Solenicola setigera is the first characterized member of the abundant and cosmopolitan uncultured marine stramenopile group MAST‐3

Culture‐independent molecular methods based on the amplification, cloning and sequencing of small‐subunit (SSU) rRNA genes are a powerful tool to study the diversity of prokaryotic and eukaryotic microorganisms for which morphological features are not conspicuous. In recent years, molecular data from environmental surveys have revealed several clades of protists lacking cultured and/or described members. Among them are various clades of marine stramenopiles (heterokonts), which are thought to play an essential ecological role as grazers, being abundant and distributed in oceans worldwide. In this work, we show that Solenicola setigera, a distinctive widespread colonial marine protist, is a member of the environmental clade MArine STramenopile 3 (MAST‐3). Solenicola is generally considered as a parasite or an epiphyte of the diatom Leptocylindrus mediterraneus. So far, the ultrastructural, morphological and ecological data available were insufficient to elucidate its phylogenetic position, even at the division or class level. We determined SSU rRNA gene sequences of S. setigera specimens sampled from different locations and seasons in the type locality, the Gulf of Lions, France. They were closely related, though not identical, which, together with morphological differences under electron microscopy, suggest the occurrence of several species. Solenicola sequences were well nested within the MAST‐3 clade in phylogenetic trees. Since Solenicola is the first identified member of this abundant marine clade, we propose the name Solenicolida for the MAST‐3 phylogenetic group.     

New Species of Saprobic Labyrinthulea (=Labyrinthulomycota) and the Erection of a gen. nov. to Resolve Molecular Polyphyly within the Aplanochytrids

A culture of a unicellular heterotrophic eukaryote was established from pollen‐baited seawater acquired from the nearshore environment in Tromsø, Norway. Light microscopy revealed the production of ectoplasmic nets and reproduction by biflagellated zoospores, as well as binary division. After culturing and subsequent nucleotide extraction, database queries of the isolate's 18S small ribosomal subunit coding region identified closest molecular affinity to Aplanochytrium haliotidis, a pathogen of abalone. Testing of phylogenetic hypotheses consistently grouped our unknown isolate and A. haliotidis among the homoplasious thraustochytrids. Transmission electron microscopy revealed complex cell walls comprised of electron‐dense lamella that formed protuberances, some associated with bothrosomes. Co‐culturing experiments with the marine fungus Penicillium brevicompactum revealed prolonged interactions with hyphal strands. Based on the combined information acquired from electron microscopy, life history information, and phylogenetic testing, we describe our unknown isolate as a novel species. To resolve molecular polyphyly within the aplanochytrids, we erect a gen. nov. that circumscribes our novel isolate and the former A. haliotidis within the thraustochytrids.     

60 Identification of diatom‐lytic bacterium sk‐02 and its capability to degrade stephanodiscus hantzschii

In order to control harmful algal blooms, many biological approaches have been tried. Specially, there have recently been discussions concerning the roles of bacteria in algal bloom dynamics. Then, algicidal bacteria are expected as an agent considerate for harmful algal blooms control. Development of these organisms as biological control agents involves isolation from environmental samples. With the aim of develop eco‐technology controlling water blooms in fresh waters, we isolated the diatom‐lysing bacteria from the sediments of Lake Seokchon and Pal¡¯tang River‐Reservoir. A soft agar‐overlay technique was used to isolate the diatom lytic bacteria. The SK‐02 showed a diatom lytic activity against Stephanodiscus hantzschii. Taxonomic identification including 16S rDNA base sequencing, and phylogenetic analysis indicated that the isolate SK‐02 had a 99.20% homology in its 16S rDNA base sequence with Pseudomonas putida. The nature of these diatom‐lying components is still under investigation. These results suggest that the indigenous bacteria isolated from the sediments may have a potential in the application and development of eco‐technology controlling harmful water blooms in the fresh water environments.     

Morphological Identities of Two Different Marine Stramenopile Environmental Sequence Clades: Bicosoeca kenaiensis (Hilliard, 1971) and Cantina marsupialis (Larsen and Patterson, 1990) gen. nov., comb. nov.

