Metabarcoding reveals potentially mixotrophic flagellates and picophytoplankton as key groups of phytoplankton in the Elbe estuary.

In estuaries, phytoplankton are faced with strong environmental forcing (e.g. high turbidity, salinity gradients). Taxa that appear under such conditions may play a critical role in maintaining food webs and biological carbon pumping, but knowledge about estuarine biota remains limited. This is also the case in the Elbe estuary where the lower 70 km of the water body are largely unexplored. In the present study, we investigated the phytoplankton composition in the Elbe estuary via metabarcoding. Our aim was to identify key taxa in the unmonitored reaches of this ecosystem and compare our results from the monitored area with available microscopy data. Phytoplankton communities followed distinct seasonal and spatial patterns. Community composition was similar across methods. Contributions of key classes and genera were correlated to each other (p < 0.05) when obtained from reads and biovolume (R2 = 0.59 and 0.33, respectively). Centric diatoms (e.g. Stephanodiscus) were the dominant group - comprising on average 55 % of the reads and 66-69 % of the biovolume. However, results from metabarcoding imply that microscopy underestimates the prevalence of picophytoplankton and flagellates with a potential for mixotrophy (e.g. cryptophytes). This might be due to their small size and sensitivity to fixation agents. We argue that mixotrophic flagellates are ecologically relevant in the mid to lower estuary, where, e.g., high turbidity render living conditions rather unfavorable, and skills such as phagotrophy provide fundamental advantages. Nevertheless, further findings - e.g. important taxa missing from the metabarcoding dataset - emphasize potential limitations of this method and quantitative biases can result from varying numbers of gene copies in different taxa. Further research should address these methodological issues but also shed light on the causal relationship of taxa with the environmental conditions, also with respect to active mixotrophic behavior.

A three-genome ultraconserved element phylogeny of cryptophytes.

Cryptophytes are single celled protists found in all aquatic environments. They are composed of a heterotrophic genus, Goniomonas, and a largely autotrophic group comprising many genera. Cryptophytes evolved through secondary endosymbiosis between a host eukaryotic heterotroph and a symbiont red alga. This merger resulted in a four-genome system that includes the nuclear and mitochondrial genomes from the host and a second nuclear genome (nucleomorph) and plastid genome inherited from the symbiont. Here, we make use of different genomes (with potentially distinct evolutionary histories) to perform a phylogenomic study of the early history of cryptophytes. Using ultraconserved elements from the host nuclear genome and symbiont nucleomorph and plastid genomes, we produce a three-genome phylogeny of 91 strains of cryptophytes. Our phylogenetic analyses find that that there are three major cryptophyte clades: Clade 1 comprises Chroomonas and Hemiselmis species, Clade 2, a taxonomically rich clade, comprises at least twelve genera, and Clade 3, comprises the heterotrophic Goniomonas species. Each of these major clades include both freshwater and marine species, but subclades within these clades differ in degrees of niche conservatism. Finally, we discuss priorities for taxonomic revision to Cryptophyceae based on previous studies and in light of these phylogenomic analyses.

Phytoplankton dynamics in the Mar Menor, a Mediterranean coastal lagoon strongly impacted by eutrophication.

The Mar Menor hypersaline coastal lagoon has suffered serious degradation in the last three decades attributable to nutrient pollution. In 2015, the lagoon experienced an intensive bloom of cyanobacteria that triggered a drastic change of its ecosystem. Our analyses indicate that phytoplankton in 2016-2021 did not present a seasonal variability pattern; the community was mainly dominated by diatoms and punctually reached abundance peaks above 107 cell L-1 along with chlorophyll a concentrations exceeding 20 µg L-1. The predominant diatom genera during these blooms were different as well as the nutrient conditions under which they were produced. These high diatom abundances are unprecedented in the lagoon; in fact, our data indicate that the taxonomic composition, time variation patterns and cell abundance of phytoplankton in 2016-2021 differ notably in comparison to the data published before 2015. Consequently, our results support the finding that the trophic status of the lagoon has changed profoundly.

The closest lineage of Archaeplastida is revealed by phylogenomics analyses that include Microheliella maris.

