Draft genome sequence of the coccolithovirus Emiliania huxleyi virus 202

Emiliania huxleyi virus 202 (EhV-202) is a member of the Coccolithoviridae, a group of viruses that infect the marine coccolithophorid Emiliania huxleyi. EhV-202 has a 160- to 180-nm-diameter icosahedral structure and a genome of approximately 407 kbp, consisting of 485 coding sequences (CDSs). Here we describe the genomic features of EhV-202, together with a draft genome sequence and its annotation, highlighting the homology and heterogeneity of this genome in comparison with the EhV-86 reference genome.     

Draft genome sequence of four coccolithoviruses: Emiliania huxleyi virus EhV-88, EhV-201, EhV-207, and EhV-208

The Coccolithoviridae are a group of viruses which infect the marine coccolithophorid microalga Emiliania huxleyi. The Emiliania huxleyi viruses (known as EhVs) described herein have 160- to 180-nm diameter icosahedral structures, have genomes of approximately 400 kbp, and consist of more than 450 predicted coding sequences (CDSs). Here, we describe the genomic features of four newly sequenced coccolithoviruses (EhV-88, EhV-201, EhV-207, and EhV-208) together with their draft genome sequences and their annotations, highlighting the homology and heterogeneity of these genomes to the EhV-86 model reference genome.     

Draft genome sequence of the coccolithovirus EhV-84

The Coccolithoviridae is a recently discovered group of viruses that infect the marine coccolithophorid Emiliania huxleyi. Emiliania huxleyi virus 84 (EhV-84) has a 160 -180 nm diameter icosahedral structure and a genome of approximately 400 kbp. Here we describe the structural and genomic features of this virus, together with a near complete draft genome sequence (~99%) and its annotation. This is the fourth genome sequence of a member of the coccolithovirus family.     

Evolutionary history of the Coccolithoviridae

We recently determined the genome sequence of the Coccolithoviridae strain Emiliania huxleyi virus 86 (EhV-86), a giant double-stranded DNA (dsDNA) algal virus from the family Phycodnaviridae that infects the marine coccolithophorid E. huxleyi. Here, we determine the phylogenetic relationship between EhV-86 and other large dsDNA viruses. Twenty-five core genes common to nuclear-cytoplasmic large dsDNA virus genomes were identified in the EhV-86 genome; sequence from eight of these genes were used to create a phylogenetic tree in which EhV-86 was placed firmly with the two other members of the Phycodnaviridae. We have also identified a 100-kb region of the EhV-86 genome which appears to have transferred into this genome from an unknown source. Furthermore, the presence of six RNA polymerase subunits (unique among the Phycodnaviridae) suggests both a unique evolutionary history and a unique lifestyle for this intriguing virus.     

Viral trans-dominant manipulation of algal sphingolipids

Emiliania huxleyi is the host for the coccolithovirus (EhV), which is responsible for the demise of large oceanic blooms formed by this alga. The EhV-86 virus genome sequence has identified several genes apparently involved in sphingolipid metabolism. Recently, an unusual glucosylceramide from E. huxleyi infected with EhV-86 was isolated, implicating sphingolipids in the lysis of this alga. However, the EhV-86-encoded genes contain only a subset of the activities required to generate the novel sphingolipid, implying that its synthesis is the result of coordinated interactions between algal- and viral-encoded biosynthetic enzymes. Here, we discuss the likely role for EhV-86 open reading frames (ORFs) in the synthesis of novel sphingolipids and also consider the concept of the trans-dominant manipulation of lipid metabolism.     

Genomic presence of recombinant porcine endogenous retrovirus in transmitting miniature swine

Background

The Coccolithoviridae is a recently discovered family of viruses that infect the marine coccolithophorid Emiliania huxleyi. Following on from the sequencing of the type strain EhV-86, we have sequenced a second strain, EhV-163.

Results

We have sequenced approximately 80% of the EhV-163 genome, equating to more than 200 full length CDSs. Conserved and variable CDSs and a gene replacement have been identified in the EhV-86 and EhV-163 genomes.

Conclusion

The sequencing of EhV-163 has provided a wealth of information which will aid the re-annotating of the EhV-86 genome and identified a gene insertion in EhV-163.     

Proteomic analysis of the EhV-86 virion

Background

Emiliania huxleyi virus 86 (EhV-86) is the type species of the genus Coccolithovirus within the family Phycodnaviridae. The fully sequenced 407,339 bp genome is predicted to encode 473 protein coding sequences (CDSs) and is the largest Phycodnaviridae sequenced to date. The majority of EhV-86 CDSs exhibit no similarity to proteins in the public databases.