Although environmental DNA surveys improve our understanding of biodiversity, interpretation of unidentified lineages is limited by the absence of associated morphological traits and living cultures. Unidentified lineages of marine stramenopiles are called “MAST clades”. Twenty‐five MAST clades have been recognized: MAST‐1 through MAST‐25; seven of these have been subsequently discarded because the sequences representing those clades were found to either (1) be chimeric or (2) affiliate within previously described taxonomic groups. Eighteen MAST clades remain without a cellular identity. Moreover, the discarded “MAST‐13” has been used in different studies to refer to two different environmental sequence clades. After establishing four cultures representing two different species of heterotrophic stramenopiles and then characterizing their morphology and molecular phylogenetic positions, we determined that the two different species represented the two different MAST‐13 clades: (1) a lorica‐bearing Bicosoeca kenaiensis and (2) a microaerophilic flagellate previously named “Cafeteria marsupialis”. Both species were previously described with only light microscopy; no cultures, ultrastructural data or DNA sequences were available from these species prior to this study. The molecular phylogenetic position of three different “C. marsupialis” isolates was not closely related to the type species of Cafeteria; therefore, we established a new genus for these isolates, Cantina gen. nov.     

EVOLUTIONARY ANALYSES OF THE NUCLEAR‐ENCODED PHOTOSYNTHETIC GENE psbO FROM TERTIARY PLASTID‐CONTAINING ALGAE IN DINOPHYTA1

Although the dinophytes generally possess red‐algal‐derived secondary plastids, tertiary plastids originating from haptophyte and diatom ancestors are recognized in some lineages within the Dinophyta. However, little is known about the nuclear‐encoded genes of plastid‐targeted proteins from the dinophytes with diatom‐derived tertiary plastids. We analyzed the sequences of the nuclear psbO gene encoding oxygen‐evolving enhancer protein from various algae with red‐algal‐derived secondary and tertiary plastids. Based on our sequencing of 10 new genes and phylogenetic analysis of PsbO amino acid sequences from a wide taxon sampling of red algae and organisms with red‐algal‐derived plastids, dinophytes form three separate lineages: one composed of peridinin‐containing species with secondary plastids, and the other two having haptophyte‐ or diatom‐derived tertiary plastids and forming a robust monophyletic group with haptophytes and diatoms, respectively. Comparison of the N‐terminal sequences of PsbO proteins suggests that psbO genes from a dinophyte with diatom‐derived tertiary plastids (Kryptoperidinium) encode proteins that are targeted to the diatom plastid from the endosymbiotic diatom nucleus as in the secondary phototrophs, whereas the fucoxanthin‐containing dinophytes (Karenia and Karlodinium) have evolved an additional system of psbO genes for targeting the PsbO proteins to their haptophyte‐derived tertiary plastids from the host dinophyte nuclei.     

Freshwater diatom biogeography in the Canadian Arctic Archipelago

Aim Document the biogeographical distributions of diatoms in the Canadian Arctic in relation to environmental factors. Location The Canadian Arctic Archipelago. Methods Diatoms were extracted from lake sediments and treated using standard methods. Rarefaction‐estimated species richness, diatom concentrations (valves cc^?1), and diatom abundance were mapped using a Geographic Information System. The physical and chemical parameters of the lakes were measured. Results A total of 326 taxa from 63 genera were found in 62 lakes of the study area. Up to 85 and as low as eight taxa were identified in any one lake, and rarefaction‐estimated species richness correlated with lake size. Nearby lakes could have greatly different diatom assemblages. Many taxa showed limited distributions. Response surfaces and ordination techniques indicate that a large number of taxa prefer colder regions of the Arctic while the abundance of others was influenced by lake water chemical parameters such as TKN and SiO₂. Main conclusions Although many taxa were common and found across the study area, diatom assemblages showed regional differences in the Arctic. Some taxa were not found in either the northern or southern parts of the Archipelago and others were restricted to particular regions. Newly delineated genera showed interpretable geographical patterns and could be related to environmental factors, suggesting that this more natural grouping may enhance our understanding of diatom ecology. Geographical, physical, and chemical factors are needed to explain diatom distributions in the Arctic.     