By clarifying the phylogenetic positions of 'orphan' protists (unicellular micro-eukaryotes with no affinity to extant lineages), we may uncover the novel affiliation between two (or more) major lineages in eukaryotes. Microheliella maris was an orphan protist, which failed to be placed within the previously described lineages by pioneering phylogenetic analyses. In this study, we analysed a 319-gene alignment and demonstrated that M. maris represents a basal lineage of one of the major eukaryotic lineages, Cryptista. We here propose a new clade name 'Pancryptista' for Cryptista plus M. maris. The 319-gene analyses also indicated that M. maris is a key taxon to recover the monophyly of Archaeplastida and the sister relationship between Archaeplastida and Pancryptista, which is collectively called 'CAM clade' here. Significantly, Cryptophyceae tend to be attracted to Rhodophyta depending on the taxon sampling (ex., in the absence of M. maris and Rhodelphidia) and the particular phylogenetic 'signal' most likely hindered the stable recovery of the monophyly of Archaeplastida in previous studies.

Cascading effects in freshwater microbial food webs by predatory Cercozoa, Katablepharidacea and ciliates feeding on aplastidic bacterivorous cryptophytes.

Heterotrophic nanoflagellates (HNF) are considered as major planktonic bacterivores, however, larger HNF taxa can also be important predators of eukaryotes. To examine this trophic cascading, natural protistan communities from a freshwater reservoir were released from grazing pressure by zooplankton via filtration through 10- and 5-µm filters, yielding microbial food webs of different complexity. Protistan growth was stimulated by amendments of five Limnohabitans strains, thus yielding five prey-specific treatments distinctly modulating protistan communities in 10- versus 5-µm fractions. HNF dynamics was tracked by applying five eukaryotic fluorescence in situ hybridization probes covering 55-90% of total flagellates. During the first experimental part, mainly small bacterivorous Cryptophyceae prevailed, with significantly higher abundances in 5-µm treatments. Larger predatory flagellates affiliating with Katablepharidacea and one Cercozoan lineage (increasing to up to 28% of total HNF) proliferated towards the experimental endpoint, having obviously small phagocytized HNF in their food vacuoles. These predatory flagellates reached higher abundances in 10-µm treatments, where small ciliate predators and flagellate hunters also (Urotricha spp., Balanion planctonicum) dominated the ciliate assemblage. Overall, our study reports pronounced cascading effects from bacteria to bacterivorous HNF, predatory HNF and ciliates in highly treatment-specific fashions, defined by both prey-food characteristics and feeding modes of predominating protists.

The complete plastid genome of a marine microalgae Cryptophyceae sp. CCMP2293 (Cryptophyta).

In this study, we present the complete plastid genome of Cryptophyceae sp. CCMP2293. The circular genome is 139,208 bp in length and contains 142 protein-coding genes (PCGs), 30 transfer RNA (tRNA) genes, 6 ribosome RNA (rRNA) genes, and 1 transfer-messenger RNA (tmRNA) gene. The overall nucleotide composition is: 33.6% A, 32.5% T, 16.8% C, and 17.1% G with a total A + T content of 66.1%. The phylogenetic tree was constructed to explore the taxonomic status of Cryptophyceae sp. CCMP2293, which is closely related to G. theta and R. salina.

Revision of the Genus Chroomonas HANSGIRG: The Benefits of DNA-containing Specimens.

For years the genus Chroomonas was defined as being a cryptophyte with rectangular periplast plates, with a gullet and with biliprotein types PC 630 or 645. In phylogenetic trees the genus proved to be paraphyletic. Moreover, cells with hexagonal periplast plates were found in an SEM preparation from material of the type species C. nordstedtii. In this study, material of Hansgirg's C. nordstedtii was subjected to PCR and to sequencing of two short DNA tags. These tags allowed for an unambiguous identification of the real C. nordstedtii in the phylogeny of the blue-green cryptophytes. The genus Chroomonas corresponds to subclade 1, whereas subclades 3 and 4 do not belong to Chroomonas, if Hemiselmis is maintained. Additional examination by light and scanning electron microscopy and by spectrophotometry demonstrate that subclade 1 comprises only cells with hexagonal periplast plates and PC 630, whereas rectangular periplast plates are found only in subclades 3 and 4. Consequently the genus Chroomonas and its type species, C. nordstedtii, are revised and two novel species, C. debatzensis and C. gentoftensis sp. nov., are described.

