverse and generally dominated by Caudovirales, yet appearing distinct from sample to sample. With a level of caution, we report the unexpected finding of Phycodnaviridae, which infects algae and plants, and trace amounts of insect-infecting Iridoviridae. Results from Kebrit Deep revealed stratification in the viral communities present in the interface: the upper-interface was enriched with viruses associated with typical marine bacteria, while the lower-interface was enriched with haloviruses and halophages. These results provide first insights into the unexplored viral communities present in deep-sea brines of the Red Sea, representing one of the first steps for ongoing and future sampling efforts and studies.

Deep sequencing of amplified Prasinovirus and host green algal genes from an Indian Ocean transect reveals interacting trophic dependencies and new genotypes.

High-throughput sequencing of Prasinovirus DNA polymerase and host green algal (Mamiellophyceae) ribosomal RNA genes was used to analyse the diversity and distribution of these taxa over a ∼10 000 km latitudinal section of the Indian Ocean. New viral and host groups were identified among the different trophic conditions observed, and highlighted that although unknown prasinoviruses are diverse, the cosmopolitan algal genera Bathycoccus, Micromonas and Ostreococcus represent a large proportion of the host diversity. While Prasinovirus communities were correlated to both the geography and the environment, host communities were not, perhaps because the genetic marker used lacked sufficient resolution. Nevertheless, analysis of single environmental variables showed that eutrophic conditions strongly influence the distributions of both hosts and viruses. Moreover, these communities were not correlated, in their composition or specific richness. These observations could result from antagonistic dynamics, such as that illustrated in a prey-predator model, and/or because hosts might be under a complex set of selective pressures. Both of these reasons must be considered to interpret environmental surveys of viruses and hosts, because covariation does not always imply interaction.

Isolation and characterization of a virus infecting the freshwater algae Chrysochromulina parva.

Water samples from Lake Ontario, Canada were tested for lytic activity against the freshwater haptophyte algae Chrysochromulina parva. A filterable lytic agent was isolated and identified as a virus via transmission electron microscopy and molecular methods. The virus, CpV-BQ1, is icosahedral, ca. 145nm in diameter, assembled within the cytoplasm, and has a genome size of ca. 485kb. Sequences obtained through PCR-amplification of DNA polymerase (polB) genes clustered among sequences from the family Phycodnaviridae, whereas major capsid protein (MCP) sequences clustered among sequences from either the Phycodnaviridae or Mimiviridae. Based on quantitative molecular assays, C. parva׳s abundance in Lake Ontario was relatively stable, yet CpV-BQ1׳s abundance was variable suggesting complex virus-host dynamics. This study demonstrates that CpV-BQ1 is a member of the proposed order Megavirales with characteristics of both phycodnaviruses and mimiviruses indicating that, in addition to its complex ecological dynamics, it also has a complex evolutionary history.

Tsv-N1: A Novel DNA Algal Virus that Infects Tetraselmis striata.

Numbering in excess of 10 million per milliliter of water, it is now undisputed that aquatic viruses are one of the major factors shaping the ecology and evolution of Earth's microbial world. Nonetheless, environmental viral diversity and roles remain poorly understood. Here we report the first thorough characterization of a virus (designated TsV) that infects the coastal marine microalga Tetraselmis striata. Unlike previously known microalgae-infecting viruses, TsV is a small (60 nm) DNA virus, with a 31 kb genome. From a range of eight different strains belonging to the Chlamydomonadaceae family, TsV was only able to infect T. striata. Gene expression dynamics revealed an up-regulation of viral transcripts already 1 h post-infection (p.i.). First clear signs of infection were observed 24 h p.i., with the appearance of viral factories inside the nucleus. TsV assembly was exclusively nuclear. TsV-N1 genome revealed very different from previously known algae viruses (Phycodnaviridae). Putative function and/or homology could be resolved for only 9 of the 33 ORFs encoded. Among those was a surprising DNA polymerase type Delta (only found in Eukaryotes), and two genes with closest homology to genes from human parasites of the urogenital tract. These results support the idea that the diversity of microalgae viruses goes far beyond the Phycodnaviridae family and leave the door open for future studies on implications of microalgae viruses for human health.

Infection of phytoplankton by aerosolized marine viruses.

Marine viruses constitute a major ecological and evolutionary driving force in the marine ecosystems. However, their dispersal mechanisms remain underexplored. Here we follow the dynamics of Emiliania huxleyi viruses (EhV) that infect the ubiquitous, bloom-forming phytoplankton E. huxleyi and show that EhV are emitted to the atmosphere as primary marine aerosols. Using a laboratory-based setup, we showed that the dynamic of EhV aerial emission is strongly coupled to the host-virus dynamic in the culture media. In addition, we recovered EhV DNA from atmospheric samples collected over an E. huxleyi bloom in the North Atlantic, providing evidence for aerosolization of marine viruses in their natural environment. Decay rate analysis in the laboratory revealed that aerosolized viruses can remain infective under meteorological conditions prevailing during E. huxleyi blooms in the ocean, allowing potential dispersal and infectivity over hundreds of kilometers. Based on the combined laboratory and in situ findings, we propose that atmospheric transport of EhV is an effective transmission mechanism for spreading viral infection over large areas in the ocean. This transmission mechanism may also have an important ecological impact on the large-scale host-virus "arms race" during bloom succession and consequently the turnover of carbon in the ocean.