Determining diatom ecotones and their relationship to terrestrial ecoregion designations in the central Canadian Arctic Islands

Ecotones are key areas for the detection of global change because many are predicted to move with shifts in climate. Prince of Wales Island, in the Canadian Arctic Archipelago, spans the transition between mid‐ to high‐Arctic ecoregions. We analyzed limnological variables and recent diatom assemblages from its lakes and ponds to determine if assemblages reflected this ecotone. Limnological gradients were short, and water chemistry explained 20.0% of diatom variance in a redundancy analysis (RDA), driven primarily by dissolved organic carbon, Ca and SO₄. Most taxa were small, benthic forms; key taxa such as planktonic Cyclotella species were restricted to the warmer, southern portion of the study area, while benthic Staurosirella were associated with larger, ice‐dominated lakes. Nonetheless, there were no significant changes in diatom assemblages across the mid‐ to high‐Arctic ecoregion boundary. We combined our data set with one from nearby Cornwallis Island to expand the study area and lengthen its environmental gradients. Within this expanded data set, 40.6% of the diatom variance was explained by a combination of water chemistry and geographic variables, and significant relationships were revealed between diatom distributions and key limnological variables, including pH, specific conductivity, and chl‐a. Using principal coordinates analysis, we estimated community turnover with latitude and applied piecewise linear regression to determine diatom ecotone positions. A pronounced transition was present between Prince of Wales Island and the colder, more northerly Cornwallis Island. These data will be important in detecting any future northward ecotone movement in response to predicted Arctic climate warming in this highly sensitive region.     

Morphological and genetic diversity of Beaufort Sea diatoms with high contributions from the Chaetoceros neogracilis species complex

Seventy‐five diatom strains isolated from the Beaufort Sea (Canadian Arctic) in the summer of 2009 were characterized by light and electron microscopy (SEM and TEM), as well as 18S and 28S rRNA gene sequencing. These strains group into 20 genotypes and 17 morphotypes and are affiliated with the genera Arcocellulus, Attheya, Chaetoceros, Cylindrotheca, Eucampia, Nitzschia, Porosira, Pseudo‐nitzschia, Shionodiscus, Thalassiosira, and Synedropsis. Most of the species have a distribution confined to the northern/polar area. Chaetoceros neogracilis and Chaetoceros gelidus were the most represented taxa. Strains of C. neogracilis were morphologically similar and shared identical 18S rRNA gene sequences, but belonged to four distinct genetic clades based on 28S rRNA, ITS‐1 and ITS‐2 phylogenies. Secondary structure prediction revealed that these four clades differ in hemi‐compensatory base changes (HCBCs) in paired positions of the ITS‐2, suggesting their inability to interbreed. Reproductively isolated C. neogracilis genotypes can thus co‐occur in summer phytoplankton communities in the Beaufort Sea. C. neogracilis generally occurred as single cells but also formed short colonies. It is phylogenetically distinct from an Antarctic species, erroneously identified in some previous studies as C. neogracilis, but named here as Chaetoceros sp. This work provides taxonomically validated sequences for 20 Arctic diatom taxa, which will facilitate future metabarcoding studies on phytoplankton in this region.     

Evaluating diatom community composition in the absence of marked limnological gradients in the high Arctic: a surface sediment calibration set from Cornwallis Island (Nunavut,Canada)

Fossil diatom assemblages preserved within the sedimentary record in Arctic lakes provide the potential to reconstruct past changes in important limnological variables. During the summers of 1992 and 1993, we examined previously unstudied freshwater ecosystems on Cornwallis Island, Arctic Canada, with the specific objectives of (1) documenting the limnology and modern diatom assemblages from this region, and (2) determining which environmental variables most influence diatom species distributions. The Cornwallis Island study sites displayed the least amount of variance in measured water chemistry variables in comparison to nearly all of our labs’ previous freshwater surveys in the Arctic. The small limnological gradients precluded the development of a statistically robust diatom inference model, but perhaps more importantly, allowed us to explore variations in diatom composition in the absence of marked variations in water chemistry. Diatom species turnover was minimal, with the most common diatom taxa being Achnanthidium minutissima, Nitzschia perminuta, N. frustulum, with lesser percent abundances of Chaemaepinnularia soehrensis, Navicula chiarae, Psammothidium marginulata, and A. kryophila. A small number of study sites differed from the majority with respect to water chemistry (e.g., coastal sites with high specific conductivities) and habitat availability (e.g., ephemeral ponds with extensive moss habitats), and these sites had markedly different diatom assemblages. These data reinforce previous observations that water chemistry and other climate-related factors are the primary environmental controls influencing diatom distributions at high latitudes. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Developmental and Ultrastructural Characterization and Phylogenetic Analysis of Trypanosoma herthameyeri n. sp. of Brazilian Leptodactilydae Frogs