Phenology of cryptomonads and the CRY1 lineage in a coastal brackish lagoon (Vistula Lagoon, Baltic Sea).

Cryptomonadales have acquired their plastids by secondary endosymbiosis. A novel clade-CRY1-has been discovered at the base of the Cryptomonadales tree, but it remains unknown whether it contains plastids. Cryptomonadales are also an important component of phytoplankton assemblages. However, they cannot be readily identified in fixed samples, and knowledge on dynamics and distribution of specific taxa is scarce. We investigated the phenology of the CRY1 lineage, three cryptomonadales clades and a species Proteomonas sulcata in a brackish lagoon of the Baltic Sea (salinity 0.3-3.9) using fluorescence in situ hybridization. A newly design probe revealed that specimens of the CRY1 lineage were aplastidic. This adds evidence against the chromalveolate hypothesis, and suggests that the evolution of cryptomonadales' plastids might have been shorter than is currently assumed. The CRY1 lineage was the most abundant cryptomonad clade in the lagoon. All of the studied cryptomonads peaked in spring at the most freshwater station, except for P. sulcata that peaked in summer and autumn. Salinity and concentration of dissolved inorganic nitrogen most significantly affected their distribution and dynamics. Our findings contribute to the ecology and evolution of cryptomonads, and may advance understanding of evolutionary relationships within the eukaryotic tree of life.

A New Heterotrophic Cryptomonad: Hemiarma marina n. g., n. sp.

We report a new heterotrophic cryptomonad Hemiarma marina n. g., n. sp. that was collected from a seaweed sample from the Republic of Palau. In our molecular phylogenetic analyses using the small subunit ribosomal RNA gene, H. marina formed a clade with two marine environmental sequences, and the clade was placed as a sister lineage of the freshwater cryptomonad environmental clade CRY1. Alternatively, in the concatenated large and small subunit ribosomal RNA gene phylogeny, H. marina was placed as a sister lineage of Goniomonas. Light and electron microscopic observations showed that H. marina shares several ultrastructural features with cryptomonads, such as flattened mitochondrial cristae, a periplast cell covering, and ejectisomes that consist of two coiled ribbon structures. On the other hand, H. marina exhibited unique behaviors, such as attaching to substrates with its posterior flagellum and displaying a jumping motion. H. marina also had unique periplast arrangement and flagellar transitional region. On the basis of both molecular and morphological information, we concluded that H. marina should be treated as new genus and species of cryptomonads.

Diversity of nitrogen assimilation pathways among microbial photosynthetic eukaryotes.

In an effort to better understand the diversity of genes coding for nitrogen (N) uptake and assimilation pathways among microalgae, we analyzed the transcriptomes of five phylogenetically diverse single celled algae originally isolated from the same high arctic marine region. The five photosynthetic flagellates (a pelagophyte, dictyochophyte, chrysoph-yte, cryptophyte and haptophyte) were grown on standard media and media with only urea or nitrate as a nitrogen source; cells were harvested during late exponential growth. Based on homolog protein sequences, transcriptomes of each alga were interrogated to retrieve genes potentially associated with nitrogen uptake and utilization pathways. We further investigated the phylogeny of poorly characterized genes and gene families that were identified. While the phylogeny of the active urea transporter (DUR3) was taxonomically coherent, those for the urea transporter superfamily, putative nitrilases and amidases indicated complex evolutionary histories, and preliminary evidence for horizontal gene transfers. All five algae expressed genes for ammonium assimilation and all but the chrysophyte expressed genes involved in nitrate utilization and the urea cycle. Among the four algae with nitrate transporter transcripts, we detected lower expression levels in three of these (the dictyochophyte, pelagophyte, and cryptophyte) grown in the urea only medium compared with cultures from the nitrate only media. The diversity of N pathway genes in the five algae, and their ability to grow using urea as a nitrogen source, suggest that these flagellates are able to use a variety of organic nitrogen sources, which would be an advantage in an inorganic nitrogen - limited environment, such as the Arctic Ocean.

