Class Ulvophyceae Research Articles

Complete mitochondrial genome and phylogenetic analysis of the marine microalga Symbiochlorum hainanensis (Ulvophyceae, Chlorophyta)

Symbiochlorum hainanensis Gong et al. (2018). is a unicellular green alga belonging to Ulvophyceae, Chlorophyta, and considered as an important species in coral-algae symbiont areas. In this study, we revealed firstly the mitochondrial genome sequence of the S. hainanensis. This mitochondrial genome was a circular DNA molecule of 59,508 bp, including 24 transfer RNA genes, 3 ribosomal RNA genes, and 31 protein-coding genes. The GC content of the genome was 35.4%. The phylogenetic tree suggested that S. hainandiae was a sister to the OUU clade within the class Ulvophyceae. The mitochondrial genome structure and gene content of S. hainanensis supported that S. hainanensis was a new unidentified green alga in Ulvophyceae.

Ulva: An emerging green seaweed model for systems biology.

Green seaweeds exhibit a wide range of morphologies and occupy various ecological niches, spanning from freshwater to marine and terrestrial habitats. These organisms, which predominantly belong to the class Ulvophyceae, showcase a remarkable instance of parallel evolution toward complex multicellularity and macroscopic thalli in the Viridiplantae lineage. Within the green seaweeds, several Ulva species ("sea lettuce") are model organisms for studying carbon assimilation, interactions with bacteria, life cycle progression, and morphogenesis. Ulva species are also notorious for their fast growth and capacity to dominate nutrient-rich, anthropogenically disturbed coastal ecosystems during "green tide" blooms. From an economic perspective, Ulva has garnered increasing attention as a promising feedstock for the production of food, feed, and biobased products, also as a means of removing excess nutrients from the environment. We propose that Ulva is poised to further develop as a model in green seaweed research. In this perspective, we focus explicitly on Ulva mutabilis/compressa as a model species and highlight the molecular data and tools that are currently available or in development. We discuss several areas that will benefit from future research or where exciting new developments have been reported in other Ulva species.

Characterization of the chloroplast genome of Symbiochlorum hainanensis (Ulvophyceae, Chlorophyta) and its phylogenetic analysis

Symbiochlorum hainandiae S.Q. Gong & Z.Y. Li, 2018 is a unicellular green alga belonging to Ulvophyceae, Chlorophyta, and plays important roles in coral reef ecosystem. In this study, high-throughput sequencing technology is used to sequence and assemble the chloroplast genome of S. hainandiae. The complete chloroplast genome of S. hainandiae was 158, 96 bp with the GC content of 32.86%. A total of 126 genes were identified, including 98 protein-coding genes, 26 tRNA, and 2 rRNA genes. The inverted repeat region was lost in the complete chloroplast genome of S. hainandiae. The phylogenetic analysis supports that S. hainandiae is a new sister lineage to the genus Ignatius within the class Ulvophyceae.

Ulosarcina terrestrica gen. nov., sp. nov., a New Ulvophycean Sarcinoid Alga from the Russian Far East

Many filamentous and sarcinoid terrestrial or freshwater green algae that were previously assigned to the Chlorophyceae are members of lineages belonging to the class Ulvophyceae. One of these lineages is the Planophila-clade (Ulotrichales). Some genera in this clade share similar morphology: cell packages forming branched pseudofilaments, uniseriate or sometimes biseriate filaments, often embedded in common mucilage. During a study on soil algal diversity in the temperate monsoon climate zone in Russia (Primorsky Territory, Vladivostok), we isolated a strain of sarcinoid green alga and examined it using an integrative approach. SSU and ITS rDNA sequence data, morphological characteristics, and life cycle features differentiated this strain from closely related genera of the order Ulotrichales and led us to describe it as Ulosarcina terrestrica gen. et sp. nov.