Results

Proteomic analysis by 1-DE and then LC-MS/MS determined that the virion of EhV-86 is composed of at least 28 proteins, 23 of which are predicted to be membrane proteins. Besides the major capsid protein, putative function can be assigned to 4 other components of the virion: two lectin proteins, a thioredoxin and a serine/threonine protein kinase.

Conclusion

This study represents the first steps toward the identification of the protein components that make up the EhV-86 virion. Aside from the major capsid protein, whose function in the virion is well known and defined, the nature of the other proteins suggest roles involved with viral budding, caspase activation, signalling, anti-oxidation, virus adsorption and host range determination.     

病毒介导的海洋球石藻(Coccolithophores)鞘脂类代谢调控

海洋球石藻(Coccolithophores)是一种全球广泛分布且具有重要生态功能的真核浮游植物,有些种类是大洋和近岸常见的赤潮种。自然海域中,病毒感染是导致球石藻死亡和赤潮消亡的一个关键因素。基于一株海洋球石藻Emiliania huxleyi及其特异性裂解病毒全基因组测序注释的结果,研究者们发现病毒可能通过基因横向转移从宿主基因组中获取了一系列与鞘脂类代谢相关的关键酶基因,进而在一定程度上掌控了宿主鞘脂类代谢,大量合成、积累病毒性鞘脂类物质,并最终诱导宿主细胞以凋亡的形式死亡。因此,病毒介导的宿主鞘脂类代谢在调节病毒与宿主间相互作用中具有重要意义。本文着重综述海洋球石藻病毒与宿主间的基因横向转移、病毒介导的宿主鞘脂类代谢特点及其生态学意义,以期深入了解海洋球石藻病毒与宿主间复杂的相互作用关系。     

142 Insight Into the Mitochondrial Genome of Emiliania Huxleyi (Haptophyta)

Emiliania huxleyi (Lohmann) Hay et Mohler 1967 (CCMP373) is the most abundant representative of the Haptophyta, and can be found in oceanic and neritic waters from subpolar to tropical latitudes. Blooms of this coccolithophorid may reach cell densities of 2.106 ml⁻¹ and emit vast amounts of DMS (dimethyl sulfide), with the potential to affect the global climate. We report here the DNA sequence of more than 21 kb of the mitochondrial genome of this species, out of an approximate total of 30 kb. Preliminary annotation of the genome using database searches identified at least 16 genes. The data were also compared to the unpublished mitochondrial genome of Pavlova lutheri , another member of the haptophytes, and some important differences were identified. Although the gene content of E. huxleyi mtDNA seems to closely resemble that of P. lutheri mtDNA, the gene order differs substantially. Access to the P. lutheri and E. huxleyi mitochondrial genomes will permit the comparative analysis of two deeply diverging members of an ancient and ecologically significant lineage. Among other potential applications, the data will help to clarify phylogeny within the haptophytes as well as to determine the phylogenetic position of this division in relation to other groups of algae, such as heterokonts, dinoflagellates and cryptophytes.     

Dynamics and genotypic composition of Emiliania huxleyi and their co-occurring viruses during a coccolithophore bloom in the North Sea

We studied the temporal succession of vertical profiles of Emiliania huxleyi and their specific viruses (EhVs) during the progression of a natural phytoplankton bloom in the North Sea in June 1999. Genotypic richness was assessed by exploiting the variations in a gene encoding a protein with calcium-binding motifs (GPA) for E.?huxleyi and in the viral major capsid protein gene for EhVs. Using denaturing gradient gel electrophoresis and sequencing analysis, we showed at least three different E.?huxleyi and EhV genotypic profiles during the period of study, revealing a complex, and changing assemblage at the molecular level. Our results also indicate that the dynamics of EhV genotypes reflect fluctuations in abundance of potential E.?huxleyi host cells. The presence and concentration of specific EhVs in the area prior to the bloom, or EhVs transported into the area by different water masses, are significant factors affecting the structure and intraspecific succession of E.?huxleyi during the phytoplankton bloom.     

Virus succession observed during an Emiliania huxleyi bloom

Denaturing gradient gel electrophoresis was used as a molecular tool to determine the diversity and to monitor population dynamics of viruses that infect the globally important coccolithophorid Emiliania huxleyi. We exploited variations in the major capsid protein gene from E. huxleyi-specific viruses to monitor their genetic diversity during an E. huxleyi bloom in a mesocosm experiment off western Norway. We reveal that, despite the presence of several virus genotypes at the start of an E. huxleyi bloom, only a few virus genotypes eventually go on to kill the bloom.     