Rhinomonas nottbecki n. sp. (cryptomonadales) and molecular phylogeny of the family Pyrenomonadaceae.

The cryptomonad Rhinomonas nottbecki n. sp., isolated from the Baltic Sea, is described from live and fixed cells studied by light, scanning, and transmission electron microscopy together with sequences of the partial nucleus- and nucleomorph-encoded 18S rRNA genes as well as the nucleus-encoded ITS1, 5.8S, ITS2, and the 5'-end of the 28S rRNA gene regions. The sequence analyses include comparison with 43 strains from the family Pyrenomonadaceae. Rhinomonas nottbecki cells are dorsoventrally flattened, obloid in shape; 10.0-17.2 µm long, 5.5-8.1 µm thick, and 4.4-8.8 µm wide. The inner periplast has roughly hexagonal plates. Rhinomonas nottbecki cells resemble those of Rhinomonas reticulata, but the nucleomorph 18S rRNA gene of R. nottbecki differs by 2% from that of R. reticulata, while the ITS region by 11%. The intraspecific variability in the ITS region of R. nottbecki is 5%. In addition, the predicted ITS2 secondary structures are different in R. nottbecki and R. reticulata. The family Pyrenomonadaceae includes three clades: Clade A, Clade B, and Clade C. All Rhinomonas sequences branched within the Clade C, while the genus Rhodomonas is paraphyletic. The analyses suggest that the genus Storeatula is an alternating morphotype of the genera Rhinomonas and Rhodomonas and that the family Pyrenomonadaceae includes some species that were described multiple times, as well as novel species.

Phytoplankton assemblages in lateral lagoons of a large tropical reservoir / Assembleias fitoplanctônicas em lagoas laterais de um reservatório tropical de grande porte

This study aimed to analyse the composition and ecological attributes of the phytoplankton assemblages in four lateral lagoons and in the main channel of Rosana Reservoir (Paranapanema River, SE Brazil). Fieldwork was carried out in September and November/2004 and January, March, May and August/2005. A total of 283 taxa was identified. Zygnemaphyta was the most specious group, followed by Chlorophyta and Bacillariophyta. Higher richness, abundance and biomass were observed in the lagoons when compared with the river-reservoir sampling point, especially during the rainy period. Cryptophyceae and Bacillariophyceae dominated numerically. Cryptomonas brasiliensis Castro, Bicudo and Bicudo was the main species of the phytoplankton in terms of abundance and frequency of occurrence. The dynamics of the most important taxa are discussed and the results showed that the phytoplankton assemblages are mainly influenced by meteorological factors and nutrient availability (the main driving forces). Correlation analyses indicated that the assemblage abundance was limited by nutrient (nitrogen and phosphorus). The phytoplankton abundance influenced positively the zooplankton abundance, what indicates the prevalence of bottom-up control routes in the lateral lagoons system. The results validate the hypotheses that lateral lagoons have a prominent ecological role on the phytoplankton diversity, as already previously demonstrated for fish and zooplankton. Therefore, the incorporation of the lateral lagoons in environmental programmes should be a target strategy for the conservation of the regional aquatic biota, minimising the negative impact of the dam.