Tracking Iodine Decrease in Commercially Sold Caulerpa racemosa (Forsskål) J. Agardh (Chlorophyta, Ulvophyceae) during Storage

Iodine is known as an essential mineral for thyroid hormone production in humans. It is water- soluble and diffuses into the atmosphere, where it is thought to break ozone molecules. Caulerpa racemosa (Phylum Chlorophyta, Class Ulvophyceae, Order Bropsidales) is a common dietary seaweed believed to be a good source of iodine in the Philippines. As of this writing, there is no record of iodine concentration measurements in harvested C. racemosa from Philippine waters. There is an expected decrease in iodine concentrations from harvested C. racemosa through time. But how fast it decreases and how much of it remains in the seaweed if stored for days is also unknown. This study measured iodine concentration in C. racemosa samples from an area in the Philippines and calculated the daily changes in iodine level concentrations of harvested C. racemosa that were stored. Titration analyses were done every 24 h, with factors such as the algal age, temperature, light, among others, kept constant. Storage conditions were adapted from how local vendors store leftover seaweed merchandise. Results revealed a decrease in iodine levels over time (y = –20.438x + 228.99; R² = 0.8937). The initial concentration (at dry weight) of iodine measured was 196 ppm and dropped to 94.8 ppm on the seventh day. Iodine levels of freshly picked and stored C. racemosa were also compared with the recommended daily iodine intake for humans.

Metatranscriptomic Identification of Diverse and Divergent RNA Viruses in Green and Chlorarachniophyte Algae Cultures.

Our knowledge of the diversity and evolution of the virosphere will likely increase dramatically with the study of microbial eukaryotes, including the microalgae within which few RNA viruses have been documented. By combining total RNA sequencing with sequence and structural-based homology detection, we identified 18 novel RNA viruses in cultured samples from two major groups of microbial algae: the chlorophytes and the chlorarachniophytes. Most of the RNA viruses identified in the green algae class Ulvophyceae were related to the Tombusviridae and Amalgaviridae viral families commonly associated with land plants. This suggests that the evolutionary history of these viruses extends to divergence events between algae and land plants. Seven Ostreobium sp-associated viruses exhibited sequence similarity to the mitoviruses most commonly found in fungi, compatible with horizontal virus transfer between algae and fungi. We also document, for the first time, RNA viruses associated with chlorarachniophytes, including the first negative-sense (bunya-like) RNA virus in microalgae, as well as a distant homolog of the plant virus Virgaviridae, potentially signifying viral inheritance from the secondary chloroplast endosymbiosis that marked the origin of the chlorarachniophytes. More broadly, these data suggest that the scarcity of RNA viruses in algae results from limited investigation rather than their absence.

A siphonous macroalgal genome suggests convergent functions of homeobox genes in algae and land plants.

Genome evolution and development of unicellular, multinucleate macroalgae (siphonous algae) are poorly known, although various multicellular organisms have been studied extensively. To understand macroalgal developmental evolution, we assembled the ∼26 Mb genome of a siphonous green alga, Caulerpa lentillifera, with high contiguity, containing 9,311 protein-coding genes. Molecular phylogeny using 107 nuclear genes indicates that the diversification of the class Ulvophyceae, including C. lentillifera, occurred before the split of the Chlorophyceae and Trebouxiophyceae. Compared with other green algae, the TALE superclass of homeobox genes, which expanded in land plants, shows a series of lineage-specific duplications in this siphonous macroalga. Plant hormone signalling components were also expanded in a lineage-specific manner. Expanded transport regulators, which show spatially different expression, suggest that the structural patterning strategy of a multinucleate cell depends on diversification of nuclear pore proteins. These results not only imply functional convergence of duplicated genes among green plants, but also provide insight into evolutionary roots of green plants. Based on the present results, we propose cellular and molecular mechanisms involved in the structural differentiation in the siphonous alga.

Bacteria-induced morphogenesis of Ulva intestinalis and Ulva mutabilis (Chlorophyta): a contribution to the lottery theory

The green marine macroalgae of the class Ulvophyceae (Ulvophytes) are common algae distributed worldwide particularly in intertidal areas, which play a key role in aquatic ecosystems. They are potentially valuable resources for food, animal feed and fuel but can also cause massive nuisance blooms. Members of Ulvaceae, like many other seaweeds, harbour a rich diversity of epiphytic bacteria with functions related to host growth and morphological development. In the absence of appropriate bacterially derived signals, germ cells of the genus Ulva develop into ‘atypical’ colonies consisting of undifferentiated cells with abnormal cell walls. This paper examines the specificity of bacteria-induced morphogenesis in Ulva, by cross-testing bacteria isolated from several Ulva species on two Ulva species, the emerging model system Ulva mutabilis and the prominent biofouler species Ulva intestinalis. We show that pairs of bacterial strains isolated from species other than U. mutabilis and U. intestinalis can fully rescue axenic plantlets generated either from U. mutabilis or U. intestinalis gametes. This laboratory-based study demonstrates that different compositions of microbial communities with similar functional characteristics can enable complete algal morphogenesis and thus supports the ‘competitive lottery’ theory for how symbiotic bacteria drive algal development.