Characterization of a Nitrogen-Regulated Protein Identified by Cell Surface Biotinylation of a Marine Phytoplankton

The biotinylating reagent succinimidyl 6-(biotinamido)hexanoate was used to label the cell surfaces of the cosmopolitan, marine, eukaryotic microorganism Emiliania huxleyi under different growth conditions. Proteins characteristic of different nutrient conditions could be identified. In particular, a nitrogen-regulated protein, nrp1, has an 82-kDa subunit that is present under nitrogen limitation and during growth on urea. It is absent under phosphate limitation or during exponential growth on nitrate or ammonia. nrp1 is the major membrane or wall protein in nitrogen-limited cells and is found in several strains of E. huxleyi. It may be a useful biomarker for examining the physiological state of E. huxleyi cells in their environment.     

Molecular dynamics of Emiliania huxleyi and cooccurring viruses during two separate mesocosm studies

In this study we used denaturing gradient gel electrophoresis, sequencing analysis, and analytical flow cytometry to monitor the dynamics and genetic richness of Emiliania huxleyi isolates and cooccurring viruses during two mesocosm experiments in a Norwegian fjord in 2000 and 2003. We exploited variations in a gene encoding a protein with calcium-binding motifs (GPA) and in the major capsid protein (MCP) gene to assess allelic and genotypic richness within E. huxleyi and E. huxleyi-specific viruses (EhVs), respectively. To our knowledge, this is the first report that shows the effectiveness of the GPA gene for analysis of natural communities of E. huxleyi. Our results revealed the existence of a genetically rich, yet stable E. huxleyi and EhV community in the fjordic environment. Incredibly, the same virus and host genotypes dominated in separate studies conducted 3 years apart. Both E. huxleyi-dominated blooms contained the same six E. huxleyi alleles. In addition, despite the presence of at least six and four EhV genotypes at the start of the blooms in 2000 and 2003, respectively, the same two virus genotypes dominated the naturally occurring infections during the exponential and termination phases of the blooms in both years.     

Development and validation of a molecular technique for the analysis of archived formalin-preserved phytoplankton samples permits retrospective assessment of Emiliania huxleyi communities

The coccolithophores, particularly the species Emiliania huxleyi (Lohmann) Hay & Mohler, account for the bulk of global calcium carbonate production and as such play a fundamental role in global CO2 cycling and the carbonate chemistry of the oceans. To evaluate the response of this functional group to the effects of climate change, we undertook a feasibility study to determine whether a retrospective approach could be used on archived coccolithophore datasets. We demonstrate for the first time a technique for the extraction of E. huxleyi nucleic acids from archived formalin-fixed samples of the long-term Continuous Plankton Recorder. Molecular analysis of a nine year old formalin-fixed sample reveals the presence of a diverse population of E. huxleyi genotypes within a developing coccolithophore bloom. In addition, E. huxleyi sequences were amplified from a number of formalin-fixed samples, the earliest of which was collected in August 1972. This molecular assay promises the possibility of studying global variations in the distribution and genetic make-up of E. huxleyi communities over extensive periods of time.     

Effects of HCO-3 on Surface Calcification and CO2 Fixation in Marine Emiliania huxleyi

Emiliania huxleyi is a ubiquitous species with the largest biomass in marine planktonics. When cells of E. huxleyi were grown in ESM with additional 20 mmol/L HC03- , coccolith scales were formed on cell surface and the weight ratio of calcium carbonate fixed in coecoliths to organic substance in cells was about 2.47: 1. It was only about 0.05: 1 in cells grown in ESM without HCO3- addition, where no coccoliths were observed under scanning electron microscope, and the content of lipid reached 18.1% of dry. cell weight. It was demonstrated that the HCO3- concentration was the key factor to control the calcification on cell surface. Therefore, in addition to the pathway of photosynthesis for CO2 fixation, calcification on cell surface forming coccoliths is an alternative pathway for fixing dissolved inorganic carbon in E. huxleyi. Moreover, being rich in lipids, E. huxleyi cells produced high content of hydrocarbons including extractable organic matter, saturates and aromatics under pyrolysis at 300℃. Among those, the yield of saturates from E. huxleyi reached as high as 2.8%, 6-15 times that from other algae. All these suggest that E. huxleyi is a good experimental system for studies on the optimization of environment through carbon cycle and renewable energy in algal biomass.