We described the phylogenetic affiliation, development in cultures and ultrastructural features of a trypanosome of Leptodacylus chaquensis from the Pantanal biome of Brazil. In the inferred phylogeny, this trypanosome nested into the Anura clade of the basal Aquatic clade of Trypanosoma, but was separate from all known species within this clade. This finding enabled us to describe it as Trypanosoma herthameyeri n. sp., which also infects other Leptodacylus species from the Pantanal and Caatinga biomes. Trypanosoma herthameyeri multiplies as small rounded forms clumped together and evolving into multiple‐fission forms and rosettes of epimastigotes released as long forms with long flagella; scarce trypomastigotes and glove‐like forms are common in stationary‐phase cultures. For the first time, a trypanosome from an amphibian was observed by field emission scanning electron microscopy, revealing a cytostome opening, well‐developed flagellar lamella, and many grooves in pumpkin‐like forms. Transmission electron microscopy showed highly developed Golgi complexes, relaxed catenation of KDNA, and a rich set of spongiome tubules in a regular parallel arrangement to the flagellar pocket as confirmed by electron tomography. Considering the basal position in the phylogenetic tree, developmental and ultrastructural data of T. herthameyeri are valuable for evolutionary studies of trypanosome architecture and cell biology.     

Labyrinthula diatomea n. sp.—A Labyrinthulid Associated with Marine Diatoms

Labyrinthulomycetes are mostly fungus‐like heterotrophic protists that absorb nutrients in an osmotrophic or phagotrophic manner. Members of order Labyrinthulida produce unique membrane‐bound ectoplasmic networks for movement and feeding. Among the various types of labyrinthulids’ food substrates, diatoms play an important role due to their ubiquitous distribution and abundant biomass. We isolated and cultivated new diatom consuming Labyrinthulida strains from shallow coastal marine sediments. We described Labyrinthula diatomea n. sp. that differs from all known labyrinthulids in both molecular and morphological features. We provided strain delimitation within the genus Labyrinthula based on ITS sequences via haplotype network construction and compared it with previous phylogenetic surveys.     

First report of the potentially toxic marine diatom Pseudo‐nitzschia simulans (Bacillariophyceae) from the East Australian Current

Certain species of the marine diatom genus Pseudo‐nitzschia are responsible for the production of the domoic acid (DA), a neurotoxin that can bioaccumulate in the food chain and cause amnesic shellfish poisoning (ASP) in animals and humans. This study extends our knowledge by reporting on the first observation of the potentially toxic species Pseudo‐nitzschia simulans from this region. One clonal strain of P. simulans was isolated from the East Australian Current and characterized using light and transmission electron microscopy, and phylogenetic analyses based on regions of the internal transcribed spacer (ITS) and the D1–D3 region of the large subunit (LSU) of the nuclear‐encoded ribosomal deoxyribonucleic acid (rDNA), as well as examined for DA production as measured by liquid chromatography–mass spectrometry. Although this strain was non‐toxic under the defined growth conditions, the results unambiguously confirmed that this isolate is the potentially toxic species P. simulans – the first report of this species from the Southern Hemisphere.     