Reconstructing the Phylogeny of Capsosiphon fulvescens (Ulotrichales, Chlorophyta) from Korea Based on rbcL and 18S rDNA Sequences.

Capsosiphon fulvescens is a filamentous green algae in the class Ulvophyceae. It has been consumed as food with unique flavor and soft texture to treat stomach disorders and hangovers, and its economic value justifies studying its nutritional and potential therapeutic effects. In contrast to these applications, only a few taxonomic studies have been conducted on C. fulvescens. In particular, classification and phylogenetic relationships of the C. fulvescens below the order level are controversial. To determine its phylogenetic position in the class, we used rbcL and 18S rDNA sequences as molecular markers to construct phylogenetic trees. The amplified rbcL and 18S rDNA sequences from 4 C. fulvescens isolates (Jindo, Jangheung, Wando, and Koheung, Korea) were used for phylogenetic analysis by employing three different phylogenetic methods: neighbor joining (NJ), maximum parsimony (MP), and maximum likelihood (ML). The rbcL phylogenetic tree showed that all taxa in the order Ulvales were clustered as a monophyletic group and resolved the phylogenetic position of C. fulvescens in the order Ulotrichales. The significance of our study is that the 18S rDNA phylogenetic tree shows the detailed taxonomic position of C. fulvescens. In our result, C. fulvescens is inferred as a member of Ulotrichaceae, along with Urospora and Acrosiphonia.

Plant architecture without multicellularity: quandaries over patterning and the soma-germline divide in siphonous algae.

Multicellularity has independently evolved numerous times throughout the major lineages of life. Often, multicellularity can enable complex, macroscopic organismal architectures but it is not required for the elaboration of morphology. Several alternative cellular strategies have arisen as solutions permitting exquisite forms. The green algae class Ulvophyceae, for example, contains truly multicellular organisms, as well as macroscopic siphonous cells harboring one or multiple nuclei, and siphonocladous species, which are multinucleate and multicellular. These diverse cellular organizations raise a number of questions about the evolutionary and molecular mechanisms underlying complex organismal morphology in the green plants. Importantly, how does morphological patterning arise in giant coenocytes, and do nuclei, analogous to cells in multicellular organisms, take on distinct somatic and germline identities? Here, we comparatively explore examples of patterning and differentiation in diverse coenocytic and single-cell organisms and discuss possible mechanisms of development and nuclear differentiation in the siphonous algae.

De novo transcriptome analysis of an aerial microalga Trentepohlia jolithus: pathway description and gene discovery for carbon fixation and carotenoid biosynthesis.

BackgroundAlgae in the order Trentepohliales have a broad geographic distribution and are generally characterized by the presence of abundant β-carotene. The many monographs published to date have mainly focused on their morphology, taxonomy, phylogeny, distribution and reproduction; molecular studies of this order are still rare. High-throughput RNA sequencing (RNA-Seq) technology provides a powerful and efficient method for transcript analysis and gene discovery in Trentepohlia jolithus.Methods/Principal FindingsIllumina HiSeq 2000 sequencing generated 55,007,830 Illumina PE raw reads, which were assembled into 41,328 assembled unigenes. Based on NR annotation, 53.28% of the unigenes (22,018) could be assigned to gene ontology classes with 54 subcategories and 161,451 functional terms. A total of 26,217 (63.44%) assembled unigenes were mapped to 128 KEGG pathways. Furthermore, a set of 5,798 SSRs in 5,206 unigenes and 131,478 putative SNPs were identified. Moreover, the fact that all of the C4 photosynthesis genes exist in T. jolithus suggests a complex carbon acquisition and fixation system. Similarities and differences between T. jolithus and other algae in carotenoid biosynthesis are also described in depth.Conclusions/SignificanceThis is the first broad transcriptome survey for T. jolithus, increasing the amount of molecular data available for the class Ulvophyceae. As well as providing resources for functional genomics studies, the functional genes and putative pathways identified here will contribute to a better understanding of carbon fixation and fatty acid and carotenoid biosynthesis in T. jolithus.