O objetivo deste estudo foi analisar a composição e os atributos ecológicos das assembleias fitoplanctônicas de quatro lagoas marginais e canal principal do Reservatório de Rosana (SE, Brasil). Os trabalhos de campo foram realizados em setembro e novembro de 2004, e janeiro, março, maio e agosto de 2005. O número total de táxons identificados foi de 283. Zygnemaphyta foi o grupo com maior número de espécies, seguido por Chlorophyta e Bacillariophyta. Maior riqueza, abundância e biomassa foram observadas nas lagoas, quando comparadas ao canal do rio-reservatório, especialmente durante o período chuvoso. Cryptophyceae e Bacillariophyceae dominaram numericamente as assembleias. Cryptomonas brasiliensis Castro, Bicudo e Bicudo foi a principal espécie do fitoplâncton, em termos de abundância e frequência de ocorrência. Os resultados mostraram que as assembleias fitoplanctônicas são influenciadas principalmente pelos fatores meteorológicos e nutrientes (funções de força). Análises de correlação mostraram que a abundância do fitoplâncton foi limitada por nutrientes (nitrogênio e fósforo). Por sua vez, a abundância fitoplanctônica influenciou positivamente a abundância do zooplâncton, o que indica a predominância de mecanismos de controle ascendente no sistema de lagoas laterais. Os resultados validaram a hipótese, assim como já havia sido demonstrado para peixes e zooplâncton, de que as lagoas laterais têm um proeminente papel na diversidade do fitoplâncton. A incorporação das lagoas laterais em programas ambientais seria uma boa estratégia para a conservação da biota aquática regional, minimizando o impacto negativo do represamento.

URGORRI COMPLANATUS GEN. ET SP. NOV. (CRYPTOPHYCEAE), A RED-TIDE-FORMING SPECIES IN BRACKISH WATERS(1).

The morphology, ultrastructure, phylogeny, and ecology of a new red-tide-forming cryptomonad, Urgorri complanatus Laza-Martínez gen. et sp. nov., is described. U. complanatus has been collected in southwestern European estuaries, blooming in the inner reaches of several of them. The estuarine character of the species is also supported by its in vitro salinity preferences, showing a maximum growth rate at 10 psu. U. complanatus is a distinctive species and can be easily distinguished by LM from other known brackish and marine species. Cells are dorsoventrally flattened. The plastid has two anterior lobes. One pyrenoid is located in each of the lobes, and a third one on the posterior part. Thylakoids are arranged in pairs and do not penetrate pyrenoids. The plastid is reddish due to the presence of the phycoerythrin Cr-PE545. An orange discoidal eyespot lies beneath the nucleus, in the posterior ventral face of the plastid. A long furrow runs from the vestibulum, and a gullet is lacking. The periplast is composed of an inner sheet. The nuclear 18S rDNA based molecular analysis reveals U. complanatus is not related to any of the main cryptomonad lineages. Based on ultrastructural and pigment data, the most probable relatives are those merged under the family Geminigeraceae. Its lack of derived characters, together with the presence of characters proposed in previous studies to be primitive, suggests Urgorri could be considered representative of the cryptophycean ancestral character state.

Checklist das Cryptophyceae do estado de São Paulo, Brasil / Checklist of Cryptophyceae from São Paulo State, Brazil

Usando como base de informações a monografia sobre o grupo publicada pelo projeto Flora Ficológica do Estado de São Paulo (Programa BIOTA), o acervo depositado no Herbário Científico do Estado de São Paulo "Maria Eneyda P. Kauffmann Fidalgo" (SP), a Lista de Espécies da Flora do Brasil e artigos científicos, dissertações e teses (com descrições e ilustrações) temos: nove gêneros e 39 espécies de criptoficeas, sendo 16 espécies exclusivas para o Estado de São Paulo, todas registradas em ambientes de água doce. A carência de especialistas no Estado de São Paulo e no Brasil, além de problemas de estratégia amostral, necessidade de uso da microscopia eletrônica e ausência de estudos de biologia molecular, são fatores que devem ter subestimado o conhecimento taxonômico de Cryptophyceae no Estado.

Based on the information provided by the monograph on the group published by the project Phycology Flora of São Paulo (BIOTA), the collection deposited in the Scientific Herbarium "Mary P. Eneyda Kauffmann Fidalgo " of the State of São Paulo, the List of Endangered Species of the Flora of Brazil and scientific papers, dissertations and theses (with descriptions and illustrations), there are nine genera and 39 species of cryptophytes, 16 of which are unique species to the State of São Paulo, all recorded in freshwater environments. The lack of specialists in the State of São Paulo and Brazil as well as problems in the sampling strategy, the necessary use of electronic microscopy and the absence of molecular biology studies are factors that must have underestimated the taxonomic knowledge of Cryptophyceae in the state.