Company matters: The presence of other genotypes alters traits and intraspecific selection in an Arctic diatom under climate change

Arctic phytoplankton and their response to future conditions shape one of the most rapidly changing ecosystems on the planet. We tested how much the phenotypic responses of strains from the same Arctic diatom population diverge and whether the physiology and intraspecific composition of multistrain populations differs from expectations based on single strain traits. To this end, we conducted incubation experiments with the diatom Thalassiosira hyalina under present‐day and future temperature and pCO₂ treatments. Six fresh isolates from the same Svalbard population were incubated as mono‐ and multistrain cultures. For the first time, we were able to closely follow intraspecific selection within an artificial population using microsatellites and allele‐specific quantitative PCR. Our results showed not only that there is substantial variation in how strains of the same species cope with the tested environments but also that changes in genotype composition, production rates, and cellular quotas in the multistrain cultures are not predictable from monoculture performance. Nevertheless, the physiological responses as well as strain composition of the artificial populations were highly reproducible within each environment. Interestingly, we only detected significant strain sorting in those populations exposed to the future treatment. This study illustrates that the genetic composition of populations can change on very short timescales through selection from the intraspecific standing stock, indicating the potential for rapid population level adaptation to climate change. We further show that individuals adjust their phenotype not only in response to their physicochemical but also to their biological surroundings. Such intraspecific interactions need to be understood in order to realistically predict ecosystem responses to global change.     

Comparison of Freshwater Diatom Assemblages from a High Arctic Oasis to Nearby Polar Desert Sites and Their Application to Environmental Inference Models

Arctic oases are regions of atypical warmth and relatively high biological production and diversity. They are small in area (<5 km²) and uncommon in occurrence, yet they are relatively well studied due to the abundance of plant and animal life contained within them. A notable exception is the lack of research on freshwater ecosystems within polar oases. Here, we aim to increase our understanding of freshwater diatom ecology in polar oases. Diatoms were identified and enumerated from modern sediments collected in 23 lakes and ponds contained within the Lake Hazen oasis on Ellesmere Island, and compared with diatom assemblages from 29 sites located outside of the oasis across the northern portion of the island. There were significant differences in water chemistry variables between oasis and northern sites, with oasis sites having higher conductivity and greater concentrations of nutrients and related variables such as dissolved organic carbon (DOC). Taxa across all sites were typical of those recorded in Arctic freshwaters, with species from the genera Achnanthes sensu lato, Fragilaria sensu lato, and Nitzschia dominating the assemblages. A correspondence analysis (CA) ordination showed that oasis sites generally plotted separately from the northern sites, although the sites also appear to plot separately based on whether they were lakes or ponds. Canonical correspondence analysis (CCA) identified specific conductivity, DOC, and SiO₂ as explaining significant (P < 0.05) and additional amounts of variation in the diatom data set. The most robust diatom‐based inference model was generated for DOC, which will provide useful reconstructions on long‐term changes in paleo‐optics of high Arctic lakes.     

Hemispheric‐scale patterns of climate‐related shifts in planktonic diatoms from North American and European lakes

A synthesis of over 200 diatom‐based paleolimnological records from nonacidified/nonenriched lakes reveals remarkably similar taxon‐specific shifts across the Northern Hemisphere since the 19th century. Our data indicate that these diatom shifts occurred in conjunction with changes in freshwater habitat structure and quality, which, in turn, we link to hemispheric warming trends. Significant increases in the relative abundances of planktonic Cyclotella taxa (P<0.01) were concurrent with sharp declines in both heavily silicified Aulacoseira taxa (P<0.01) and benthic Fragilaria taxa (P<0.01). We demonstrate that this trend is not limited to Arctic and alpine environments, but that lakes at temperate latitudes are now showing similar ecological changes. As expected, the onset of biological responses to warming occurred significantly earlier (P<0.05) in climatically sensitive Arctic regions (median age=ad 1870) compared with temperate regions (median age=ad 1970). In a detailed paleolimnological case study, we report strong relationships (P<0.005) between sedimentary diatom data from Whitefish Bay, Lake of the Woods (Ontario, Canada), and long‐term changes in air temperature and ice‐out records. Other potential environmental factors, such as atmospheric nitrogen deposition, could not explain our observations. These data provide clear evidence that unparalleled warming over the last few decades resulted in substantial increases in the length of the ice‐free period that, similar to 19th century changes in high‐latitude lakes, likely triggered a reorganization of diatom community composition. We show that many nonacidified, nutrient‐poor, freshwater ecosystems throughout the Northern Hemisphere have crossed important climatically induced ecological thresholds. These findings are worrisome, as the ecological changes that we report at both mid‐ and high‐latitude sites have occurred with increases in mean annual air temperature that are less than half of what is projected for these regions over the next half century.     