A Consensus Secondary Structure of ITS2 in the Chlorophyta Identified by Phylogenetic Reconstruction

The definition of species plays a pivotal role in biology. It has been proposed that Compensatory Base Changes (CBCs) in the fast-evolving Internal Transcribed Spacer 2 (ITS2) correlate with speciation and thus can be used to distinguish species. The applicability of CBC - based species concepts using ITS2, however, rests on the homology of the investigated ITS2 positions. We studied the ITS2 molecule of 147 strains of Chlorophyceae (Chlorophyta, Viridiplantae) including 26 new sequences in the order Chaetophorales, and compared their secondary structures to ITS2 in the sister class Ulvophyceae, represented by the order Ulvales. Using a phylogenetic/comparative approach, it was possible to identify 1) the first consensus structure model of the ITS2 molecule that can be applied to two classes of green algae [Ulvophyceae (Ulvales), Chlorophyceae] and 2) landmarks (the spacer regions separating the ITS2 Helices) for more robust prediction of the secondary structures in green algae. Moreover, we found that CBCs in homologous positions in these 147 strains (representing 115 validly described species) are either completely absent or mostly associated with internal branches representing higher order taxonomic levels (genera, families, orders). As reported for the Ulvales, CBCs are not diagnostic at the species level in the dataset used.

Host specificity and coevolution of Flavobacteriaceae endosymbionts within the siphonous green seaweed Bryopsis

The siphonous green seaweed Bryopsis harbors complex intracellular bacterial communities. Previous studies demonstrated that certain species form close, obligate associations with Flavobacteriaceae. A predominant imprint of host evolutionary history on the presence of these bacteria suggests a highly specialized association. In this study we elaborate on previous results by expanding the taxon sampling and testing for host–symbiont coevolution Therefore, we optimized a PCR protocol to directly and specifically amplify Flavobacteriaceae endosymbiont 16S rRNA gene sequences, which allowed us to screen a large number of algal samples without the need for cultivation or surface sterilization. We analyzed 146 Bryopsis samples, and 92 additional samples belonging to the Bryopsidales and other orders within the class Ulvophyceae. Results indicate that the Flavobacteriaceae endosymbionts are restricted to Bryopsis, and only occur within specific, warm-temperate and tropical clades of the genus. Statistical analyses (AMOVA) demonstrate a significant non-random host–symbiont association. Comparison of bacterial 16S rRNA and Bryopsis rbcL phylogenies, however, reveal complex host–symbiont evolutionary associations, whereby closely related hosts predominantly harbor genetically similar endosymbionts. Bacterial genotypes are rarely confined to a single Bryopsis species and most Bryopsis species harbored several Flavobacteriaceae, obscuring a clear pattern of coevolution.

Remarks on Bačinella Radoičić, 1959 (type species B. irregularis) and its representatives

Bacinella irregularis was described by Radoicic (Bull Serv Geol Geophys Rep Serbie 17:87–92, 1959) as an alga incertae sedis. Based on material from the Lower Aptian of the western Maestrat Basin (Spain), the genus Bacinella with its various species is revised; an emended diagnosis is given as well as a detailed description. Bacinella is interpreted as an organism that excavated complex branching galleries into biogenic hard substrates, e.g., crusts of Lithocodiumaggregatum Elliott (such as in the holotype specimen of B. irregularis). Its morphology and boring pattern is comparable to that of modern filamentous-septate euendolithic green algae of the class Ulvophyceae. These forms, however, are microfilamentous (microborers) whereas Bacinella has a larger filament diameter. Nearly all of the “B. irregularis” occurrences reported in the literature do not belong to this taxon, but represent irregular vesicular crusts (“bacinellid” fabrics = B. irregularis auct. non) that display some resemblances with modern microbial mats.