Phytoplankton assemblages in a reservoir cascade of a large tropical - subtropical river (SE, Brazil)

The phytoplankton assemblages from eight reservoirs of the Paranapanema River were studied during two consecutive years. Chlorophyceae and Bacillaryophyceae dominated in richness. The observed high number of taxa, 234, reflects the extensive sampling programme and evidences the necessity of considering the whole hydrograph basin to assess the biodiversity status of inland water ecosystems. The dams had a negative effect on phytoplankton richness, with higher number of taxa associate to riverine (non-regulated) stretches. The tributary rivers also exhibited high species richness, showing the importance of considering the lateral dimension, in addition to the longitudinal one, for aquatic biota inventories in large river basins/reservoirs. Richness and diversity were also positively influenced by the connectivity with lateral wetlands (macrophyte-dominated lakes) due to the periphyton influence. The phytoplankton abundance/biomass was not influenced by higher water retention time. Higher values occurred in the middle basin stretches (river-passage reservoirs) due to the increase in the trophic conditions. There was a positive correlation with phosphorus. Poorer light conditions in the cascade do not limit the phytoplankton biomass, with assemblages dominated by species tolerant to turbulent conditions and high mineral turbidity. Bacillariophyceae and Cryptophyceae dominated numerically. The first group (unicellular forms) was prominent in the large and oligotrophic upstream reservoirs. The second was highly abundant in the river-passage (low retention time), and more eutrophic, reservoirs. Cyanophyceae growth is probably controlled by advection processes (wash-out effect). The zooplankton does not control the phytoplankton biomass and the diversity of both groups is positively associated in the cascade. The structure of the phytoplankton assemblages showed to be a good indicator of the operationally distinct reservoirs of the Paranapanema cascade and also reflected the changes in the trophic conditions along the basin.

As assembleias fitoplanctônicas de oito reservatórios do rio Paranapanema foram estudadas durante dois anos consecutivos. Chlorophyceae e Bacillaryophyceae foram os grupos dominantes em riqueza. O elevado número de táxons observado, 234, reflete o extensivo programa de amostragem e evidencia a necessidade de se considerar a bacia hidrográfica como um todo em programas de avaliação da biodiversidade de ecossistemas aquáticos interiores. As barragens tiveram um efeito negativo sobre a riqueza do fitoplâncton, sendo o maior número de táxons associado aos trechos fluviais (não regulados). Os rios tributários também exibiram um elevado número de espécies, mostrando a importância de se considerar a dimensão lateral, além do gradiente longitudinal, para os inventários da biota aquática em reservatórios e bacias hidrográficas de grande porte. A riqueza e a diversidade também foram positivamente influenciadas pela conectividade com áreas laterais inundáveis - várzeas dominadas pela presença de macrófitas aquáticas, devido à contribuição do perifíton. A abundância e biomassa do fitoplâncton não foram influenciadas pelo tempo de retenção da água nos reservatórios. Valores mais elevados foram observados no médio Paranapanema (reservatórios fio d'água) devido ao aumento nas condições de trofia. Houve uma correlação positiva com a concentração de fósforo. Condições de menor transparência não limitaram a biomassa fitoplanctônica, sendo as assembleias dominadas por espécies tolerantes a condições de elevada turbulência e turbidez mineral. Bacillariophyceae e Cryptophyceae foram as algas numericamente dominantes. O primeiro grupo (formas unicelulares) predominou nos grandes reservatórios oligotróficos do alto Paranapanema. O segundo grupo foi muito abundante em reservatórios do tipo fio d'água (baixo tempo de retenção) e também mais eutróficos. Provavelmente o crescimento das Cyanophyceae é controlado por processos de transporte advectivo. O zooplâncton não exerce um controle sobre a biomassa fitoplanctônica e a diversidade de ambos os grupos esteve positivamente associada na cascata. A estrutura das assembleias fitoplanctônicas pode ser considerada um bom indicador das distintas condições de operação dos reservatórios em cascata do rio Paranapanema e também reflete as mudanças nas condições oligotróficas ao longo da bacia.