Lower marine fungi (labyrinthulomycetes) and the decay of mangrove leaf litter

This paper deals with the association of members of the Labyrinthulomycetes (Thraustochytriales and Labyrinthulales) with decaying or decayed leaves at an intertidal mangrove at Morib, Malaysia. Representatives of both orders of these obligately marine unicellular eukaryotes of unresolved taxonomic affinities (Chamberlain & Moss, 1998) were consistently isolated from leaves at all stages of decay from the recently fallen to those in an advanced stage of decay, but not from either green or senescent yellow leaves attached to trees. Baiting experiments using -irradiated leaf discs of Sonneratia and Rhizophora spp. immersed in the aquatic environment of the mangrove, revealed that leaf material was colonised by both labyrinthulids and thraustochytrids within 24 hours of immersion at the test site and these organisms were isolated from the leaf material throughout the 14 day study period. In vitro experiments using axenic cultures of three thraustochytrid genera inoculated onto sterile discs of Sonneratia leaves and incubated for 14 days caused loss of both biomass and structural integrity of the leaf material. Freeze fracture, followed by scanning electron microscopy of leaves inoculated with a thraustochytrid and a strain of Labyrinthula, revealed that penetration of the leaf occured after 4 days and that the thraustochytrid was associated with localised degradation of internal leaf tissues. Cellulase production by an isolate of Schizochytrium aggregatum was detected. The results of all the above investigations are discussed with reference to the role of members of the Labyrinthulomycetes in nutrient cycling in the mangrove.     

The first report of the potentially harmful diatom Pseudo‐nitzschia caciantha from Australian coastal waters

The diatom Pseudo‐nitzschia is a significant component of coastal waters worldwide and a producer of the potent neurotoxin, domoic acid. Sixteen species belonging to this genus have been reported from Australian waters, but the potentially toxic species P. caciantha has not been previously known from this region. Two clonal strains of P. caciantha were isolated from Coogee Beach, south‐east Australia, and the morphological, molecular and toxicological evidence for this species delineation were examined using light and transmission electron microscopy, phylogenetic analysis based on sequences of the second internal transcribed spacer and domoic acid production as measured by liquid chromatography–mass spectrometry. The results unambiguously confirmed that these isolates are the potentially toxic species P. caciantha , being only the second report of this species in the Southern Hemisphere. The potential for further hidden Pseudo‐nitzschia diversity in these waters is considerable.     

MORPHOLOGY AND MOLECULAR PHYLOGENY OF ANKISTRODINIUM GEN. NOV. (DINOPHYCEAE), A NEW GENUS OF MARINE SAND‐DWELLING DINOFLAGELLATES FORMERLY CLASSIFIED WITHIN AMPHIDINIUM1

The classical athecate dinoflagellate genera (Amphidinium, Gymnodinium, Gyrodinium) have long been recognized to be polyphyletic. Amphidinium sensu lato is the most diverse of all marine benthic dinoflagellate genera; however, following the redefinition of this genus ～100 species remain now of uncertain or unknown generic affiliation. In an effort to improve our taxonomic and phylogenetic understanding of one of these species, namely Amphidinium semilunatum, we re‐investigated organisms from several distant sites around the world using light and scanning electron microscopy and molecular phylogenetic methods. Our results enabled us to describe this species within a new heterotrophic genus, Ankistrodinium. Cells of A. semilunatum were strongly laterally flattened, rounded‐quadrangular to oval in lateral view, and possessed a small asymmetrical epicone. The sulcus was wide and characteristically deeply incised on the hypocone running around the antapex and reaching the dorsal side. The straight acrobase with hook‐shaped end started at the sulcal extension and continued onto the epicone. The molecular phylogenetic results clearly showed that A. semilunatum is a distinct taxon and is only distantly related to species within the genus Amphidinium sensu stricto. The nearest sister group to Ankistrodinium could not be reliably determined.