Asexual life history by biflagellate zoids in Monostroma latissimum (Ulotrichales)

Monostroma latissimum (Kuetzing) Wittrock is a monostromatic green alga of commercial importance in Japan. Here we report the serendipitous discovery of asexually reproducing specimens collected from Usa, on the Pacific coast of Kochi Prefecture, south-western Japan. Zoids were found to be biflagellate and negatively phototactic. Germination of settled zoids was observed to follow erect-filamentous ontogeny similar to that of the previously reported sexual strain. Moreover, the newly discovered asexual strain had identical sequences of nuclear encoded ITS (Internal Transcribed Spacer) region to that of the sexual strain . On the basis of this finding, we postulate that the ITS sequences may have been maintained in these conspecific strains despite the evolution in sexuality. Relationships were investigated among M. latissimum and other monostromatic taxa within the class Ulvophyceae using ITS sequences in order to understand relative phylogenetic position of this species.

The Distribution of Elongation Factor‐1 Alpha (EF‐1α), Elongation Factor‐Like (EFL), and a Non‐Canonical Genetic Code in the Ulvophyceae: Discrete Genetic Characters Support a Consistent Phylogenetic Framework

The systematics of the green algal class Ulvophyceae have been difficult to resolve with ultrastructural and molecular phylogenetic analyses. Therefore, we investigated relationships among ulvophycean orders by determining the distribution of two discrete genetic characters previously identified only in the order Dasycladales. First, Acetabularia acetabulum uses the core translation GTPase Elongation Factor 1alpha (EF-1alpha) while most Chlorophyta instead possess the related GTPase Elongation Factor-Like (EFL). Second, the nuclear genomes of dasycladaleans A. acetabulum and Batophora oerstedii use a rare non-canonical genetic code in which the canonical termination codons TAA and TAG instead encode glutamine. Representatives of Ulvales and Ulotrichales were found to encode EFL, while Caulerpales, Dasycladales, Siphonocladales, and Ignatius tetrasporus were found to encode EF-1alpha, in congruence with the two major lineages previously proposed for the Ulvophyceae. The EF-1alpha of I. tetrasporus supports its relationship with Caulerpales/Dasycladales/Siphonocladales, in agreement with ultrastructural evidence, but contrary to certain small subunit rRNA analyses that place it with Ulvales/Ulotrichales. The same non-canonical genetic code previously described in A. acetabulum was observed in EF-1alpha sequences from Parvocaulis pusillus (Dasycladales), Chaetomorpha coliformis, and Cladophora cf. crinalis (Siphonocladales), whereas Caulerpales use the universal code. This supports a sister relationship between Siphonocladales and Dasycladales and further refines our understanding of ulvophycean phylogeny.

Phylogenetic relationships and species circumscription in Trentepohlia and Printzina (Trentepohliales, Chlorophyta)

Subaerial green microalgae represent a polyphyletic complex of organisms, whose genetic diversity is much higher than their simple morphologies suggest. The order Trentepohliales is the only species-rich group of subaerial algae belonging to the class Ulvophyceae and represents an ideal model taxon to investigate evolutionary patterns of these organisms. We studied phylogenetic relationships in two common genera of Trentepohliales ( Trentepohlia and Printzina) by separate and combined analyses of the rbcL and 18S rRNA genes. Trentepohlia and Printzina were not resolved as monophyletic groups. Three main clades were recovered in all analyses, but none corresponded to any trentepohlialean genus as defined based on morphological grounds. The rbcL and 18S rRNA datasets provided congruent phylogenetic signals and similar topologies were recovered in single-gene analyses. Analyses performed on the combined 2-gene dataset inferred generally higher nodal support. The results clarified several taxonomic problems and showed that the evolution of these algae has been characterized by considerable morphological convergence. Trentepohlia abietina and T. flava were shown to be separate species from T. aurea; Printzina lagenifera, T. arborum and T. umbrina were resolved as polyphyletic taxa, whose vegetative morphology appears to have evolved independently in separate lineages. Incongruence between phylogenetic relationships and traditional morphological classification was demonstrated, showing that the morphological characters commonly used in the taxonomy of the Trentepohliales are phylogenetically irrelevant.

Natural succession of macroalgal-dominated epibenthic assemblages at different water depths and after transplantation from deep to shallow water on Spitsbergen

In the current study, we investigated the primary succession of seaweeds over different time periods at different water depths. Furthermore, we followed the succession of field-grown benthic communities of different successional age, developing on ceramic tiles, prior to and after transplantation from 8 to 0.5 m water depth. The transplantation simulated changes associated with the break up of sea-ice cover, e.g. light regime or wave exposure. For this purpose, we transplanted 12 and 21-month old communities, grown at 8 m water depth, together with a set of sterile tiles, onto rafts, floating in 0.5 m water depth. Our results describe for the first time the succession of macroalgal communities in the Arctic and give important insights into the effect of disturbance of differently aged communities. The primary succession at 0.5 m water depth was mainly driven by Bacillariophyta and filamentous green algae like Urospora sp. and Ulothrix implexa. Twelve-month old communities at 8 m water depth are dominated by members of the Ectocarpales (Phaeophyceae), like Pylaiella littoralis, P. varia, and Ectocarpus siliculosus and the green alga U. implexa, whereas the 21-month old community showed a higher cover of the green algal class Ulvophyceae and sessile invertebrates. After transplantation to near surface conditions, species composition of the communities changed, but this effect was differently strong between communities of different age.

Morphology and phylogenetic position of Spongiochrysis hawaiiensis gen. et sp. nov., the first known terrestrial member of the order Cladophorales (Ulvophyceae, Chlorophyta)

Spongiochrysis hawaiiensis gen. et sp. nov. is described from material collected at two sites on O'ahu, Hawaiian Islands. This alga produced golden-yellow crusts on the bark of Casuarina trees and consisted of globular cells with an axial stellate chloroplast. The only form of reproduction was a specialized type of autosporulation in which a budding-like division of the mother cell produced daughter cells of different sizes. Phylogenetic analyses of the 18S rRNA gene showed that Spongiochrysis hawaiiensis is a member of the freshwater clade of the Cladophorales/Siphonocladales lineage in the green algal class Ulvophyceae. On the basis of its unicellular habit and terrestrial habitat, this species is well differentiated from all other members of the same lineage and can be considered as the first known successful step of a member of the order Cladophorales into terrestrial habitats. The implications of the description of this species with regard to both green algal evolution and conservation of little-known tropical habitats are discussed. The holotype specimen of Spongiochrysis hawaiiensis is GALW015489 and isotype specimens have also been deposited in the BISH and BM herbaria.

Protist Origin of the Ulvophyceae (Chlorophyta) Revealed by SSU rDNA Analyses of Marine Coccoid Green Algae

AbstractTraditionally, brackish and marine coccoid green algae are placed as isolated species of common trebouxiophycean or chlorophycean genera (e.g. Chlorella, Chlorococcum); exclusively the marine genera Chlorocystis and Halochlorococcum are characterized by quadriflagellated zoospores and a ‘Codiolum‐stage’ (characteristic for the Ulvophyceae) in their life histories. In Chlorocystis cohnii, sexual reproduction could only be induced by presence of the tube‐dwelling diatom Berkeleya rutilans. Phylogenetic analyses of the SSU rDNA sequences of 32, mostly marine, coccoid or sarcinoid strains confirmed the monophyly of the Ulvophyceae. The class contain 11 independent well‐supported lineages; of which eight contain coccoid species. (1) The basal branch of the ulvophytes is the Oltmannsiellopsis clade (in agreement with the order Oltmannsiellopsidales emend.) which contains the quadriflagellated monads (O. viridis and O. unicellularis) and the type species of the coccoid genus Halochlorococcum (H. marinum). (2) The other species of Halochlorococcum (except H. saccatum) formed together with Chlorocystis and Chlorella salina a significant clade corresponding to the order Chlorocystidales in emended form. (3) Gomontia polyrhiza is closely related to Urospora penicelliformis within the Acrosiphonia group (Acrosiphoniales). (4) The sarcinoid Trichosarcina polymorpha and T. mucosa, and the freshwater planctonic Helicodictyon planctonicum are member of the Ulothrix clade together with filamenteous taxa. (5) The type species of Pseudoneochloris (P. marina) form together with three unidentified marine coccoid strains an independent lineage within the Ulvophyceae (called Pseudoneochloris clade). (6–8) Ignatius tetrasporus (isolated from soil), the strain CCMP 250 originating from desert soil and the marine strain CCMP 1293 isolated from Palau Islands represent three single lineages within the ulvophytes. From our results, the class Ulvophyceae can be subdivided taxonomically into at least 11 orders (including the Ulvales, Cladophorales and Dasycladales) with the largest biodiversity at the coccoid level.†Present address: Department of Molecular and Cell Biology, Brown University, Providence, RI 02912, USA.